Document zzDrvar1jYpEm5Gd7QKnpjz9B

IN THE UNITED STATES DISTRICT COURT FOR THE DISTRICT OF NEVADA NEVADA POWER COMPANY, A NEVADA CORPORATION, PLAINTIFF, VS. MONSANTO COMPANY, A FOREIGN CORPORATION; GENERAL ELECTRIC COMPANY, A FOREIGN CORPORATION; WESTINGHOUSE ELECTRIC CORPORATION, A FOREIGN CORPORATION; AND DOES I XXV, INCLUSIVE, DEPENDENTS. CV-S-89-555-LDG-LRL EXHIBITS 1-18 TO THE DEPOSITION OF ROBERT EMMET KELLY, VOLUME I TAKEN ON FEBRUARY 15, 1994 M.D. MARTIN & ASSOCIATES CERTIFIED COURT REPORTERS 2200 MARKET STREET, SUITE 412 GALVESTON, TEXAS 77550 (409) 762-2222 * FAX (409) 762-8040 WATER PCB-SD0000030390 AN ACNEFORM DERMATERGOS1S JACK W. JONES, M.D. AN1I HERBERT S. ALDEN, M.D. ATLANTA, l!A. . .. ' It is only within comparatively recent times that the dermatologist has interested himself in dermatoses associated with or caused by the daily work. With the advent of a more clearly defined appraisal of the words eczema and dermatitis and the more general use of the patch test, der- matologists have necessarily inquired more and more deeply into the innumerable chemical contacts that occur in the dav bv dav lives of their patients. As well as being conversant with dermatology, the der- matologist must have a working knowledge of the manufacture and content of the many new and old com|>ounds that are handled and applied by large numbers of persons. Hence, more and more derma- tologists are becoming industrially minded, and it is dawning on them that the solution of some of their industrial difficulties may aid them in a clearer conception of the dermatoses that are encountered in their every-day practice. Dermatcrgoses of the acne form type occurring in persons working with chemicals have been repeatedly reported in Europe and discussed as industrial "chlor-acnc." ' This term "chlor-acne" was first used by Hcrxhcimcr2 in 1899 to describe an eruption couqiosed of comedones and small sebaceous pustules that occurred on the arms and faces of workers manufacturing chlorine gas electrolytically. using carbon electrodes. It was natural to assume that the chlorine was the causative agent. Uettman E had observed in March 1897 two patients whose skins were diffusely pigmented, dark, rough and dry. in whom were observed small tenacious comedones associated with numerous small follicular abscesses. Subsequently he rejnirted twenty-one additional cases with similar symptoms in workmen who were engaged in cleaning out an acid tower used in the manufacture of hydrochloric acid. At no time did these men come in contact with free chlorine, but numerous loose Read at the Fifty-Eighth Animal Meeting of the American Dermatological Association, Inc., White Sulphur Springs, \V. Ya.. May 2. 19.15. 1. White, Prosser: The Dermatcrgoses. cd. 4. New York, Paul P>. Hoclier. Inc., 1934. 2. Ilcrxhcinicr: Miinc.hcn. med. Wchnschr. 46:278, IS99. 3. Bettman: Deutsche med. Wchnschr. 27:437, 1901. t 1 < j : j : j j ; i j 5 j ! j j | | 1 j | ? WATER PCB-SD0000030391 JO N US-A1.1)UN--A CNUl:0 A'.l/ DUUM ATURGOSIS 1023 derivatives of coal lar were present. Herxheimcr later absolved the chlorine lmt considered the trouble to lie due to various chlorbenzenes. From time to time other re|Kirts of the occurrence of "chlor-acne" have been made in the European journals, each author continuing the error of Ilerxheimer in assuming that the cause of the condition was chlorine. Some of these reports have been indefinite as to the chemical conditions under which the men worked, except that they were exjiosed to chlorine gas. 1 lowever, some authors have suspected various chlor benzene derivatives, such as hexa-chlorlienzene, hexa-chlorethvlene, para-nitrobenzene, pcrchlornaphthalcnc ("jterna") and probably others.'1 Chlorinated naphthalene was first indicted by Wauer r' in 1918 as a cause of acncform eruptions of the skin, the condition being called Rcnuikraiiklieit. II is observations were confirmed by Koelsch " and recently by Teleky.7 The latter reported an exhaustive study of a large number of cases occurring in workers engaged in the manufacture of pcrchlornaphlhalcne, in which the workers were excised to fumes of the molten mass and also to the sublimated dusts. In approximately one-half of the workers exposed there developed comedones of a particularly heavy and tenacious type with resultant sebaceous abscesses, containing heavy yellow pus and serum. Considerable improvement resulted when the chlorine content of the mass was decreased from 30 per cent to 8 per cent, and Teleky believed that the disease was due directly to the halogen content of the substance. He mentioned the similarity of the disease to tccraaie (tar acne) but carried the antilogy no further. Individual predisposition appeared to alter receptivity. Some authors have leaned to the view that the acncform eruption is due to direct external contact with the chemicals (notably Bettman, ITallopcau, Jacquet, Fumouze, Teleky and Koelsch8). Others (Leh mann, Ilerxheimer, Jacobi, Both and Robert!l) considered it to be a -I. Occupation and Health: Encyclopedia of Hygiene, Pathology and Social Welfare, International Labour Office, boston, World Peace Foundation, 1925, brochures 34 and 285. 5. Wauer, cited in Occupation and Health: Encyclopedia of Hygiene, Pathol ogy and Social Welfare, International latlmur ("Hike, Itoston, World Peace Founda tion, 1925, brochure 34. 6. Koelsch, F., in Ulhnann, K.; Oppenheiin, M., and Rille, J. H.: Injuries to the Skin, Leipzig, Leopold Voss, 1920, vol. 2, p. 303. 7. Teleky: Klin. Wchnschr. 6:845 (April 30); 897 (May 7) 1927; 7:214 (Jan. 29) 1928. 8. hettman; ITallopcau; Jacquet: Fumouze; Teleky, and Koelsch, cited in Occupation and Health: Encyclopedia of Hygiene, Pathology and Social Wel fare. International I.alxiur Office, boston, World Peace Foundation, 1925, brochure 285. 9. Lehmann; Herxheimcr; Jacobi: Roth, and Robert, cited in Occupation and Health: Encyclopedia of Hygiene, Pathology and Social Welfare, Inter national Labour Office, boston, World Peace Foundation, 1925, brochure 285. WATER PCB-SD0000030392 1024 arch mis or nr.RMAroi.oaY and syi'iulolocy dermatitis due to the absorption of chemical comixumds by the lungs or gastro-iuleslinal tract, with elimination by .way of the sebaceous glands. The majority of writers on the subject, particularly Prosser White,1 have seemed to feel that chlorine as such has little to do with the formation of the comedones and cysts, and they have rcjwatedly referred to tar and products of the distillation of tar as the prime causative factors. Prosser White rebuked authors for using the term "chlor-acnc" at all and expressed the Itclief that the process is one of the manifold cutaneous reactions produced by tar and its derivatives. Recently we have had the opportunity to study, with the full cooi>eration of the manufacturers, an outbreak of acne form eruption occurring in a group of workers engaged in the manufacture of chlorinated di-phenyl. This study has brought out some |wints in the production of this unusual dermatosis that wc feel are important in solving its exact cause. The following case is typically illustrative of the disease as it occurred in the men working in this plant. RKl'ORT OF A CASK History.--O. D., a Negro aged 26, began work in the distillation of chlorinated di-phenyl in April 1930 and worked regularly until the latter part of the year 1933. Alxnit May of 1933, lie noticed the appearance of blackheads on hi? face, neck, arms and legs. These areas itched slightly. In a short time blackheads began to appear on the chest, hack and lower part of the abdomen, around the navel and on the scrotum and penis. Many of these blackheads swelled and became infected, discharging thick pus. The areas healed with difficulty and often left scars. The condition seemed to he progressive until Novemlter 19.13. When seen in December 1933 the patient complained of lassitude, loss of appetite and loss of libido and said that his cutaneous condition seemed to lie improving. Physical Examination.--On examination lie seemed in good general health. II is complaint of lassitude was not borne out by anything more than the usual tempera ment of the Negro toward work. On the forehead, extending within the hair line, and on the cheeks, chin, nose and neck were numerous small, very black, tenacious comedones, their distribution best described as being "|cppercd" within the skin. Many of the comedones surmounted firm shotlike cysts, which in some areas contained viscid yellow pus. The pustular elements were more noticeable on the neck. Similar shotlike comedones and cysts had appeared on the shoulders, midportion of the hack and chest, with an occasional large cyst. A peculiar peppering of the skin with tenacious carliou-cnlnrcd comedones was apparent around the umbilicus and lower portion of the abdomen. The scrotum and penis were involved in a similar process, the former being given more to the formation of cysts. The outer surfaces of the forearms and anterior thighs showed similar but fewer comedones. The whole eruption was acne form hut differed from acne particularly in the lack of a selwrrheic appearance of the skin and in the peculiarly deep black of the comedones as well as the general peppered distribution in areas not usually involved in acne vulgaris. A general physical examination had revealed nothing of importance. WATER PCB-SD0000030393 JONES-AUJEN--ACNECORM DERMATERGOSIS 1025 Treatment.--lie was instructed to scrub his skin thoroughly before and after his hours of work, to wear fresh clothing each day during work and to use a lotion made up as follows: zinc sulfate 4 Gin., potassium sulfurata 4 Gin. and distilled water to make 125 cc. On recommendation he. later reported for weekly doses of roentgen radiation according to the usual manner of treating acne vulgaris. Microscopic Examination.--A tiortiou of the skin of the chest was removed for microscopic study (fig. 1). The chief feature of the sections was noted in the hair follicles and sebaceous glands, in which there were cystic dilatation, Sumnufry of the Symptoms ami Treatment in Sixteen Cases of Acneform Eruption CllSt! Age Ill 22 2 28 ;ik 22 J\V 5F 28 OS :w 7Ji 20 bit 1!) i/D* 2<i 10U :;7 11G r<; 12P 20 1.211 :(7 tut 15 F 22 Hip 20 Knee WIlllC While While While While While Negro Negro Negro While While Willie While N`*gro While Negro Tyik* of Ski/f Kelmrrhcie; previous iieiie Average Seborrheic Average lry Average Seborrheic; previous acne Seborrheic Average Seborrheic Average Ury Seborrheic Average Average Seborrheic Selorrheic TyjH' of iCrii|tion THffnsc comedones; few cysts PHTiise comedones; few cysts Hi ITuse comedones; large cysts and pustules Pflfusc comedones; large cysts it ml uhsccsscs PHTuhc comedones; few cysts on face ami neck 1M IT use comedones; abscesses on neck; severe,eysls Frylhemnlous dif fuse eomedones; few snudl eysls Few scattered com edones; occasional cyst Diffuse comedones; cysts; smuil abscesses Scattered comedones; occasional abscess Sea l lere*l comedones; occasional ubsecss Few comedones; occasional cyst Occasional comedone Very few comedones Scattered coimalones PilTosc comedones; few cysts buck mnt fueii Time of Exposure 0 months 5 iiioiilhs SlKttiul Treatment Tiieiston; roentgeno therapy Noue Throughout lncishm; drainage 10 months None Throughout None 10 mouths S mouths Incision; drainage None 5 mouths None Throughout Roentgcnotheruiiy i) mouths None Throughout Roentgeno therapy 2 months None Throughout 12 months Throughout ? None None Roentgeno therapy Roentgeno therapy " Tills cuss Is riiiorlsil in detail. destruction of the hair, marked thinning and atrophy of the epithelium of the follicles and a heavy plug of keratinized material which partly filled the cystic cavity. In some areas there was a superficial plug at the surface opening; others showed the surface open and the plug deeply situated. There was no purulent exudate. There were a zone of moderately dense connective tissue surrounding the enlarged follicles, slight edema and infiltration by lymphocytes but no leukocytes. Slight edematous changes were noted in the occasional sebaceous glands' present, hut they were strikingly few. The sweat glands were normal. 1026 ARCHII'liS OI; DHKM ATOl.OGY ANH SYI'IIU.OLOGY Fig. 1.--Section of skin from the chest, showing histologic changes in the formation of an acncform eruption. WATER PCB-SD0000030395 JONliS-ALDGN--ACNLilHUMl DliRMATUUGOSIS 1027 Of the twenty-four men working in the manufacture of chlorinated di-phenyl within the period from the late summer of 1932 to Octol>er, 1933, twenty-three were retried to have had an acneform eruption on the face and hotly. Of the twenty-three, sixteen were examined. These men presented eruptions of acneform character similar in type and dis tribution to that in the case referred to hut varying in severity (table). In many patients numerous small sebaceous abscesses developed, par ticularly around the collora line, which exuded heavy, tenacious pus, Fig. 2.--Characteristic appearance and location of an acneform eruption. and tlie remaining ulcers were indolent, leaving in their wake much scarring. In two of the cases - particularly, many large abscesses developed on the neck and hack. Until very recent times di-phenyl and chlorinated di-phenyl were only laboratory curiosities, and their commercial manufacture was unknown. In the experimental stage of manufacture the apparatus was necessarily crude, and experimenters and workers were exposed for long periods to fumes and dusts containing much chlorine, both free and combined. In the early experiments, as well as in the early manufacture in large WATER PCB-SD0000030396 mix .ih'cmrns or imrM.rroi.ocY syi'iiii.ology quantities. the di-phenyl was made by healing benzene derived from crude coal tar. When heated in a suitable medium benzene (C,,H,,) becomes di-phenyl (C12H,,,). This substance is then chlorinated at various saturation j>oints, by exposing it under suitable conditions to free chlorine gas. Distillation of the resulting chlorinated di-phenyl10 results in a purified commercial product (fig. .V). The commercial l>enzcne always has a small quantity of impurities, hut some of the cruder commercial benzenes contain relatively large quantities of these impurities, such as xylene, toluene and paraffin. On the heating of the Fig. 3.--Chart illustrating the process of manufacture of chlorinated di-phcnvl. cruder benzenes, the toluenes and xylenes, when present in large quan tities, may produce the chemical substance styrene, which has the formula C,. Hr, CTI: CH,. Styrene when chlorinated (it being insepa rable from the di-phenyl) is an unstable compound dropping its chlorine atoms very readily. It is probable that other complex hydro carbons max' lye formed from these impurities, which on chlorination become quite as unstable as the styrene. One of these compounds may be chlorcthyl benzene (C I ln Cl I-CI-CII..,). 10. Chlorinated di-phcnvl consists of a mixture of di-phcnyl chlorinated at various saturation points: nona-chloro-di-phcnyl (Cm lICl.i -C CU) etc. > i 11 i i WATER PCB-SD0000030397 JOS'ES-.I LI)US'--.1CN ti!()AM I 1)HA'MATliJ<COS1S 1029 In the experimental stage and in the early manufacture of chlorinated di-phenyl, the men working were exjiosed for long periods to these chlorinated products. As the demand for the finished product increased, quantitative manufacture was s|>ecded up rapidly, and ojx'u stills and heating units were of necessity used until letter equipment could l>e designed and made. Hut since no physical trouble had developed in the workers previously there was no apparent hazard connected with manufacture. On or almut March 1933 the electrical properties of the chlorinated di-phenyl produced in the plant the workmen of which were examined fell below the sj>ccifieations, and the color of the product deepened. About one month later an acneform eruption was observed on the faces and arms of several of the workers, which became progres sively worse, gradually becoming apparent among all the workers engaged in the process at that time. Although one of the men had had a slight acneform eruption on his face in January, little thought was given to it until the eruption appeared in the others. In the summer of 1932, six or eight months previous to the general outbreak of the condition, owing to a difference in price, the crude benzene was pur chased from another source, and it was not until the purchase of this particular benzene was discontinued, in October 1933, that the finished product came up to the standard. During the jieriod from March 1933 until October 1933, the eruption continued among the men. and it was present as already described when the men were examined in December of the same year. The appearance of the eruption coincidentally with the production of a |Mior grade of chlorinated di-phenyl seemed to indicate a relation ship. On due chemical investigation styrene was assumed to be the offending substance causing the low grade of the finished product. It seems that it may be readily assumed that our problem as to the causative agents of the acneform eruption was connected with the chlorinated products of the impurities present in the crude l>enzene. However, with the exception of styrene, these products are not known. The problem hence becomes difficult of exact solution, and it is neces sary to resort to a reasonable deductive hypothesis. The exact conditions under which the men contracted their eruption cannot he satisfactorily duplicated experimentally since prolonged mild contact seems to be a deciding factor. Repeated patch tests made on numerous persons with the finished product, chlorinated di-phenyl, and the foreign substances styrene, styrene di-chloride and chlurethylbenzene, have not resulted in an acneform eruption except in one instance. Repealed patch tests made with chlorethylbenzene in the same area produced on the third attempt an erythematous follicular eruption, which persisted only a few days but which was liv no means typical. WATER PCB-SD0000030398 1030 ARCHIVES OF DERMATOLOGY AND SYPHILOLOGY We could not duplicate this in oilier instances. Since styrene, chlorinated styrene, chlorethylbenzene, xylene, toluene and their probable products are for the most part lipid solvents, they will produce death and denudation of the epithelium, and one always obtains a chemical burn in rcs|xmsc to a patch test made with the styrene and styrene di-chloridc. Small quantities mixed loosely with an inert powder or dissolved in ether produce a mild erythematous reaction hut no acucform eruption. Patch tests made with the finished product, chlori nated di-phenyl, at no time produced any erythema or evidence of irritation; hence the chlorinated di-phenyl can be absolutely absolved as an irritating agent. In looking over the consecutive history of the development of the eruption, we found it apparent that the men were in some degree exposed to the chlorinated impurities in the benzene for some time before the eruption actually appeared, and that to duplicate the conditions we should have to make related patch tests daily for long periods in the same person, a procedure which is not feasible. As a group, the workers in the dusts and vapors, which were pre sumably saturated with chlorinated hydrocarbons, were not cleanly, making little effort to bathe after work, and sweating and rubbing enhanced the liability to deixisition of the substances on the skin. Theoretically, the chlorinated hydrocarbons, presumably styrene di-chloridc and chlorethylbcnzcne, deposited on the skin gather around the hair follicles and sebaceous glands and arc gradually rubbed in. Being lipid solvents, they dissolve sebum and in the reaction, which is modified by the heat of the body and the water present, give olt hydro chloric acid, which causes death or irritation of the cells. This in turn causes an excess of cell growth, inflammation, sebaceous plugging and an acneform eruption (fig. 4). This process repeated over a long period may result in severe sebaceous infection, sebaceous abscess and scarring. One would naturally expect this process to occur in the seborrheic areas as well as where clothing might rub the particles into the skin. Also one would expect the process to enhance an already existing acne vulgaris. This occurred in cases 1 and 6 (table 1). the cases in which there was the worst involvement with sebaceous abscesses. In the beginning an attempt at prevention of the condition was made by being cs|>ecially careful that all men engaged in the manu facture of chlorinated di-phenyl should have a thorough bath after working hours and that they should wear freshly laundered clothing before starting work. They were also instructed to apply night and morning veterinary white lotion to the alTected parts. Some of the patients having the most numerous cysts and follicular infections were instructed to report for roentgen treatment administered in the manner usually employed in treating acne vulgaris. Following the WATER PCB-SD0000030399 JQNES-ALDEN--ACNEEORM DERMATIIRGOSIS 1031 change in the type of benzene used and the employment of enclosed distilling apparatus and ventilation fans, there was noted a gradual improvement in the acne form eruption. Those juitients receiving roent gen radiation improved much more rapidly, hut it cannot !>e said that this form of treatment gave unusual results. The final treatment, of course, was removal of the patient from the offending chlorinated hydrocarltons, and then cleanliness and relief from any infection. All the men showing evidence of cutaneous disease, so far as is known, are greatly improved, or the condition has entirely cleared except for an occasional comedo and a few scars. Fig. 4.--Process of manufacture of chlorinated di-phenyl containing impurities which cause an acneform eruption. When one recalls the cases reported by the European investigators, it is altogether plausible that the chemical process outlined here occurred in most instances. The slow liberation of chlorine in the presence of a lipid solvent (usually chlorinated hydrocarbons) was present in most cases. Tt is doubtful whether the disease is influenced by the quantity of chlorine present; it is apparently more due to the formation of unstable chlorinated hydrocarbons as well as to the liability of the workers to close contact without a thorough cleansing of the skin and a change front saturated clothing. It is also probable that the disease has little to do with tar itself but more with the products formed by the :i' I!.- i WATER 10.12 a item ms oi; dukmatolocy and svfiiii.olooy chlorination of tar. 1 lence, the disease cannot lie called "chlor-acne" or "tar acne." That it is not caused hy internal absorption--is evident when one considers the following facts: .First, acute intoxications by chlorine rarely occur, since suffocation and death hy pulmonary edema ensue even in the presence of relatively small quantities of chlorine gas in the air. Second, when ingested, chlorine and the chlorinated hydro carbons are so rapidly oxidized and chlorine so readily combines with sodium that opportunity for acne-like eruptions, such as occur in bromine intoxication, is not possible. In retrospect, we are intrigued by the hypothesis that these theoreti cal considerations may have some relation to acne vulgaris. The past quarter of a century has seen an enormous increase in the use of coal for heating homes as well as in industry. The smoke stacks of the cities have belched an ever increasing stream of coal smoke--smoke heavily laden with carbonaceous products containing numerous hydro carbons. it is well known that coal smoke contains hydrochloric acid. Is it not possible that at least some of the acne vulgaris may have a part of its origin in the constant contacts with the chlorinated hydro carbons of coal smoke? SUMMARY We have recorded herein observations concerning an industrial dermatosis of an acneform type. The type of eruption and the causative factors are analogous to those in the previously re|>orted cases siioken of in the European literature as instances of "chlor-acue," tar acne or PeniaJcrankheit. Although exact duplication of the conditions under which the workers acquired the dermatosis cannot be accomplished hy experimental means, reasonable deductions as to the cause of the difficulty can be made. The eruption occurred during the manufacture of chlorinated di-phenyl from benzene. The dermatosis occurred at a time when the benzene used contained excessive quantities of toluene, xylene and paraffin as impurities. The beating and chlorination of these impurities probably resulted in the production of styrene di-cldoride and chlorcthylbenzcne, which, we believe, on contact with the skin produced the acneform eruption by the slow lilieration of hydrochloric acid and not by internal absorption of any chemical. While many chlorinated hydrocarl ions, as brought out hy Prosser White,1 are lipid substances producing death or irritation of the cells by the liberation of chlorine, this is not true in all instances, since the finished pnxluct. chlorinated di-phenvl. did not, either experimentally or actually, produce any' cutaneous or sebaceous irritation. Irritation apparently de] tends on the ease with which chlorine atoms arc released from the hydro carbon base. We believe that these dermatoses should be lalicled acne form dcrmatcrgoscs resulting from certain unstable chlorinated WATER PCB-SD0000030401 JON US-A LI)E.\'--A CM lil;0 KM DliRMATERGOSIS 1033 hydrocarbons and should not he spoken of as "chlor-acne" or tar acne. The suggestion has been made that some of the acne vulgaris of the skin may he the result of the same type of irritation from soot or coal smoke. ABSTRACT 01' DISCUSSION Dit. Oi.ivkh S. Okmsiiv, Chicago: 1 should like to add an ex|>ericiice 1 had three months ago that is oi interest in this connection. I think that Dr. Jones' paper is extraordinarily interesting' and valuable, and the small group of cases 1 am going to report may add to his troubles rather than solve any of the problems. About three months ago in Michigan, I saw a patient, through the courtesy of Dr. Milton G, Butler, who presented an extraordinary acneforni eruption. This patient was 18 years old and had been observed by Dr. Butler. The eruption in these various cases consisted of acneforni pustules and comedones of variable size and depth, with scars, some of which were keloidal. The eruption occurred on the fitce and neck, particularly over the posterior portion, and extended well up into the scalp. The trunk, arms, thighs and legs were also involved. Two oi the patients presented on the hack of the neck deep |K-riiotlicular pustular lesions, with keloidal scars, such as are seen in Kaposi's dermatitis papillaris capillitii. Both deep and superficial atrophic scars, such as are seen in deep lesions leaving acne, were present in other portions of the neck, face and trunk. The comedones were prominent in all areas, including the legs. They were of variable size, and many were large and deep. The age of the affected |>alicnts varied from 18 to 50 years, the younger ones being more severely affected. The condition was apparently produced by contact with :m ingredient in it fungicide produced by a chemical company for destroying fungi in lumlter. The particular chemical in this fungicide that apparently was responsible for the eruption was sodium tetrachlor-ortho-phenylphenale, which is a yellow powder. Only those who came in contact with it bad the eruption. Those who handled the other ingredients of the fungicide and did not come in contact with this particular chemical had no difficulty. As there were apparently no chlorine, bromine or tar radicals in the formula, it is difficult to determine the active factor in the production of the lesions and also whether the eruption was caused hv local action by contact or through absorption and internal effects. The eruption was not concerned in any way with acne, and no patient in whom the eruption developed had had acne previously. Dr. Marion B. Sdi.ziikrc.kr, New York: I think that Dr. Jones has con tributed a study that will be very interesting when continued. I had an opportunity of seeing a group of patients with acne of external origin about a year and a half ago. The patients were all employed in a factory making radio condensers. Pitch tar and a type of wax (halowax, a chlorinated naphthalene) were used in these condensers. The acne which developed was like that described by Dr. Jones and also included comedones on the legs, as did that which Dr. Ormsby lias mentioned. I saw six patients from this one small factory. They were all in tbe age of puberty or the early twenties, and the group included persons of lioth sexes. I believe tbe most ini|>ortant point in this study is the possible connection between acneforni dermatoses distinctly due to external irritants and true acne vulgaris. It is interesting that many derivatives of tar are closely allied to the estrogenic hormone, that certain components of tar are Ixith estrogenic and acnegenic and that the estrogenic hormone must perhaps l>e incriminated in the produc tion of acne vulgaris. There can he no doubt that acne is often due to stimulation or I: WATER_PCB-SD0000030402 io.M archirns on hi-rmatoi.ocv asi> svnmr.oi.ocv irritation of the piloschaccous apparatus and that hormonal as well as external inllucnccs can produce this stimulation. In the patient suffering from acne, wlio has, of course, come in contact with the estrogenic hormone during inlra-uterinc life and then has had no further contact with this hormone (or the closely related testicular hormone) until lie or she begins to form his or her own hormones, one can readily conceive the |Kilhogcncsis of acne vulgaris as being based on an excessive stimulation of the hair follicles, a stimulation produced by the sudden new formation of the hormones at puberty. It is as though the follicles, after being sensitized to the hormones during fetal contact and then passing through a period of freedom from contact, react excessively to the new and massive contacts with the products of the gonads at puberty. I have discussed this subject in great detail elsewhere. Dr. H. G. Irvinf., Minneaimlis: 1 wish to comment on one point and to compliment Dr. Jones on an excellent piece of work and his presentation of it. There was a difference between his situation and that which Dr. Tunuacliff and I encountered (Arcii. Df.rmat. & Svi'ii. 33: .106 |Feh| 19.16). In his case the manufacturers called on the medical profession to help solve the problem. Our study was undertaken where lay |>coplc were in charge. It shows clearly the need of medical advice in the production of various chemical preparations. Dr. Jack W. Jones, Atlanta. Ga.: I am very grateful for the discussion. I did not have time to go into the various preventive measures attempted in con trolling the outbreak or the treatment. In most of the cases the condition has finally cleared up. Regarding Dr. Sulzberger's discussion, we studied this outbreak and the literature rather carefully for a year and a half or two years, and we are con vinced that the etiology in (his particular group can lie ascrilrcd to external irritation and not to any internal complications. Our idea is that the unstable chlorinated hydrocarbons give off chlorine very easily, whereas the chlorine that is fixed in the benzene ring is comparatively stable. When these unstable chlorinated hydrocarbons come into contact with the skin and enter the hair follicles, they are broken down slowly, giving off chlorine to form hydrochloric acid. We feel that this is the primary cause of the trouble. WATER PCB-SD0000030403 u SYMPOSIUM ON CJ.CRTAJN Cl J LOR J NATCH HYDROCARBONS Harvard School or Puituc Hkaltu, Boston*, Mass. ' ' . JuKK-30, iy.37 . ' Program ]. The problem of possible systemic cfleets from certain chlorinated hydro carbons. . Cecil K. Deism:?., M.mjl-ixini: Kicxji Waukcn a:; Ojuxvuai: A. Hcexrrr rj iltrxerj ,tLit! tf I'uUif Htjtli mj lA'^JriRK/ ef fliUrtir, ' }Urt*t4 UtCittl S;L*i.', L.<tL-m, Hot1. 2. Demonstration of laboratory n'.clhods for ventilating operations employing .chlorinated hydrocarbons. A discusssion of permissible concentrations of such compounds in the air. . . C.P. Vaclou, M_M.E. ' . 4 tUitect Pttftntr tf fuJuiUit! Wjjvu, limnJ Sdas-t tf I'vl-'.k Hullk '' . AM> |*ini i. r*fi.jir.^ Ch.H. . ' . .* * fttftutt tf Jljiitnt, JlenttJ .Wi.vi.-/ /'i.Yc 17tsLi ' ' 3. A description of the lesions found in the liver of animals subjected to .various chorinatcd hydrocarbons. ' | Gbanyxld: A. Bennett, M.D. Aittrik in /*<*&;?* Htnuti McIuaI SkLsA . .. ' . i* c. t (TIic recent! and third papers will appear rbxwhrre. Copies * of them wit! be sent to alt who at tended the symposium.} * . . 1 DEPOSITION i EXHIBIT |^ ............. ".......... " - v . .1 WATER PCB-SD0000030404 f tiwlfciirlinoi^h^'iiii>f>lTliTITiltl~%f THE JOURNAL OF . INDUSTRIAL HYGIENE AND TOXICOLOGY ,. ; ' Vomjue 19 SEPTEMBER, 1937 Number 7 - . THE PROBLEM OF POSSIBLE SYSTEMIC EFFECTS FROM . CERTAIN CHLORINATED HYDROCARBONS* Cecil IL Drinker,' Madeleine Field Warren and Granville A. .' Bennett Dtparlmtnt of Physiology, Harvard School of Public Htallh and Department of Pathology, -f 4 Harvard Medical School, dotton. Matt, < f*7'S :"\UIAV-~*.~. ' HE use of chlorinated naphtha rapidly and thoroughly as possible.! Tlenes and compounds of allied pharmacological possibilities is In brief these cases were as follows: . Patient 1. Male, age 21. The previous extremely wide, and with the steamdeydical history of this mar. was ia no way growth of the use of electricity is cer significant except for the fact that he had tain to expand much farther.- For an attach of jaundice about 6 weeks prior to years it has been known that many of these compounds cause a troublesome acne, and there is a large literature his fatal illness. Late in December, 193G, he became badly constipated and had much abdominal pain and distention. When ad mitted to the hospital he was slightly jaun upon this phase of the subject. Our diced and was evidently very ill. He was investigations have not been concerned somewhat anemic and his skin, particularly with chloracne but with the possibility of systemic effects following ingestion or inhalation of such products. In the upon the arms, face, chest and back, showed many pustules. He died after a brief period in the hospital, and at autopsy was found to spring of 193G, the Halowax Corpora tion, a division of the Bakelite Cor poration, called our attention to three fatal cases of jaundice in workmen using chlorinated naphthalenes and chlorinated diphenyl, and requested that the subject i>e investigated as * Received for publication Juno SO, 1037. t Tha Haiowax Company makes many products besides chlorinated naphthalenes, and it has come to our knowledge that all of these products arc indiscriminately called "halowaxca" by purchasers and users, and arc lumped together as possible causers of aene and even of systemic disease. Since "haiowax" is merely a trade designation, care should be taken to describe compounds by their chemical names and thus avoid condemnations which arc both troublesome end misleading. 2S3 jtyssjr. rtrw At WATER PCB-SD0000030405  ' 1-*S`i3t*r, 2S4 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [Sijii. i$S7 -f'TVciic- Lava a cirrhosis of the liver with acute yel low atrophy superimposed upon it. This mao. had been exposed to low cooceatratioos of vapors arising from a mixture of tetra and pentachlomaphthalenes, together with approximately 10 per. cent of a refined chlo who had been exposed to trichlornaphthalene. Of these 13 were found to have mild digestive complaints, anorexia, nausea and vertigo, but Courtois-Suffit remarks finally, "Ab rinated diphenyl. While both he and others engaged in the same work had chloracne, there were no other disturbances of health in fellow workmen, nor was there any pre cipitating cause for the acute yellow atro sorption is certainly possible and we have for proof of it some of the diges tive and general complaints which have been due to it. But they appear phy such as treatment with arsphenamine to be of little consequence considering or exposure to dangerous concentrations of carbon tetrachloride. " Patitni 3. This was a young man who died in February, 1926, after an acute illness characterized by jaundice. Ha had been the mildness of the digestive troubles and the absence of respiratory phe nomena." In Touraine's cases the exposure was exposed to fumes arising from a mixture of penta and hexachiornaphthalenea. There is no record of chloracne. The patient worked with a large number of other people of whom but one (Patient 3), a close friend, had significant illness. to a trichlornaphthalene, whereas the American cases of acute yellow atrophy were exposed to compounds of higher chlorination. Our own experiments indicate that tricnlomaphthalenes re Paiient 3. Another young man employed with Patient 2. Ha became jaundiced in March, 1SS6, and died after an illness of 2 weeks. A careful autopsy resulted in a diagnosis of acute yellow atrophy of the liver Here again no history could be ob quire enormous dosage, far beyond anything encountered in industry, in order to produce liver damage. Teleky (1227) reported a number of cases of chloracne in persons exposed to tained as to a precipitating cause, and there was no record of preceding attacks of jaundice. chlorinated naphthalenes with a chlo rine content ranging from 14 to 53 per cent. He found that the lower the In addition to these three very re chlorine content the lass the acne. cent fatalities, we have learned of four Mittelsiiidt (1935) examined a number other possible cases, none of them of cases of chloracne due to trichlor fatal. All of these have had jaundice naphthalene and reported a number of and the entire group consists of iso vague general complaints but nothing lated individuals who have been picked in the nature of serious disease. Re out of large groups having the same garding his animal experimentation, exposure. In but one instance, Pa Lehmann (1919) reported that ani tient 1, is there record of antecedent mals led chlorinated naphthalenes re disturbance of health, and the general fused to cat after a time and that, health of fellow workers has been good. whether poisoned by inhalation or by Such cases have not been reported feeding, at death showed "peculiar" in the medical literature and only occa lesions in the liver. Flinn and Jarvik sionally can one find reference to sys (1936) gave subcutaneous injections of temic effects of any sort. For exam enormous doses of chlorinated naph ple, Courtois-Sufiit (1934) reports on thalenes dissolved in paraffin oil to work done by Touraine and his asso rabbits. The compounds used were ciates (1934) who examined 00 workers as follows: * * *;?`v..v<..*<:;y-' . ;v*SS*L V ~- atl4sS&tH --'rSSS*! "-a*#?. I i ; :I ' 'A. V i'.. . 1' ! WATER_PCB-SD0000030406 rot. 1$, no. 7\ EFFECTS OF CHLORINATED HYDROCARBONS 285 1- A mixture of tri and tetrachlor- nificant in relation to the production naphthalcne. of acne. In the work of Flinn and 2. A mixture of tctra and penta- Jarvik (1936) tire compounds produc chlomaphthalenc. ing serious liver injury were the most 3. A mixture of penta and hexa- highly chlorinated of those tested, chlomaphthalene. - though the chlorine contents as given In addition, sublimates from (2) and by analysis vary surprisingly little. (3) were collected in oil and injected 3. There are no published figures subcutaneously. upon the amounts of various chlorin None of the animals receiving (1) or ated naphthalenes in the air which will the sublimate from (2) died, and even produce injury of any sort, and while after 2 months were quite normal when the work of Lehmann (1919) and of autopsied. The first death in the ani- Flitm and Jarvik (1936) point to the m:Us receiving (3) occurred on the 12th liver as a possible site of injury this day and the last died on the 20th day. indication rests upon such extreme Those receiving the sublimate from (3) dosage as to fail to apply directly to were even more severely affected. human exposure. Autopsy in these animals revealed striking changes in the liver, not, as EXPEHIXtENTAL WoUK described, entirely characteristic of In appraising the possible toxicity of acute yellow atrophy but sufficiently auy substance met in industry it is suggestive to cause the authors to con first necessary to determine the prin clude that "certain chlorinated naph cipal route of absorptiou. In the ease thalenes or impurities contained in of the compounds under consideration them are capable of producing yellow there can be no doubt that inhalatiou atrophy of the liver in the rabbit." is their chief means of entering the At the beginning of their paper, body. They arc used hot in a great Flitm and Jarvik (1930) mention the variety of operations and volatilize in fact that there liave been three cases varied degree. They are often applied of acute yellow atrophy of the liver in solution in such volatile solvents as in men working with chlorinated naph carbon tetrachloride and toluene. The thalenes but give no details in regard amounts reaching the air under such to them. These cases are undoubtedly circumstances are hardly detectable. the same as those described in the It will however lie shown that carbon beginning of this paper. tetrachloride adds to the toxicity of One may summarize the meagre the chlorinated naphthalenes and allied literature upon systemic effects from compounds, and if there is possibility these substances as follows: of inhaling these compounds in other 1. With the exception of the men parts of the factory then inhalation of tion of acute yellow atrophy by Flint! carbon tetrachloride adds a decided and Jarvik (1930) then; are no reports hazard. Under such circumstances or even suggestions of serious effects solvents such as toluene should be upon human beings. used. . 2. There is evidence (Teicky, 1927) Observation in a number of plants that the degree of chlorination is sig causes us to foci that oven though WATER PCB-SD0000030407 2S6 JOURNAL OF INDUSTRIAL HYCIEXEAND TOXICOLOGY [Sepl. 1337 workmen may be extremely dirty and careless, comparatively little of these waxes is eaten. They are tenacious substances, insoluble in water, and if they get on the hands they stick there and arc not transferred to the food. Skin absorption is the third possible means of entering the body. It may occur but at best must be slight when compared to the steady inhalation of finely divided or gaseous material in the air. tion. They, thrive upon a diet very similar to man, and in the case of these chlorinated compounds it is possible that diet may be very significant. Finally, their normal characteristics have been described so well as to make the detection of abnormalities both easy and certain. Method of eapasure.--The inhalation experiments were carried out in four large air-tight wooden boxes, each capable of holding ten rat cages, size Fig. 1. Front view and inflow end oi two 'ooxes with rat cages in place and doors open. Inhalation experiments are then the mast important sources of information, but to them we have added a certain number of observations upon ingestion anti subcutaneous injection of various compounds. Inhalation Experiments Annuals.--White rats have been employed throughout. They permit the use of a large mind ter of animals in a relatively small inhalation installa 22' x 22' x 14', in two tiers of five cages each. When the experiment was not in progress the doors were opened wide and the cages kept in place (figure 1). At oho center of one end of each box --the inflow end--air was introduced through a pipe. 7 inches in diameter (figure 2). Each box was equipped with an individual variable-speed elec tric blower which blow the air through several feet of 7-iuch pipe before enter- WATER PCB-SD0000030408 10, no. 7] EFFECTS OF CHLORINATED HYDROCARBON'S ins the end of the box. An orifice chlorinated diphenyl were placed in the meter was placed in the pipe line, and bottom of each flask, and melted in the the flow of air in cubic feet per minute electric heater.. Fresh samples were could be adjusted and read off directly used everj' other day, but it was often from a calibration curve. A vane found necessary to add 20 gm. of new deflector placed at the entrance of the material even after one run since so pipe into the box was adjusted to as much had sublimed. Whatever the sure a uniform distribution of the case, the collected sublimate was stream of air to the two tiers of cages. At the opposite end of the box--the outflow end--the air from each box was exhausted through a 7-inch pipe fitted with a damper and connected to a large central exhaust fan (figure 3). About 4 inches from the entrance of the 7-inch pipe into the box, the fumes of the substances listed were intro duced into the inflowing air. Spe cially designed pyrex glass flasks (figure 4), 72 inches long and with a diameter of 1$ inches, were used to hold the heated waxes. These flasks were made with' a side arm and tube that extended to the bottom. The top of the flask was ground, and into this fitted a short tube H inches in length. This short tube was inserted into a large rubber stopper that fitted tightly into a hole cut out of the 7-inch pipe on the under Side (figure 2). The flask in turn was placed in an electric heater made to cover it completely below the side arm and ground glass top. Rubber tubing connected the side arm with a compressed air reser voir and a gentle stream of air blown Fic. 2. Electric heater for maintaining chlorinated compounds at a constant tem perature in place at inflow end of box. through the melted wax kept it in motion and assured uniform heating. always removed from the upper part Into each flask was inserted a long stem, of the flask and a clean top used each centigrade thermometer which was day. No sample was ever used for kept in place and could lie rend at any more than two runs. time above the 7-inch pipe through The flask plus the contents was care which it passer! (figure 2). fully weighed at the beginning of the Approximately 30 gin. of pulverized run and at the end, and the loss in chlorinated naphthalenes or 20 gm. of weight used to calculate the average mfv r-lHi- i --3^*2fS.'S~- WATER_PCB-SD0000030409 .. ___ -,/'ihTi TTr*~JTrifii~t irfllitiiTTi^1 f)iHiiThiT^nTi*lPi? 2SS JOUIIXAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [Sept. 1037 amount in a cubic meter of air per minute as determined by a series of Pic. 3. Outflow end of two boxes showing connections to exhaust system. \. * F:c. 4. Py.-ox glass flask flowmeter readings. Tito figures ob tained were not absolute because of slight variations in the air flowing through the boxes and because of deposition of material on the ther mometers and on the inside surfaces of the box, but they checked well with direct determinations through air samples. Approximateiy 1 hour was allowed for the wax in the flasks to melt and come to a constant temperature. At that time the box doors were tightly closed, air bubbled through the flasks, and the blowers turned on. This was the beginning of the exposure period. By means of rheostats on the fans and dampers in the outflow pipes, the amount of air flowing through the boxes was adjusted and an attempt was made to keep the four boxes as uniform as possible--usually between 165 and 175 c.f.m. In the first group of experiments the following substances were tested: 1. A mixture of trichlomaphtha- lencs plus a trace of tetrachlor- napthaiene. Chlorine content 49.9 per cent. 2. A mixture of penta aud hexa- chlornaphthalenes. Chlorine ' content 62.6 per cent. 3. A mixture of 90 per cent penta and hexachlornnphthalenes plus 10 per cent refined chlorin ated diphenyl. Chlorine con tent 63.0 per cent. 4. Chlorinated diphenyl. Chlorine content 05.0 per cent. The compounds were selected as rep resenting a certain range in chlorina tion :mu also because of their industrial importance. In each instance SO ani mals were exposed, 10 rats being placed in each cage. They were fed Purina WATER_PCB-SD0000030410 eol. -o. ?\ EFFECTS OF CHLORINATED HYDROCARBONS 289 Dog Chow supplemented by lettuce, eggs, milk and cod liver oil. This first group of experiments was begun on July 1, 1936 and the last exposure to number 2 (penta and hexacblornaphthaIer.es) was on November 16th. The exposure to the other three compounds ceased upon November ISth. On October 15th representa tive animals were taken out of expo sure from groups 1, 2, and 4 and were killed after 2 months on December 15th, in order to see whether this clear wax concentrations were somewhat higher at the inflow end of the boxes. Preliminary runs showed that once properly adjusted, wax concentrations in the air remained very uniform from day to day, but to insure absolute safety readings of temperature, air flow, etc. were made every night be tween 10:00 and 12:00 pm. as well as on starting and stopping. At dif ferent times during the course of the experiment tests for free chloride were made but were uniformly negative and TABLE 1 Conditions Maintained in Inhalation Experiment from Jttlt 1 to November 16 and 18 lUTXXUb tA. xen AY. COKCXjmUTlOK or Aim mx IOtTzA-S. nsoms AY. BAAT ex* VCSVAX r. c. m*./cu. nv rongg feamr bourt Trichloraaphthalenea plus traces oi tetrachlomaphthalene................... 49.9 150-100 1.31 High 2.60 1896 16 Low 0.10 Penta and hexachlornaphthalenes.... 62.6 160-170 1.16 High 2.19 1S64 Low 0.51 16 $G% penta and hex&chlornaplithalenes plus 10% chlorinated diphenyl. 63.0 165-173 1.37 High 3.17 Low 0.64 1896 16 Chlorinated diphenyl......................... 65.0 165-173 0.57 High 1.19 1S96 Low 0.22 16 period would bring about recovery in the affected livers. Animals from group 3 were similarly removed from exposure on October 4th and killed for examination on December 4th. The average length of exposure was 1G hours daily for G days a week. Each morning at about 9:00 exposure ceased, and between this time and 4:00 p.m. the rats were cleaned, fed, weighed, etc. In order to secure uni formity of exposure the cages wore shifted on a regular schedule, since showed that under the temperatures used no decomposition occurred. Table 1 shows the temperatures at which the various substances were held, the average concentrations per cubic meter of air, and the average exposures from July 1 to November 1G and IS. The concentration of ddorinated naph thalenes and chlorinated diphenyl in the air af workrooms.--Tabtc 1 shows that animals have been exposed to varied concentrations of the substances under mr -T*tfihVfrrrfr 290 JOURNAL OF INDUSTRLVL ETGIENE AND TOXICOLOGY [Sept. 1937 test. The concentrations employed may be regarded as fairly representa tive of industrial experience. Prior to the initiation of inhalation cspcrimerits a number of estimates of chlo rinated hydrocarbons in the air of dif ferent factories were made and the in question is passed over heated plati num in an electrically heated quartz tube and the effluent gas scrubbed in a column of glass beads moistened with sodium carbonate containing a trace of sodium sulfite. The beads are then washed down and the chloride deter- Fiq. 5. Dimensioned sketch of combustion tube and absorption apparatus. concentrations chosen for inhalation experiments depended on these exami nations. The technic of analysis and the apparatus employed were the work of Frederick R. Millhiser and William F. Hemperiy. The method and apparatus used for determining concentrations of ehiorinated hydrocarbons in air were adapted from well-known procedures. The air mined nephclometrically as silver chloride. Tcb'oens (1937) has recently de scribed a method and apparatus which should be equally satisfactory for this work. In figure 5 we show a dimen sioned sketch of the combustion tube and absorption apparatus as used by us in both our laboratory and field problems. The absorption tube is WATER_PCB-SD0000030412 sol. IS, no. 7] EFFECTS OF CHLORINATED HYDROCARBONS 291 somewhat easier to wash down than is have been exposed for 1G hours to an Tebbcus's but the essentials of the two atmosphere constantly impregnated devices are the same.* with the substance under test, whereas In both cases conversion of the chlo human exposure is usually a variable rinated hydrocarbons to hydrochloric quantity, intense for a short time and * add and subsequent absorption as then negligible. It is our opinion at sodium carbonate should be complete. the present time that the concentra In our case concentrations in air are tions of chlorinated hydrocarbons used apt to be very low--the objective is' in our experiments would be dangerous less than I mgm. per cu. m. Conse for workers in the case of compounds quently the amount of silver chloride above trichlornaphthalene in chlorina actually formed in the final reaction is tion. Fortunately it is easy to venti so small that it can not be determined late processes of manufacture which either gravimetrically or by titration. require these substances and to reduce For this reason we have been forced to air contamination practically to the use the nephelometric procedure which vanishing point. Such treatment of is sensitive to concentrations as low the problem at once removes both the as 0.1 mgm. per cu. m. of air. -possibility of systemic poisoning and It is doubtful if the sampling rate the annoyances that arise from cases should exceed 1 liter per minute as of acne. conversion of the chlorinated com pound to hydrochloric acid and its Results of Inhalation Experiments. subsequent absoqjtion are not efficient 1. Animals exposed to a mixture unless the velocity of the gas stream is of trichlomaphthalenes plus small low. Another precaution to be noted amounts of tetrachlomapthalene. is that the method is not selective-- Living animals were apparently en any cidorinated substance will be tirely normal. Autopsies performed determined. Furthermore it does not near the end of exposure seemed to distinguish between solid particles and show slight swelling of the liver, and gases but determines them all alike and microscopic examination occasionally the results must, of course, be com showed swollen and hypergranular liver puted in terms of total chlorine. cells. The changes were, however, At the present time we possess in never more than slight. ` formation as to the amounts of chlo 2. Animals exposed to a mixture of rinated naphthalenes in the air of 30 penta and hexacnlornaphthaienes. different plants, and in a number of No abnormalities were observed in the instances the measurements have been living animals. Rats were killed and repeated several times. Frequently autopsied every G weeks. In the first the amounts have been greater than animals sacrificed liver changes were those used in our experiments, but it observed. Tuese were swelling of must be remembered that the rats ceils, slight granulation and hyaliniza- * The combustion and absorption appa ratus suited to field sampling is now made by Willson Products, luc., Reading, Pa. rho equipment includes tiowmeter and suction pump with a carrying ease. tion. In September and October these conditions were somewhat more ad vanced, and in November the process became stationary. There were highly 292 JOURNAL OF INDUSTRLAL HYGIENE AND TOXICOLOGY [Sept. 1337 granular cells, hyaline inclusions and exposed to these compounds without mitotic figures, but no more than 2 illness of any sort. months previously. i 3. Animals exposed to a mixture of The functional appraisal of the liver 90 per cent penta and hexachlor- damage caused by certain chlorinated naphthalenes plus 10 per cent refined naphthalenes and by chlorinated di chlorinated diphenyl No abnormali phenyl.--There are no tests of Ever ties were seen in the living animals. function useful in such minor degrees . After 6 weeks the Evers showed of Ever damage as were produced in - changes similar to those in the animals these experiments. Indeed the ani- . .. ; exposed to penta and hexachlomaph- mals resisted the injury so perfectly _\.v thalenes. These advanced in grade as to display no abnormaEties except - during August and September and upon histological examination of the .. 1 vr. then became stationary. Ever. The situation was perhaps . A Animals exposed to chlorinated similar to that met in industry, where, : diphenyl. No abnormalities were seen barring acne, the health of workers in - in the living rats. After 6 weeks' ex these compounds has been good with . . posure there was slight Ever damage the exception of the fact that in iso which advanced during the next 2 lated instances jaundice has occurred months. The changes consisted in which upon at least three occasions slight to moderate swelling of the Ever has gone on to acute yellow atrophy. _ cells, an increased granularity and During recent years this disease has - many mitotic figures. Eyaiinizaticn been seen following administration of * was always present as a result of carbon tetrachloride, arsphenamine inhalation of chlorinated diphenyl. and cincophen. In the case of carbon . Summary of the first inhalation ex- tetrachloride it is known that a low - perimenL--In these experiments care calcium diet and alcohol favor the pro- .. ful observation of appearance, body . duction of Ever damage. For the weight, activity, blood, and urine acute yellow atrophy that occasionally - showed no abnormalities of any sort. complicates use of the other two drugs Yet after 6 weeks' exposure ail the no cause can be assigned. One cannot .' compounds with chlorination above produce acute yellow atrophy with . trichlornaphthaiene caused minor de arsphenamine, but somehow or other grees of liver damage, and no changes this now and then does happen to pa whatsoever in other organs. There tients under antisyphilitic treatment. ' was no acute yellow atrophy or any It occurred to us that something of thing suggesting it except that a slight the same sort might be involved in this degree of liver damage was always problem. The human cases have been . present and was quite clear in the liver scattered and few. They have been sections examined microscopically. isolated instances out of large groups . Ibis damage had no detectable effect of healthy employees svno have had ' on the health of the animals. They equal degrees of exposure. It was our ' held their weight, ate and behaved idea that perhaps many of these people . normally, being in every respect similar got liver changes such as existed in our to the many people who have been rats, changes not recognizable through WATER PCB-SD0000030414 Ml. IS, no. 7] EFFECTS OF CHLORINATED HYDROCARBONS 293 any means other than autopsy exami nation. If, upon the substratum of such changes, they got an acute liver disturbance, acute catarrhal jaundice, not a very common disease but one 1 died Nov. 10th between 5:00 and 10:30 pun. 5 died Nov. 11th between 4:30 and 10:30 p.m. ` 1 died Nov. 11th between 4:30 p.m. and 9:00 a.m. Nov. 12th. which any of us may experience, would this relatively innocuous disease go over to acute yellow atrophy? Knowing that our inhalation rats had liver changes, but changes too 2 died Nov. 12th about 1:00 a.m. 1 died Nov. 13th between 5:00 and 10:00 pun. 4. Chlorinated diphenyl. Fed at 10:00 a.m., Nov. 10,1936. 10 rats. 1 killed Nov. 10th at 10:30 p.m. (almost slight to cause recognizable symptoms, we decided to test their resistance to substances known to cause liver de struction. Carbon tetrachloride and alcohol were selected. Having infor dead). 2 died Nov. 11th between 4:30 and 10:30 p.m. 1 died Nov. 12th before S:30 a.m. (stiff)* 1 died Nov. 13th between 9:30 and " 11:00 aun. mation that 1.0 cc. of carbon tetra chloride plus 1.0 cc. of ethyl alcohol per kilogram would kill 14 per cent of normal w'nite rats, we reduced the dose to 0.75 cc. carbon tetrachloride and 1 died Nov. 16th between 10:15 p.m. Nov. 15th and 9:00 aun. Nov. 16th (stiff). Controls: Fed at 9:30 a.m., Ncv. 12,1936. 10 rats. 1 killed Nov. 13th for normal liver.. 0.75 cc. of ethyl alcohoL This tabulation summarizes into the This mixture was given by stomach facts that: `. tube to the following groups of ani 1. No normal rats were killed by mals and with the results found in the carbon tetrachloride and ethyl alcohol. following tabulation. 2. No trichlomaphthalene exposed Rats fed 0.75 ee. per kym. each of carton tetrachloride and ethyl alcohol by stomach lube rats were killed, and this finding agrees with our inability to find lesions of moment in the livers of the animals that inhaled this substance. 1. 'frichlomaphlhulcne plus trace of tctra- 3. The penta and hexachiorinated chlomavhlhaler.e. F1 at 9:00 a.si., Nov. 10, 1926. 10 raw. No deaths. 2. Pcnt/i and hexachlamaphthalencs. Fed at 9:30 a.m., Nov. 10, 1925. 10 rats. 3 died Nov. 11th between 4:30 and 10:30 p.m. ,- naphthalene, the mixture of these with 10 per cent chlorinated diphenyl, and "finally the latter substance alone seem to have produced conditions lowering the resistance to an agent capable of 1 died Nov. J2th at 3:20 p.m. 1 died Nov. 12th at 4:30 p.m. 1 died Nov. 13th between 9:30 and 11:20 am. 1 died Nov, iSLh between night of Nov. 14th and 12:30 p.m. Nov. 15th. 2 producing serious liver disease. 4. The sole lesion produced by this test of liver function was acute yellow atrophy usually accompanied by jaundice. died Nov. 16th between 10:15 p.m. Nov. 15th and 9:00 a.m. Nov. ICth (stiff). 3. 50 per cent penia and hczachlomapkthalcnes and 10 per cent chlorinated diphenyl. This test of liver function was ac complished with a substance which itseif is an organic chloride and curi ously enough it is the only substance Fed at 10:00 a.m., Nov. 10, 1936. 10 raw. wc were able to find that was effective. ! ^`TTT'Tri*^^*;*?***** L!4*!sS&xHt*?3&s$3S5*i -if ^ i TI 4 irj j, ii:ii]j!h ^&==si li i. -JJ II:f i4/$jB5<Zc5wa^-->rrs^v ^ i 3| ;' ZJLi'.'l >*...*SSL -hJ i. ?4 ; J-i is -jgSSggSp|i;|: IsliSsssa5; 4 i y i 'TSOa'jsn^hiSStfi-CTaSvtrSai .tm,. , )SSS2?IWJi.-5^2!=S..Sr.l<: . SgbsgSSg ..j {it ! is^ag^'^Kj-p 35i5i3??iK.:se i ill 1 . isgsgtac^sr rt; * i<; Hvs5i?3S? * i. m ! uriforansm *; j r*. j '2 ! !'l t.. Saggjrtsr* . r\ i r S;r-ar^<j^SvS/ej--S-s4a=^r-ra#?- *-_.' i: rM.ij ?i:: _ ?*g^?Sj?r`r ^ ! "I j` ' r^^^3rfci "aITsasV-r^--'a.T; :t4a- 1<; >-:1!1 '!1r::-:' -* Ji-1 ? J-n :* i ~\ 1 < j ;;.l -;: -sggsc^ ' r. ! .'S' - - j : > * y i-5/"i5 ^ .4 ;; * ** 7: i I. rv;V'.CI -** 5 i .?j-~*-r 7=^ .- v".._` *' ' ;-* ,,'^,,cr7J'7:-VrrSu^,*>v` ' '- ' t:.* .. '* ' *--r ^ 204 JOURNAL OP INDUSTRIAL HYGIENE AND TOXICOLOGY [Sept. 1S37 In ^animals with the minor degree of liver injury which has been described we were unable to produce acute yellow atrophy or any variety of liver effect with arsphenamine, cincophen, butyl chloride, ethylene chloride and tetrachlorethyiene, but carbon tetrachlo ride was uniformly effective in dis closing the existence of liver damage. Recovery from User damage following removal from exposure.--Animals ex posed to trichlomaphthalenes plus traces of tetrachlomaphthalene being practically normal on removal from exposure need no consideration. In the case of the penta and hexachlor- signs of liver injury will be many months in returning to normal. The effect of high concentrations of trichlomaphthalenes with traces of Leirachlomaphthalene.--The first group of inhalation experiments showed that this material in concentrations -averag-ing 1.31 mgm. per cu. m. of air was relatively innocuous, judged both by direct observation and by the carbon tetrachloride test. This is an in teresting fact since such compounds cause acne, though less potently than substances of higher chlorination. In order to explore the matter further one of the inhalation boxes was arranged TABLE 2 Conditions Maintained during Inhalation of High Concentrations op Trichlornaphthalenes plus Tetrachlornaphthalsne HATtlUL . Trichlomaphthalenes plus traces of tetrachlomaphthalene TtStf. AT. COKCCmtATXOX or aix in xoz TOTAL AT. SAXLT tXToaone rsrosritm c. frtfVcB* m. 137-200 10.97 ra/194 High 16.40 Low 5.7S lavra 1232 koura 16 naphthalene, rats after 2 mouths' re moval from exposure still showed swollen liver cells, increased granu larity, hyalinization and mitotic fig ures. The condition was not advanced as compared with rats killed at the time of removal from exposure but on the other hand 2 months were insuffi cient for recovery. The same Gildings were true of the mixture of pencil and liexachioriruitcd naphthalenes and chlorinated diphenyi and for chlorinated diphenyl alone. Apparently the changes induced in the iiver cells by those substances are exceedingly persistent and one must expect that an individual allowing any so that fumes from four glass contain ers were delivered to the airline instead of one. This resulted in the conditions shown in tabie 2. The animals sub jected to these conditions showed no clinical effects of any sort. After 1 month the liver ceils were slightly swollen and over-grantiiar and there worn occasional mitotic figures. These changes were similar to die early effects of more highly chlorinated compounds, and progressed only slightly during the third and fourth mouths. When rats in this condition were given carbon tetrachloride ami alcohol, in some instances their livers showed massive central necrosis and WATER PCB-SD0000030416 jsig if- vol. iS, no. 7] EFFECTS OF CHLORINATED HYDROCARBONS 295 in others this did not develop. Fifty rats were used in this experiment and one may conclude that while the trichlornaphtlialen.cs are in no degree as toxic as those of higher chlorination they are not entirely free from effects upon the liver if high enough concen trations arc inhaled over long periods. The creels of high concentrations of penta and, hexachlornaphthalenes.-~ There can be no doubt as to the dam age done the liver by these compounds but in the concentrations so far re ported no symptoms were caused that could be recognized clinically and nothing approaching acute yellow the liver after 35 days of freedom from inhalation. It is thus evident that penta and hcxachloroaphthalcncs arc markedly toxic compounds and that recovery from their effects is extremely slow. The effects of penta. and hexach.hrmphdhalenes, the mixture of these with 10 per cent chlorinated diphenyl, arid of chlorinated diphenyl alone, when inhaled in low concentrations through an 8-hour day instead of a 16-hour day as in the first experiment.--Since steady human exposure to any of the compounds tested would invariably be for 8 hours rather than 16-hour periods, a further TABLE 3 Coxditioxs ILuxtaixed ochixg Inualatcox or High Coxcextuations of Pexta axs HEXACHIOKXAPirrHALEXXS MATXttLifc Penta and hcxacbiornaphthalenea........ rruf. c. 137-1SS AT. COSCrXTBATtO* or xin tv box Plfjcu* m. 3.S3 High 14.0 Low 5.75 TOTAi AT. OATbT SXVOBOSS sxroeoAs kV9 60S toura 16 atrophy occurred except through the use of the carbon tetrachloride test. On December 1, 1936, SG rats were subjected to the conditions sumjnarized in tabic 3. This experment was terminated on January 21. All of the animals lost weight and appetite and dcatlis began after 8 days of expo sure. Fifty-five rats died, most of them heavily jaundiced, 8 were killed for microscopic examination, 9 were killed by the carbon tetrachloride test and 8 lived through the period of exposure. The livers of the animals killed for examination showed marked fatly degeneration, central in type w.th necrosis of liver ceils. Surviving animals showed pronounced changes in inhalation experiment was arranged under the conditions shown in table 4 which are quite comparable to those in table 1 except for the shorter inhala tion periods. Eighty rats were used for each of the three compounds tested. None at any time showed the slightest evidence of illness. Microscopic ex amination beginning after 6 weeks' exposure showed swollen liver colls, excess granulation, hyniinc inclusions and occasional mitotic figures. These changes did not progress and were very similar to those in the animals exposed for 16 hours under the conditions set forth in table 1.* The carbon tetra chloride test was uniformly fatal to them and one must conclude that con- 8381S1 u5.*i5 tJ v~ ; i ' VIITS 5 :2 \V: I r i- fl-tn ** i* a- ... ** t :.v-V- : ; - iC iv l> * *1^ ; t* i*Vjt v* r' ~^ WATER PCB-SD0000030417 V.. .liSiMlGiiif* Y.Vm jy'M*tfSi*gsi5Ws*4t5SiSipf .t i papgS -ill Jjfej hi ISSgiSS :I IS! }i -ggpg 3il|i ^esr#5 '.il if 'r3a*fw5uo>!tawUPCM(?ac.^J* I[ '`j-lt '-!i i im'- s-SSIskSwS. ',-S i MSi lii i: fjEgasI. $ . '< ^ejfSggS if {j Hfesirli I <3 Si. *-^^Svt2" *" * 4 ;? i W ! .-- jE *i $ i : 5svrgt$gg - i j !; WP&ZTl \ yj \ \ H5Sa|S2*i^ -) j | ! * SgTOtiSs * ifit i`. 'f J.: ^ , . ,'ii ^ '< ' --Tr 1ii.*f* '4.i. --!?*. a--itTi - ?kc-c *: i i * cgy^4 * ; ..ri ;u ^ ' ^3^a --v ":"'-] t*: 296 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY {Sept. 1037 centrations of these compounds such as wore employed cause a certain de gree of liver damage even if inhaled for but 8 hours daily. This injury is resisted successfully by the rats just as was the 16-hour injury but it is none the less there and destroys the ability of the animal to resist the carbon tetrachloride test. Summary of i7ihalalion experiments. The findings that have been described briefly will be amplified upon the pathological side in a further paper by Dr. G. A. Bennett. What has been given is, however, sufficient to indi men never inhale enough of any of these substances to get acute yellowatrophy. They may, however, acquire a substratum of liver damage upon which acute yellow atrophy may de velop. Experience in a number of plants has shown how easy it is to reduce concentrations of these com- . pounds practically to the vanishing. point, and every effort should be made to attain such conditions. Gross Feeding Experiments The various compounds used were ground as finely as possible and mixed with a standard balanced ration for TABLE 4 Conditions Maintained during Inhalation or Low Concentrations of Three Compounds during S-Hour Instead of 16-Hour Periods . 1UTZXUI. TINT. AT. COXCXCTJLATtOSI or Atm is sox TOTAL 1 AY. DAltY EXfOSCAV KUr&mmo Punta ann h*Tnr*Mnms.phfchalftnfl............... c. 149-193 *w*/cu. m. 1.44 90% penta and hexachlorn&phthalenes ^0% riniLted diphenyl............. 150-197 1.65 Chlorinated diphenyl.................................... 153-174 0.93 ran$ High 2.53 Low 0.42 hours 920 High 3.30 Low 0.56 Eigh 3.23 Low 0.03 912 920 i` kauri s 8 s .cate that compounds more higlily chlorinated than trichlornaphthaiene are capable of causing liver injury when inhaled steadily in quite low . concentrations. It is an extraordinary thing that even the most searching examination fails to show injury in any --other region. It is not easy to grade | the toxicity of the different compounds I tested, but the chlorinated diphenyl is I certainly capable of doing harm .in very I low concentrations and is probably the J most dangerous. Industrial experi- I ence combined with these experiments make it appear probable that work- the white rat. The food was placed in a single container, at which the 10 animals in a cage had an equal e the amount of the compounds aucca being reduced as the number cf rats lessened during the feeding period. The experiments were designed 'ri : an idea as tc the possible :o::i'i:;- the compounds selected and, if tome::;' was observed, some idea as to the -ute or sites of damage. The experiments were- successful in both respects, and indeed one may anticipate that future appraisals of the possible toxicity of chlorinated hydro- ~'^^i~7CSru7)--q WATER PCB-SD0000030418 eoi. 10, no. 7] EFFECTS OF CHLORINATED HYDROCARBONS 297 carbons may often be made by simple rats had died. -The last one was obvi feeding and do not require the elabo ously ill and was killed for autopsy rate apparatus and the expense neces examination. 'All animals lost weight sary for inhalation experiments. The from the beginning and were ill. At essential reason for this opinion resides autopsy the liver alone was affected, in the identity of the lesions produced the lesions observed being similar to by both methods, and the fact that the those that have been described for different compounds so far studied tetra and pentachlomaphthalenes but seem to arrange themselves identically worse. so far as toxicity is concerned which 90 per cent penta and hesachlor- ever way they are administered. naphthalenes and 10 per cent chlorinated Trichlamaphlhalene plus traces of diphenyl.--Feeding began May 4, Ictracklomaphthalene.--Feeding began 1936 on a 3 gm. dosage. On May on May 4, 1936 and continued 16th feeding was stopped but all until November 2,1936. In the begin the animals went on to death, the last ning 3 gm. of this mixture were added dying on June 8th. An autopsy the to the food for 10 rats each day, liver lesions were extremely severe and and this concentration was maintained of the usual type. throughout the experiment. All the On account of the high toxicity of animals were killed except the last the 3 gm; dosage, four rats were given which died on Nov. 2. There was no a 0.5 gm. dose every other day. Feed loss of weight and no appearance of ing began June 24, 1936 and the last wasting illness of any sort. The single animal was killed Sept. 11th. There death was due to some variety of re were no deaths but all the animals lost spiratory infection which had no rela weight. At autopsy the liver as usual tion to the material inhaled. was the single organ affected, the Histological examination showed lesions being characteristic .and ex slight changes in the liver but nothing tensive. of great significance. Chlorinated diphenyl--Feeding be Tctra and pcntachloniaphthalcnes.-- gan May 4, 1936 on a 3 gm. dosage Feeding began on June 29, 1936. A and was discontinued on May 10th. dosage of 0.5 gm. daily was employed. Seven of the 10 rats were dead by May All the animals sickened gradually and 12th. The three remaining rats were either killed or died by August gained in weight but were sacrificed 20, 1936. for autopsy purposes on July 8th. At autopsy both grossly and histo The liver changes began at once. logically the liver was the single organ There were no changes in other organs. affected. The liver colls showed sweil- A second group of 10 rats was fed a ing, hypergranuiation, hyaline inclu much smaller dose--0.5 gm. every sions and vacuoiation. Hero and other day. Feeding began May 20, there necrotic ceils were found. There 193G. The first rat died on May 29th was a tremendous accumulation of fat. and four more before June 24th. The Paula and hesachiomaphihaJcncs.-- remaining were sacrificed. Those rats Feeding began May 4, 193G, a 3 gm. that died showed losses in weight, dosage being used. By June G, nine while those sacrificed had recovered * ikdbEt *T1iiTCl 298 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [Sp<. 1037 their initial less in weight and were and rabbits, the dosage being calcu gaining. The liver lesions were similar lated on the basis of 4 mgm. per cu. m. to those found in rats fed penta and of air. Again similar results were hexacHornaphthalenes plus 10 per obtained. In all such experiments cent chlorinated diphenyl but not so there must of necessity be differences marked. in the degree of effect but invariably Summary of gross feeding experi ments.--Of the various materials fed rats in large doses trichlomaphthalene the liver was the sole organ affected and the lesions were those already described many times. * plus traces of tetrachlomaphthalene Discussion . was quite innocuous. Tetra and pentachlomaphthalene showed definite liver damage. Penta and hexachlcrnaphthalenes caused a similar grade of injury. The addition of chlorinated diphenyl to penta and hexachiornaphthalenes increased the toxicity. Chlo rinated diphenyl alone produced liver lesions but in the dosage used was less effective than when mixed with highly chlorinated naphthalenes. In no case did the compounds used produce acute yellow atrophy but the lesions ob served indicate this might be possible if one found a dosage which could act for the proper period of time. These experiments leave no doubt as to the possibility of systemic effects from the chlorinated naphthalenes and chlorinated diphenyl. ' As in the case of the effects upon the skin, the degree of chlorination seems to determine the systemic toxicity, and it is a striking thing that when trichlomaphthalene b reached systemic effects are never marked and are produced with the greatest difficulty. It is most remark able, too, that all the compounds tested attack the liver and . the liver alone. During the past few months we have determined the organically combined chloride in the livers of ani Feeding Precise Doses by Stomach Tube The compounds employed were sus pended in gum acacia. -In figuring the dosage the total amount a man of 50 kg. would inhale in an S-hour day assuming an air concentration of 20 mgro. per cu. m. was first calculated and reduced to milligrams per kilo gram. The rats and rabbits received this dose each day. The compounds used were those employed in the gross feeding experiments and the results were essentially similar though the lesions were less severe. Subcutaneous Injections mals very' severely poisoned by penta and hcxachlomaputhalenes but have found no increase over normal figures, though the livers, as determined libtologically, were very severely affected. At the present time we are condtxring inhalation experiments on a chlorin ated diphenyl containing 55 per cent of chlorine instead of G4 per cent as in the case of the experiments reported in this paper and on a compound with a chlorine content between tri and tetrachlomaphtiiulttie. V/o are also determining the degree to which, the diet may increase or decrease toxicity, thus being suggested by similar work The same gum acacia suspensions upon carbon tetrachloride. were injected subcutaneously into rats In the basis of these experiments 1 WATER PCB-SD0000030420 vol. 13, no. n EFFECTS OF CHLORINATED HYDROCARBONS'. 299: ana on many field determinations of different compounds in the air of work rooms, it appears safe and it is cer tainly easily attained, to ventilate so that the air breathed does not contain more than 0.5 mgm. per cu. m. of any of these compounds above trichlornaphthaienc. In the case of the latter compound concentrations of 10.0 mgm. are permissible. We know from many examinations in many different plants that such concentrations have been greatly exceeded during the past 20 years, and wc are conscious of the fact that our rat exposures have been inexorably constant whereas human exposure is never so ordered. Time and careful observation may change these opinions as to standards, but today we are convinced they are safe. Impregnating tanks and other arrangements utilizing the chlorinated hydrocarbons arc easy to hood and to safeguard. .Compared with benzene, lead tetraethyl and many other com pounds, these .substances are very little toxic and operations employing them can easily be safeguarded. It may be argued that where possible tricnlomaphthalene should be used, but this compound will cause acne and if employed very carelessly might do more. Furthermore, higher chlorina tion is often essential for highly prac tical reasons. The solution consists in thoroughly adequate ventilation plus good housekeeping around all wax containers. . . . A final word of caution bears upon the use of carbon tetrachloride as a wax solvent. Obviously this compound adds readily to the toxicity of the highly chlorinated waxes. If carbon tetrachloride is used, ventilation should be excellent, but in our opinion it would be better to dispense with car bon tetrachloride and depend on other solvents, especially upon those con taining no chlorine. BIBLIOGRAPHY Cocp.TOis-ScFTTr: Etude sur l'intoxication protessior.nello par le trichloronaph- talonc. Ann. dc med. legale, 14, 422 (1954). Ahstr. o paper by Tourajne, A., axd Mstrsl, B.: Dermatoses pratessionnclles par la naphtaline et ses derivds. Prat. m4d. frang., IS, 335 (1934). ' Funk, F. B., and Jarvik, N. E.: Action of certain chlorinated naphthalenes on the liver. Proe. Soe. Exper. Biol. & Med., SS, 113 (193G).' Lehmann, X. B.: Kurzcs Lehrbuch der ' Arheits- und Govrerbchygione. S. Hir- zei, Leipzig, 1919 (p. 251). ' Mittelstadt, 0.: Gewerbeschadigungea durch Haltax (Trichlomaphthalin). Inaug. dissert., Jena, 1935. . Tebbens, B. D.: Portable combustion ap paratus for field determinations of chlorinated hydrocarbons. This J., . 10, 204 (1937). Telezt, L.: Die Pernakrankheit (Clilor- acne). Klin. Wchnschr., 6, 897 (1927). Tocraine, A., Solents, Mntrel, B., and Aubrun : Cinrpiantc-quatre caa de hematites par trichloronaphtalinc. Bull. Soc. dermat. ct syph., 41, 255 (1934). 'C2s:5ir*j?<}t' -tji i; i?ll l;4|- -jZa?5\i:z , gSgas&ga H : ^^jSSSSsWaeftStgS&If I{H* ii: jSeSSgre^g f! *|-i*, j <! TX^C^.ir,,, j~ "j * r _S5*g5?r*3 i*5l( ?!: TarJS&.mza- s si gafagj 3i?| *>.* -*? /!*- i I*.f ze:^r*L>'J;*iSf.?zi j*? *!?I*l>h; 1 I> .` J i:j --.*. v a 4 4 !'-r}* * T ; g(jS&$ : l j t? *~r| :;1 1 mV2h> -.1 ' - 1-i ?; tjrvW':*4i =-I r/Scva^a :.*,--5 >5* >* ftTu4-'i . yv ,. - *-,*.* ^A~*.--V-*- V *--V '1. m- tI { i 'i j 300 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [Sept. 1937 Discussion Dr. Cecil K. Drinker: We have had incline them to. First of all I shall ask quite a long morning, but since Dr. von Oet* Dr. Sayers if he will open the discussion tingen has to leave, I am going to ask him for us. to say a word. Dr. R. R. Savers (Chief, Division of Da. W. F. ton Oettinosn (Director, Industrial Hygiene, National Institute of Haskell Laboratory of Industrial Toxical- Health, U. S. Public Health Service, Wash- -gy, Wilmington, Delaware): I wish to con ington, D. C.): Dr. Drinker, Ladies and gratulate Dr. Drinker and his group on the Gentlemen: It gave me a great deal of pleas-, very interesting and excellent work on the ure to have the- opportunity to listen to the subject. I also wish to congratulate the excellent and interesting discussion and Salowax Corporation on the fact that they presentations made this morning. Dr. didn't spare any effort to elucidate the prob Drinker, and I might add, the members of lem so that the dangers which apparently the Halowax Corporation, have discussed are connected with the use of this material this matter with us, as well as the manu can be controlled. - facturers of the diphenyls. ' I personally believe that there is no tojdc We have been doing some work but not material which cannot be.used.safely if it is nearly as extensive ss that carried on here. handled properly. In order to handle such The work that we have been doing seems to . 'materials properly during the differen t oper- confirm the work that Dr. Drinker has car aUonsTt'is absolutely _esseatiai"to"be famll- ried on. I think there are certain parts of it iarlntlf thTfoxicity^aad especially with the that we can very well keep in mind.. prnechanismpf.the action.--Since there is no f~ We are.deaiing with toxic substances, but `"animal which corresponds in all Its physio we_are_deaiing wnthToxic substances each. 1 logical characteristics to the human being and _eyery. day. in industry*--harmful sub it is important that such compounds be stances, hazardous substanceiFTKaftheae studied with different species of animals so carTb'e controlled is well known, no matter that one can get a cross-section of the poten What they are', and that we can use them and tial dangers. j use them in. a safe manner. Dr.~Drinker In addition, this fact illustrates the im "has called your attention to the fact that portance of the pre-employment and peri this apparently has been done and can be odical examination of workers in such work. done in any of the plants where halowax is As was pointed out by Dr. Drinker, there used. I have information from one or two are individuals who are apparently more of the other men who are in the audience ' susceptible to certain toxic agents than that in plants where damage has been done others. On the other hand, we have to try --skin damage and apparently liver damage to detect the very first signs of incipient to some of the workers--the manufacturers _ poisoning in order to prevent more serious have since corrected the conditions and ap damage. In the present instance it is very parently have adequate engineering con difficult to find these very first symptoms trol supplemented with adequate medical because there are no adequate methods, but controL I trust that in time the scientific profession After all, the proof of the pudding Is in will supply the medical profession with the eating. If the man does not get the dis adequate methods which will allow the ease your engineering control is successful. physician to dutermine incipient damage or If he does get the disease, no matter what the organ before serious clinical symptoms you have done in the wny of engineering, become manifest. you haven't been successful. We must go Thank you. hand in hand. Neither profession alone Afternoon Session can be successful. We have got to work together whether we like it or not. I hap Dr. Cecil K. Dninker: In this discus pen to like it. sion I will call cn one or two individuals and There is one other thing. Asjaraa mate- I shall expect others to speak as their notes riala are concerned I think we ought to C9 - ' . . ' *" C T nit .yw /i-.rs WATER PCB-SD0000030422 oi. 19, no. 7] EFFECTS OF CHLORINATED. HYDROCARBONS 301 smart enough to use every one of them aae- number of so-called cases. All I con say is ceufuily. If thia.material is,particularly a confession of ignorance on our part at that adsptad_ar-au:artain-purpoe-w.should.uae time and I was very glad to see that your R.^ Another material that was used pre digging into the literature as thoroughly as vious to this for insulation purposes, which you have hasn't elicited any further infor was flammable, caused a disaster in which mation than we were able to get, basically. approximately 167 men lost their lives. I About all we could find was about the two say approximately 167 because they were cat3 that our friend, Lehmann, experi never just sure whether it was 167 or 168. mented with. We got in touch with him and It might have been one or two more but they ho confessed that the amount that he hod were quite sure that 167 lost their lives. It given the eats and his method of determin happened when they had insulated a coble ing it were probably subject to very serious with this material. They had an armored scientific criticism, and we agreed with him electric cable. It was covered with lead on that point. . armor on the outside. It was 1200 feet long We felt, and I am very glad that it is what and was being lowered into a mine shaft ap you feel as a result of your experiments, proximately 3000 feet deep. It was lashed that if we put in an efficient ventilation sys to a steei rope. It dipped and fell to the tem sufficient to prevent the dermatitis, we bottom of the shaft. When the foreman was probably wouldn't have liver damage. inspecting it he sot it on fire with his car That is what we did entirely empirically. bide cap-lamp. The carbon monoxide from I think that is about all I have to say. I the ere killed those men that were in that have been delighted to be here and to have mine. Many of course escaped. ' as many members of my sue here-as I could Here wo have one harard, a fire haxard. jack out of the office. We have been de Now we have another one, a health hazard lighted to listen to the discussion. due to the toxic materials. We have to evaluate them. Certain of these materials, Dr. Cecil K. Drinker: May we now I am informed, are also flammable. I am make the discussion general and these who not just sure how flammable they ore but have questions please volunteer. flammability probably decreases as your chlorination increases. But your toxicity Mr. Manfred Bowditch (Director, also increases aa your chlorination in Division of Occupational Hygiene, Massa creases, according to Dr. Drinker, and I chusetts Department of Labor and Indus think that checks with other findings. We tries, Boston, Mass.): 1 would like to second have these hazards to balance but we con very strongly what Dr. Sayers has said with control them. I feel quite confident that regard to the approach to .his problem in we can. proper control rather than b y prohiojUun, Perhaps we can find other substances that whieh seems to me wholly unnecessary jet are better even than this material. At the only in this ease but in the case ox other present time I am advised that we have no toxic substances with which we have to deal. better. I am advised that in ships, such aa The problem of proper ventilation seems to submarines, it is quite important to have a me relatively simple from what I have heard non-flammable substance for the insulation of the situation thus far. The problem of of their electrical wire. maintaining that ventilation properly does not eeem to me quite so siinpie. I think it .On, Cecil X. Drintcse: Dr. Gray, would is going to bo quite necessary' that there you care to say a word? should lie a very adequate inspection of the plants using the materials and I can Dit. Albert S. Guat (Director, Bureau assure you that that will be the case hero in of Occupational Diseases, Suite of Con Massachusetts. necticut Department of Health, Hartford, When this trouble first broke in this state Ccnn.j: Dr, Drinker, [ don't think there is a little over a year' ago wc issued a quite anything that I can say except that wc used strongly worded warning letter to ail plants this material in Connecticut and we had n using these substances and it has ail along 302 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [Sept. 1037 bean my plan to follow that up with a fur of dollars and cents before you ever sold ther letter bringing the situation as we now any. That is the problem we have had in know it, as a result of Dr. Drinker's work, this cose. It has been on the market for to the uttention of users in this state. In 25 years. Until within the past 4 or 5 years that letter we shall unquestionably advo there has never been any intimation that it cate the concentrations that Dr. Drinker would cause any systemic effects. Thou has suggested this morning. sands and thousands of workmen have dealt One point that I think is worth comment with millions and millions of pounds of cer ing on in that connection is the question of tain of these materials, particularly the tri- identification of the substances. It is ex chlornaphthalenes. Then we come to the tremely important from the state adminis higher stags, combined with chlorinated- trative point of view that employers using diphenyl and other products, and suddenly these substances should know what they are this problem is presented to us. using. They are identified at present only We had asked various authorities in by numbers. If we list those numbers in a terested in public health, going bock over a warning letter which we send out there are period of 15 to 20 years, to investigate it likely to be changes in those numbers. The but there wasn't much enthusiasm for it. Haiowax Corporation is a progressive con Mr. Bowditch suggested that we take it up cern and it is going to be putting out bigger with the Drinker brothers at this institu and better halowaxes all the time. The tion, which we did. You know the results company's cooperation in this present inves of that work- . tigation has been such that I feel assured Now so- for os these changes are con that they will take the proper steps to. see cerned, they are beyond our control, to a that we in the state offices and also the users certain extent. We will manufacture a of the products will know what we are using. product--let's call it 1234. It has certain I wonder whether Mr. Brown would care chemical and physical characteristics. It to say anything about that. is supplied to a.cable manufacturer. It is composed of certain constituents. There bin. Sandfobd Bbown (President, Halo- may be seme tetrncldornaphthr.lene, tri- wax Corporation, New York, N. Y.): There chlomaphthsJene, paraffin, a little pitch are some aspects of this situation on which or bitumen, and possibly some chlorinated I think I can enlighten the medical and diphenyl. It does a certain job but he state and civic authorities, with respect to wants it to a little more plastic or he wants the commercial and practical aspects. If its viscosity or the specific gravity changed. you go into the research laboratories of any Possibly by a change in those constituents large chemical manufacturing company of only 1 per cent we can get that particular today you will find anything from one to a property. We can't sell it to him as the thousand different new chemical products same product so we put a new number on it. which have not yet been put on the market. Basically, however, if the variation in the Some of those may or may not be toxic. chlorinated naphthalene or the chlorinated The problem so far as the chemical manu diphenyl constituent hasn't changed, the I facturer is concerned b a question of timing. toxic property of that will not change. You have heard this excellent presentation When it comes to a question of cooperat given this morning by the Drinker brothers ing with the state authorities in that con ns to the work that they have done here. nection, if there wore some major change Should you take a product of which you made in a product we would have no hesi have developed, say, 5 or 10 gm. aud spend tancy in advising them-and we would also $50,000 on research to determine whether or advise our customers. Virtually every con \ not it is toxic or should you wait until you sumer of these materials at some time or have determined whether you have a market another lias been given their technical or for it? chemical designation along with their vari If you are producing only one hundred ous properties, whether they ho physical or substances a year you can see .that that chemical. would run into boxcar numbers in the way Those are some of the practical problems "m"jup iy|i St i WATER PCB-SD0000030424 vol. 10, no. n EFFECTS OF CHLORINATED HYDROCARBONS 303 1 with which all manufacturers of chemicals there was no substitute and there is none j arc confronted today, particularly in the today in spite of all the efforts we have made synthetic organic field, where the develop through our own research laboratories to ment is so rapid that our sales departments find one. can't even keep up with the research depart But we did develop--and I was most ments sometimes, in knowing what they are closely associated with it--and set up a doing. routine for bringing these men back to nor * mal health conditions. A number of them Mr. F. R. Kaiiier (Assistant to Man were sent to Dr. John H. Stokes and to Dr. ager, York Wireworks, General Electric Co., 0. H. Perry Pepper of the University of York, Pa.): I cm certainly pleased to have Pennsylvania Hospital, and the others to this opportunity to say a few words with Dr. Isaac R. Pels at Johns Hopkins. reference to the experience we have had at Through their recommendations and studies ' our York plant. I perhaps should say that we employed a trained nurse and two local again in this case experience alone has been physicians and you might say established a the best teacher. I hare lived with the small hospital and its facilities at the plant. problem at York with the men who went Through the application of quartz light, through the experience from its beginning. x-ray, mechanical removal of comedones It is only 1} years ago that we had in the and the treatment of pustules that de neighborhood of 50 to GO men afilicted with veloped in later stages, an utterly strict various degrees of this acne about which you routine where the worst cases were adminis all know. Eight or ten of them were very tered to each day for a period of 15 to 20 severely afflicted--horrible specimens as far minutes, another group who were less seri as their skin condition was concerned. One ous three times a week, and still another man died and the diagnosis may have attrib group once a week, we have in this year and uted hia death to exposure to halowax a half brought each and every man back to vapors but we are not sure of that. There a normal skin condition. Those who were was an atrophied condition found as a result very seriously afflicted do show scars, but of the autopsy but we are not definitely sure otherwise their skin is as healthy in appear that it was or was not connected with his ance as my own. I tell you wo are very work. Knowing the man as l did when he proud of the fact that they are still all em was employed, with the superficial examina ployed and the amount of baiowax that we tion that lie had, he appeared to me very are using today is even greater in quantity thin, pallid in his appearance, and I would and in types than we were using a year and not say from ray poor knowledge of the a half ago. physical make-up of the human being that With the adequate ventilation system we he presented a healthy appearance. How have installed, with the routine for change ever, it was only for a G montha' period from of clothing from street clothing to work the time of his employment that he com clothing when they come to work and the plained of this constipated condition and we reverse of that process, with the assurance advised that ho see his home physician. It that a shower will be taken before the street rapidly developed to the point where he was clotbing is again put on, we have found no in the hospital and in a very short time he recurrence of this skin trouble. Each and died. every man working with halowax products, More serious than that perhaps is the fact either from solution, from soiid compounds that we had 50 other meu in very bad condi or handling the wire insulated and treated tion as far ns the acne was concerned. The with it, is examined twice yearly with a com first reaction that several of our executives plete physical examination, including blood had was to throw it out--get it out of our analysis and efforts to determine any liver plant. They didn't want anything like damage. that for treating wire. But that was easily However, there is the point which was said but not so easily done. Wc might just very dciiniteiy brought out this morning. ns we!! have tiirown our business to the four We do nnt know as yet when this thing winds and said, `^We'll close up," because starts. I beiieve it would be of groat help \..i.*\ tJ J-- -* '' TVl'V-'- V. 'zz si s WATER PCB-SD0000030425 WfXMM X ' t\i .* A* *<1; :,**eyS3. :: I 155-afet -. .Vo-u~rr-.i'iK`Si ;4^S>s' s:'j7gi ggS*l - nr^.r ggi . _2$iS* 3S*$ ?ji "'"$?% V.K 304 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY tS?t. !SS7 to ua all La industry If some very funda Springfield, Mass.): Do I understand you mental work was done on humans, as Dr. to say that those men who were afflicted Jones mentioned before we came into this are back on the same Job? meeting, soma control in fundamental work to determine some method whereby we may Mb. F. R. Kameb: They all return ed to determine the inception of any liver dis normal skin health, as I say, end are work order. ing on the same job. We have had our men examined in groups three times since the trouble developed and Db. Cecil X. Dbixseb: Dr. Schwartz, nothing has shown up in any one man. We you have seen a good deal of this general have learned and appredate the fact thatwe subject. must handle hazardous materials, but we have learned how to handle them. It has Da. Louis Scsw.urrz (Medical Director, been a very great experience for us and for Dermatoses Investigations, United States myself, and again the best teacher. Public Health Service, New York, N. Y.): I am primarily interested in dermatitis aud Da. Cecil K. Dbineeb: Can you recol it was only because of my researches in that lect the air concentration that you are main field that I happened to come across infor taining now with your present ventilating mation given to me by Dr. Gray that there equipment? were some cases of yellow atrophy of the liver attributed to halowax. I can only Mb. F. R. Kaimbb: We have had two talk from personal experience about the skin examinations made of the air by Mr. Wil cases. I know from what I have seen and liam F. Hemperly and they average from 0.2 from what I have heard from doctors who to 0.6 mgm. per cu. m. We recently dis have been treating these cases that if they covered one condition which was found as are treated like on ordinary acne vulgaris the cause of a high reading in one of the and kept away from further large exposure examinations. The compound is one that they all get well, with the same results as is not manufactured by the Halowax Cor acne vulgaris. Those who have large pus poration that is used rather infrequently, tules may have scars resulting and those perhaps not more than 50 gallons of it in a who have superficial pustules or only come 6 month period. It is in the form of a paste, dones won't have any scars. reduced with solvent to a paste form, and it As far as prevention goes, I felt even be was not known that it contained a chlorin fore this investigation was undertaken that ated solvent in certain percentage. We dis this substance, like any other poisonous covered is only recently through the analy substance, can be handled and used in in sis of a sample and we found that that was dustry provided proper safety precautions the cause of throwing the results off to a high are taken, and I so advocated at the meeting value. It was then about S mgm. per cu. m., in Pennsylvania when this was discussed. I believe. That is going to be eliminated About ventilation and safety precau now. That is a combination of toluene and tions, I think that while we cannot, with our carbon tetrachloride solvent. When that present knowledge, detect by any chemical compound is exposed to the air the chlorin tests the early symptoms of intoxication ated solvent odor is completely masked by from this substance, the skin offers an easy the toluene. It is lost. I was able to detect way of proving whether your method of it only by removing the head of the drum ventilation is efficient or not. If there are and immediately smeiling the odor of carhon any cases of acne or of this dermatitis occur tetrachloride. As soon as that head was ring in a plant where halowax or the chlorin removed for a period of 10 seconds the tolu ated naphthalene or chlorinated diphenyls ene predominated and we had no knowledge arc used, then that shows that there is suf- of the other being in there. . ficient concentration of these substances in the air to cause plugging of the follicles and Mn. Armiun G. Bettui (Research Engi to cause a skin condition. If there is suffi neer, United American Bosch Corporation, cient concentration to do that there may be r- r^rji!. II I >1 I--pmU**me**e4^3*+*F*****^ H ! -- WATER_PCB-SD0000030426 nil Tt IWifl'Hnii in irnSfcTu^ j unit ill vol. 13, no. 7] EFFECTS OF CHLORINATED HYDROCARBONS 305 sufficient concentration to cause systemic Mb. TCarrek A. Cook: I am not quite poisoning in the few people who are hyper sure that I have in mind exactly what you sensitive to the action of these hydrocar want in that regard. In connection with bons. Everybody is not hypersensitive to sampling? them. If they were, of the thousands of people who have been handling these sub Dr. Cecil K. Drxxksb: Yes, In con stances, more would have contracted this nection with the whole technic of determin disease and it would have been reported ing the material in air, length of sampling, very much earlier. This material was used type of apparatus, and so forth. It has in Germany as early as 1914 or 1915. been suggested that we endeavor to agree While I cannot add anything to the upon some uniform procedure for that and methods outlined here for safety in the way as figures begin to come in, as they will in of ventilation and so forth, I can say that the course of the next couple of years, we if you go into a plant and hud that there is will at least have them on a comparable any acne present among the workers with basis. this material you will know that there is sufficient of the substance in the air to cause Mr. Warrek A. Coos: la that regard acne and consequently sufficient perhaps to there is no question but what the determina be dangerous to the few people who are tions add results of analyses of the concen hypersensitive to its action. tration of injurious materials in air, corre Dr. Cecil K. Driver: There is a prac lated with pathological findings or freedom tical point in regard to the question cf deter from pathological findings, are real factors mining these substances in air. My brother in further substantiating toxic limits which pointed out this morning that there are - are originally given us from animal experi several pieces of apparatus for doing this. mentation. In order to obtain data which It has seemed desirable to a number at the are of value along that line it is desirable to men who are here that during the next cou know to what extent, in terms of milligrams ple of years, while we are in a position of per cubic meter, workers arc exposed to learn ing, that we endeavor, if we can, to in breathing any potentially injurious confine our analyses to one piece of appa material. ratus, so that at least our results will be as It seems to me that one of the things that comparable as we can make them from place should be done, which many of the plants to place. and many of the organizations equipped to I don't know much about this phase of make determinations of injurious materials the subject: perhaps, Mr. Cook, you have in air should do, is to make an effort to an idea or two about that. develop one group and series of results which will represent average exposures of Mr. Wabrek A. Coos (Chief Industrial workers to chlorinated naphthalenes. In Hygienist, Bureau of Occupational Dis getting that figure I think that it is of great eases, Slate of Connecticut Department of importance that we should consider average Health, Hartford, Conn.). On the deter daily exposure. There is a tendency very mination of the halowax I wonder if it often in taking samples of air to determine wouldn't be preferable to have Mr. Har maximum exposures, to get the exposure per!/ discuss the method he has been using over a 15 or 20 minute period when the throughout the country for the determina worker may he actually tending t pot where tion of chiorinaced naphthalenes. the material is being used, whereas during the subsequent half hour he may be 40 or Dr. Cecil K. Diumxer: I think so far 50 feet away from that particular location. as that is concerned it is best to leave that Therefore, for one thing, it seems to me it to discussions with Mr. Hempcrly and Dr. would be extremely desirable to obtain in Brandt afterward. But the question of formation in as many plants and os many determining some uniformity of leeltnic for locations as possible of the average expo a while at least is. I think, a matter of gen sure, taking into consideration not only the eral interest to us. worker's maximum exposure while actually ;5 fj ir-sessggiS&sStl aUf - ` -*"** 306 JOURNAL OF INDUSTRLAL HYGIENE AND TOXICOLOGY [Sept. 1S3T at work with these materials but also his before the A. P. H. A. in October as one of much smaller exposure and possibly even the projects which we shall carry out at lack of exposure as he may go to other parts that time. of the room or even to other departments Aa I have not given this consideration, I durian the course of the day. am afraid my discussion is not as connected .Another type of determination which ' as I might wish it to be. We have a real seems to be extremely' desirable to have is problem here and one that deserves some concentrations of the chlorinated naph study. thalenes at certain reference points. I believe that blr. Heaperly very carefully Dr. Cecil K. Drinker: Would it be of duplicated these reference points in each service to the-American Public Health Asso- of the plants where he took his determina elation Committee if a sub-committee from tions. He took the samples, I believe, at this meeting, collected perhaps by my breathing level and a certain measured dis brother and those of you involved with these tance from the pots where the chlorinated - problems, were to make a recommendation naphthalenes were being handled, so that to the A. P. H. A. aa to the things you have nexs year he could go back to those same brought up and as to suitable apparatus, locations or others could take determina and let the final statement come from the tions at those same locations, which are Association after they have considered that definitely fixed and recorded, and learn material? - whether, due perhaps to clogged pipe lines or to fans which might become loaded with Mr. Wjuiren A. Coox: I think that the condensed chlorinated naphthalene, would be a splendid idea because we have there might be poorer ventilation than origi a bulk of information on this particular nally designed and higher concentrations material. A number , of individuals have of the chloriosted naphthalene than origi given much thought to the development of nally determined. apparatus, both the combustion method It seems to me that it would be an ex which we have here and also the continuous tremely desirable thing to have made avail record type of apparatus which Professor able for all of us who are making these Drinker mentioned this morning. I think determinations these reference points so that it is a sufficiently important project that in Massachusetts and in Connecticut, that a sub-committee made up from this at one plant or at another, a considerable group could give that committee a very group of workers could be taking samples at definite steer. this one particular point with reference to (Personnel of the Sub-committee: W. A. the source of generation of the vapor. Such Cook, chairman, A. L. Coleman, II. B. El results would permit a comparison of knowl kins, W. 3. Fuiton. S. W. Gurney, E. R. edge between plants. Kayhurst, W. F. Hemperiy, F. W. Sehi, and In that regard it happens that there is a A. N. Setteriind.j committee of the American Public Health Association which is charged with the devel Mr. Hervst B. Elkins (Chemist, Divi opment of chemical methods for determina sion of Occupational Hygiene, Massachu tion of atmospheric contamination such as setts State Department of Libor and Indus- . that now under consideration. It seems to tries, Boston, Mass.): In connection with me that it might he a very desirable project what Dr. Schwartz has said, I a:n not sure for that committee to cunsult with the in that I am in complete agreement as regards vestigator of the Ilalowax Corporation and the use of skin lesions as warning agents. suggest to ail of those who are doing work It is my understanding that in insulated of this type, after some discussion, what wire plants the chlorinated diphenyls are reference points could be adopted and also very commonly compounded with the chlo what should i:o done in connection with get rinated naphthalenes. It is aiso my under ting these average exposures. As a mutter standing that these .ire more likely to cause of fact our committee will discuss that very a severe skin irritation than the straight feature and pussihiy include it iu our report chlorinated naphthalenes. On the ether TM f J-L^****C*wvmMp-vmj ^....^ , WATER_PCB-SD0000030428 ml. 13, no. 71 EFFECTS OF CHLORINATED HYDROCARBONS hand, from Dr. Drinker's work one would to attempt to get the concentrations low. Judge that the straight chlorinated naph We saw this morning that the effect of these thalene was nearly as toxic at least as the products was ou the liver and that if some mixture of it with a small amount of chlo one working in these products developed rinated diphenyl. liver damage from some other source the In the condenser factory where we hod chances of his having serious trouble was our esses there is no chlorinated diphenyl increased. Therefore, inasmuch as there is used and no acne of the type described in no test to tell whether a person has had liver the reports on the insulated wire factories. disease, certainly in the preliminary medical examination of applicants for work the his Dn. R. Emmett Kelet (Monsanto Chem tory at least should be gone into very care ical Co., St. Louis, Mo.): I can't contribute fully for a history of previous liver diseases. anything to the laboratory studies but there There are other things that should be has been o.uite a little human experimenta token into consideration also. One is the tion in the last several years, especially at history of syphilis, because syphilis does our plan ts where wc have been manufactur have an effect on the liver. If there is any ing this chlorinated diphenyl. It has been history of syphilis I don't think that that our observation that although on ena occa man should be hired. A Kahn test should sion wc did have a more or less extensive be token and I think that it wouid be a good series of skin eruptions which we were never plan to take Kahn tests on men at stated able to attribute as to cause, whether it was intervals, because they might develop impurity in the benzene we were using or to syphilis after they are hired. the chlorinated diphenyl, we have never had I think those things should be done even any systemic reactions at all in our men. though we feel that if the concentration of We have examined them very closely both these vapors in the air is kept as low as was from what laboratory tests we thought recommended this morning, the chances of might help us and from the clinical view having toxic effects are very small. point. Also from chlorinated diphenyl alone there have been no cases of systemic Mo. F. R. Kaiuer: I would like to clarify poisoning reported. a point of Dr. Hookey's. I don't believe that we can transpose the You mentioned at the beginning of your laboratory results into the actual humans talk that your experience in acne condition without paying considerable attention to was more pronounced from the compounds the volatility of different substances and the of various materials with chlorouaphthnlene way they arc being used. . than with the straight chlorenaphth&ieaca. Did you mean by that, that the acne condi Dn. John A. Hookey (Consulting Der tion actually was more pronounced with the matologist, Haiowax Corporation, Detroit, compounds than with -ho straight culoro- Mich.): I have had considerable experience naphthalene or was that due to the fact that with the dermatological angle of this prob you were in a plant operated in a different lem. The experience at the Haiowax plant fashion? In Other words, you handled your at Wyandotte definitely was that the derma numerous wire compounds in more of a tosis did increase when compounds were hatch operation. That is my understand used rather than the straight chlorinated ing from Mr. Brown. naphthalenes. Is it a fact that the straight chloronaph- From the discussion wc have heard today thaiones are less harmful it: so far as acne I think the observation Dr. Drinker made is concerned or i s it because of the handling that the chances aru that dermatological of batch compounds? - problems will disappear along with the other is probably true. However, I think that Dn. John A. Hookey: That is a question certain precautions, certain insurance meas that would be difficult to answer. I imscd ures, you might say, should be taken in my statement on the fact that in taking a h:ring men who'are going to work with general survey of these eases after I started these products, even though wo are going working with them the men in the plant said ,- ' t J,*- .t* -- V - i ificiVcvtvw:.g-r43 v. ;V"v * ? ! 'rgg&Wrl .C -*' ** l!.r ' t WATER PCB-SD0000030429 ~,irT"Ti ****yifftirtiflfct. ttoBtamA3taa44MCjaMa*taaauAM3Hji^%dv<$A4aRatne^3tefeSaA&^i*i4aatteasMMAv2tfl0V0fMa 30S JOURNAL OF INDUSTRIAL HYGIENE ANDTOXICOLGGY ISepL 1937 that they developed acne more often and vulgaris is most apt to occur. The sebace more severely alter they started making ous giaads are most active and they seem to compounds. It may be that it was in the have it much more than the older people in method of the handling. whom the sebaceous glanus are not so active. Ms. F. R. Kauant: You say that the con We keep & clipping of all articles relating dition increases with the increased chlorine to industrial dermatitis in my office and I content and we are attributing the acne pri recently read an article in a German maga- i marily to the chlorine radical. If that is sine in which they report in Germany a true, what are we to associate insofar as similar acne condition from chlorinated these various other constituents are con phenols and chlorinated benzenes and solid cerned, such as various piasticizing agents? waxes. It seems that this acne-like condi How much do we know about those insofar tion is not peculiar to halowax or chlorin as development of acne is concerned? Some ated naphthalenes or diphenyls but can of these compounds have a haif dozen con occur with any condition that blocks up the stituents in them other than chlorodiphenyl sebaceous follicles of the skin. and chioronaphfcnalene. We are at the present time considering the ehioronaph- Dr. Ror.u< Meeker (Special Agent, thalene as the basic element responsible for State of Connecticut Department-of Labor, the acne condition which we have. We Hartford, Conn.}: I am full of questions. know very little about what the other mate As a statistician I am a little bit suspicious rials may or may not do. of averages, so I should think that we should take maxima os weil as averages in getting Dr. Jobx A. Hooket: I don't know at the threshold dose or the dangerous im about these other materials. Perhaps Dr. ' pregnation of the atmosphere with these Schwartz would know something more poisonous compounds. I just wanted to about them. drop that as a hint for this commit:** to consider in its work. Dr. Louis Sciiwartz: It is well known In spite of what you said at luncheon. in industry that there are many compounds Dr. Drinker, I still think it is rather impor that cause acne. For instance, paraffin tant to determine how the plugging of the acnes and oil acr.es have been known for a pores actually does occur, whether they are long time. Paraffin acnes, wax acnes, arc plugged as Dr. Schwartz has suggested, simply a mechanical plugging of the fol from the particles falling on the outside of licles of the skin by the minute particles of the skin or whether they arc plugged, as so wax that fall cn the skin. I think that the many doctors have informed me, from inside acne caused by these chlorinated naph as it were. The fumes are breathed, the thalenes and diphenyls, because it is caused substance gets into the blood stream It by both--I was down at the Swan Chemical thrown off through the pores of the :u.u Company where they make chlorinated di the pores are plugged. I think it is rather phenyls and they have acne there just the important to determine that because it game as over at Wyandotte where they make seems to me, from the slight knowledge that the chlorinated naphthalenes--I believe I have of the industrial application of ha:..- that that is caused by the same thiug, a wax compounds in insulating blocking up of the follicles of the skin, the the plugging of the skin, if it is formation of the comcdone and then the side plugging, may take place ci:..jr ~ comedune becomes infected and forms a handling a solid substance or 'rem' :-e pustuie.. `They don't all pustulate because coating on the wire after it becomes* :md. they don't all become infected. Many of I haven't heard anybody say- anything these men never have any pustules. They about those Massachusetts cases of very just have little white elevations on their severe dermatitis and I am not so sure but skin. . what there was some question of liver dam In those people who are in the acne age, age there. Nobody has said anything about the youngsters of IS, 20, 22 and 23, acne them at all. There is no question there WATER PCB-SD0000030430 vol. 19, no. 7\ EFFECTS OF CHLORINATED HYDROCARBONS 309 that the condition, whatever the complica tions might have been, came from handling cold wire, in a confined area of course. I think this needs a great deal more study, Dr. Drinker, than you and your associates have been able to give it os yet. All I am curious about is the way we con administer the protective labor laws in order to protect both the employer and the employee. I listened with very great interest to the expositions of suggested pots for the appli cation of halowax, chlorinated naphthalene and diphenyl. It seemed to me that they left very much to be desired. I don't see why the application of the halowax com pound, if it is necessary, as I assume it is, to use this rathar dangerous substance, can't be done in a practically closed pot. If you please, Dr. Drinker, I wish you would ask Mr. Reeves of the Rockbestos Products Corporation to toil how they have handled it. If I am any judge of equipment and safety devices they have como as near to eliminating entirely from the atmosphere ail contamination from fumigated or evapo rated halowax as anyone. All of the appli cation of halowax, either in the melting pots or where it is applied to the wire, is done under hooding and with ventilating appa ratus, ar.d it seems to be entirely adequate. wax we had a head start as far as the ventilating system was concerned, since ours was already installed and adaptable to a compound with a benzene solvent rather than a me!ted halowax compound. In order to keep the fumes of a highly volatile sol vent under control we did develop a prac tically complete enclosure for the container In which the compound was placed and exhausted that container in order to prevent any escape of the fumes from it. When we changed to the halowax compounds we fol lowed the same scheme of applying the compounds and used a completely enclosed pot. There is a hole in one side for the wire to go in and one on the other side for the wire to come out. There is enough exhaust suction on the enclosure to prevent any fumes from getting out of the openings where the wire enters and leaves. Of course periodically it is necessary to open the pot to replenish the compound or make adjust ments. In that case we have the ventilat ing system interlocked with the door that the operator opens so that when he opens it he increases the amount of air travel about ten-fold. Db. Royal Meeker; What is the veloc ity when the door is open? Ma. B. H. Reeves (Vice-president and Mr. B. H. Reeves; With the door wide General Manager, Rockbestos Products open, at breathing level the velocity is 200 Corporation, New Haven, Conn.): I will he 'feet. That is the linear velocity. We very glad to tell of our experience if it will check that every week on every opening at help tiie picture along. Our experience has every machine with a velometer, a small been more or less parallel with Mr. Kauner's instrument which is easy enough to operate in as much as we manufacture the same type and reads directly. Whenever the velocity of wire. We have avoided the serious pic falls below that figure the pipe connecting ture that he ran into, probably due to the that machine is taken down ar.d cleaned, fact that shortly after 1923 we were using because there will be condensation inside benzene as a solvent for asphaltic materials. the pipe to cut down the air flow and reduce We got into anemia difficulties with ben the ventilation efficiency. zene fumes and had to install a complete ventilating system to remove all those Dr. Royal Meeker; Whet do you think fumes from the machines that applied the of that trap device shown? compounds to the wire. Following the installation of that equipment we were able Mr. B. IT. Reeves: I don't know what to to get a substitute for benzene, so we got it think of that. I don't know whether it out of the picture. would be practical or not. I was going to We are still in the same (is as every one ask Dr. Drinker whether in passing the air else is hs far as finding r. substitute for halo- laden with halowax fumes through the boxes wax. There is none that we know o.f. Con of rats there was any deposit of halowax in sequently when we went to the use of halo- the enclosure. -r^=p(fc''~5ss^- * * 1 <^5*ig'ji5f. ;'; wwsia^s i i} i? m*k$ ^4i- ?;n "_2E3!M5^S!T Z2 ;1 5u f!.: raagscgg : > i. .JaiSsa.T.!];! .* >3 ^.vr.'ya nu.:i iiiS___A___ASHKS-c?V?s i-J !*;:<'.l]'Ji1 :, . p2S 4 i?1 i* j*j i-: IflSSfesrrai i ' 4 i-: JSwPSKSSf- i i .1 i.r r-__r?__T_T_'_l'+'*"* +* I*ii! ij ?I 'i-risar^s^i f >4 \k mmm aw. ZXfZia&SLS !*;:< sg*r<>| ? tukau^r^ * i . . - jfr.Try4* iJ1Mj *T'| a?r*4 -*i * -'1 **; g3a^gS IV^NOA -,.n,Va,T&^?&..52 j: J --'Jri ; :{ \^CrT .*",> _*i a LW . ,T.** >- vf; K'.' '.+. 1 ''--;`c--;v-vi*i>V c< sV^Tw'Sf-''.v-f^r'^' J:T^,ws,f r*v*"7--" r,.*~^-*.\ VniSiM. tfvferr.i grfgtfrartia ^ rig 310 JOURNAL OF INDUSTRIAL HYGIENE AND ^TOXICOLOGY [Sept. 1S27 Dr. Cecil K. Drinker: Yes. It is detectable readily through the outflow pipe, Mr. B. H. Reeves: Those in the far end probably didn't get the concentration. Dr. Cecil XL Drutker: We change the position of the boxes each day. The ones furthest away move into the nearest posttion, so tliat over a period of weeks all ere exposed equally. complaints were bona fide. We had no diffinulty in handling cold wire and the men had no akin trouble. Yet when they did the same thing with possibly the application of heat in tunnels, ships and so forth, they actually did develop trouble. We have used, of course, the icterus index in following some carbon tetrachloride cases and I wondered what you have to say about that. Is that of no value at all in the case of liver damage in this material? ' Dr. Ejsert R. Havhtost (Consultant Dr. Cecil K. Drinker: I don't think so in Industrial Hygiene, State Department of until it gets so extreme that we know it any Health, Columbus, Ohio): I would like to way. ask if it wouldn't be possible to put a cold . water line around that trap and deposit it Dr. B. L. Vosbtoch: I did write down a all before the air escapes from the trap. few questions that came to mind. Some of ` them have been answered and some are Mr. B. H. Reeves: We have tried cool- foolish but I shall ask them anyhow, ing the surface of the exhaust pipes in order It occurred to me that if trichiornaph- to concentrate the deposit in one ptace and thalene has been proved to be practically make it unnecessary to remove long lengths innocuous, isn't it a problem of research of the ventilating system. While we have largely for the Halowax Corporation to use found a slight advantage in doing so it only that one material adopted with other , hasn't at all paid for the expense. It is known chlorinated compounds to meet the much easier to take the piping down. Con- requirements of customers? That is one sequently I doubt the possibility of remov- of the foolish ones probably, but I shall ing it ail by cooling the surfaces of the trap, just ask it. It might be possible, by building a labyrinth Furthermore, how long is it reasonable to of refrigerating coils, to collect it. But expect the liver to recover after it has been then you would have the problem of remov- pretty well damaged. I talked to you and ing it from the coils after you had collected you said you hadn't worked that out with . it and it would involve rather complicated the animals as yet. ' procedure. Dn. Cecil K. Drinker: In that case we _ Dn. B. L. VosBintCH (Medical Director, set aside a certain number of animals to ` General Electric Company, Schenectady, recover, after knowing that a certain New York): We have heard about the pos- amount of damage had been done. Then sibilitv of preventing liver damage and skin we started to kill them but we invariably trouble among manufacturers of wires and ran out of rats before they ran out of lesions, cables end so forth, but like the old rhyme, Wc don't kuow how tong it lasts, every dog has fleas and the fleas have fleas, wc also have customers who use wires and Dn. B. I,. VosnoRcu: Have you given cables in tunnels, in enclosed spaces, splic- them calcium and glucose as you would with ing them together and doing all sorts of carbon tetrachloride? things. I am not at all certain that wc know what the concentration of these chlorinated Dit. Cecil K. Drinker: Not yet, but we hydrocarbons is under those conditions. are doing an experiment now with one grou p About the time we wore having so much on a very high calcium diet and another trouble at our York factory some of our group on a calcium free diet. The whole customers began complaining. Wc thought question of calcium in the diet, of giving we were having a hysteria of halowax mania milk daily to these people may be worth throughout the country. Some of their while. , WATER PCB-SD0000030432 *) vof. 10, no. 7j EFFECTS OF CHLORINATED HYDROCARBONS 311 Dr. B. L. Vosbgrgh: I just had those questions in mind. I would like to hear wiiat Mr. Brown would say as to the possi bility of using only this one innocuous chloriaated naphthalene. Mr. Saxdford Browx: That would simplify our problem exceedingly, not only from the commercial standpoint but from Use toxicity standpoint. But if we did so I think that Mr. Kaimer would be the first to object because he wouldn't be able to get the technical effects he desires in the cable. The same thing applies throughout all the industry where we are using these higher chlorinated products. The problem as we see it, and it is one which we intend to study further (we have our program for this continuous work that will be done under the direction of the Drinker brothers), is to try to find a line of demarcation and whether or not within this complicated chemical structure there is some one thing that causes the trouble. If we could take hcxachioraaphthalene and remove l per cent of some unknown constit uent with which we are not now familiar and eliminate the toxicity, that would be a lovely solution. Whether we will ever be able to attain that l don't know. That is one of the things we have in mind. As I said before, that is one of the things with which all manufacturers of these syn thetic organic products are confronted at somu stage, ei thcr in their laboratories or in commercial developments. It is largely a question of timing as to when you shoulddo the work, how much you should do, and whether you know how to do it. You have to learn that first. Dir. Cecil. K. Drixker: Unless there is some other important matter I will dcciare the meeting adjourned. Du. R. R. Sateius: May I say a word more with regard to what Dr. Ilookey stated on syphilis? Tf you are going to eliminate al! your workers who happen to develop syphilis during the time they are working for you, you are probably going to lose some very important employees. It would seem that it would be worth while to give serious consideration to seeing that conditions are proper and suitable for them to continue working for you and that they have proper treatment. In other words, I don't believe that is a good or sufficient reason for dis charging those employees who do develop syphilis. We have not found that to be true in other places and I question whether this is one of the places. I may be in error. Mr. Sandford Browx: I want to take this occasion to express on behalf of the Halowax Corporation my thanks for the excellent cooperation we have received from various state authorities and our customers as well in this investigation. I believe you have all formed the opinion, based on what you have heard this morning, that our whole motive here is to conduce this on an ethical, scientific and constructive basis. In collaboration with the Monsanto Chemical Company we have a much more comprehensive program in view to carry on. Therefore we want to continue that same type of cooperation. There is one thing that I want to bring up in that connection, which wil! have to be observed by the state authorities, when they go out to make these inspections. That is the necessity of not creating mob hysteria on the part of the workmen in the plants where these inspections arc made. Mr. Hemperiy has run into some very interest ing situations in the various examinations he Juts made. I know that he would be glad to relate them and give the benefit of his experience to the heads of any of these state departments who will have the direction of this work. Otherwise I can sea where we will be unable to get the full cooperation of not only the individual workmen but the plant foramen and the management,, and we must have that if we arc to gat die, results that wo are shooting for over the next sev eral months or years. This thing may con tinue, probably wil! continue for years. A5>'.*we'f>c2U+ tr'Ki'Z^Fz. * \f -iS *-vr* > < ; 7- SA'iJjwfe.fcSSt; -A ..fi j fU-ri. i * it. 1 * . I .5.' . 3j~X,S'.-TiA,*sr 1 a) 5 >. 5 A. iv li!.iv' * Ivi;Z: t .&,M $ i -a * |V * )V Sbr.r/**.#ewv\.r-r. V* ! w">. > C- > **i : WATER PCB-SD0000030433 Repriated froai The Journal or Iototsial Htgieke and Toxicology XqL 20, No. 3, February. 1333 l l iii i MORPHOLOGICAL CHANGES IN THE LIVERS OF RATS RESULTING FROM EXPOSURE TO CERTAIN CHLORINATED HYDROCARBONS* Granville A. Bennett, Cecil K. Drinker, and Madeleine Field Warren Department of Pathology, Harvard Medical School, and the Department of Physiology, Harvard School of Public Health, Boston, Massachusetts HLORiNATED hydrocarbons, particularly chlorinated naph thalenes and chlorinated di phenyl, have been used extensivelyin certain industries. - Their use in the manufacture and preparation of many types of electrical equipment is con stantly increasing. Although it is known that some of these compounds cause acne, only recently has the possibility of more serious systemic effects been recognized. During the spring of 1936 we were informed of the occurrence of three fatal cases of jaundice in workmen using chlorinated naphthalenes and chlorinated diphenyl. At the request of the manufacturers of these com pounds, wo undertook an investigation Received for uublication September 5, 1S37. ' This is the second of three papers read at a symposium oil chlorinated hydrocar bons siven at the Harvard .School of Public Health. Boston. June 30, 1937. The third paper will appear shortly. to determine what systemic effects, if any, would result from the adminis tration of a number of these com pounds to experimental animals. The known findings in the three fatal cases of jaundice, together with a review of the pertinent literature have already been reported (1). The present paper describes the pathological changes observed in rat3 that had been exposed to various chlorinated naphthalene compounds and to chlorinated diphenyl. Materials and Methods A detailed description of the ap paratus and technic employed by the authors in this investigation has been published (1). White rats, main tained on a diet of Purina Dog Chow, supplemented by lettuce, eggs, milk, and cod liver oil, were used throughout the experiments. DEPOSITION EXHIBIT | o 3 A - /ST -9</ ! St ! j 1 > ! ( 1r \ s / WATER PCB-SD0000030434 98 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [roi. SO, no. 2 The chlorinated hydrocarbons tested were the following: Compound A. A mixture of tri- and tetrachlomaphthalenes.* Chlorine content 49.4 per cent. Compound B. A mixture of terra- and pentachlomaphtha- lenes. Chlorine content 56.4 per cent. Compound C. A mixture of tetra- and pentachlornaphtha- lencs plus chlorinated diphenyl. * Chlorine content 43.5 per ccnt.f Compound D. A mixture of penta- and hexachlomaphtka- lenes. Chlorine content 62.6 per cent. Compound E. A mixture of penta- and hcxachlornaphtha- lenes. Chlorine content 62.6 per cent. N^Ui-UpVJUUU i* . u- vwUliAi-Ui Wi per cent penta- and hexa- chlomaphthalenes, plus 10 per cent chlorinated diphenyl. Chlorine content 63 per cent. ^ "Compound G. Chlorinated di phenyl. Chlorine content 65.0 per cent. These preparations were selected because of their relative importance in industry. It should be noted that they also represent a wide range of chlorination. AH of the above materials were administered orally in varying doses. Compounds A, D, F, and G were selected for the inhalation experiments. This compound contained only small amounts of tetrachlomaphthalenes. In this paper it will therefore Sc referred to as tricaiornaphthalcnes. t This compound consisted of a mixture of compounds B end G but in audition con tained two plasticizers which have been consiocred to be inert. Without these materials the cniorine content of this com pound would be between that of compounds B and D. Inhalation Experiments "We were most interested in the results of the iuhnlatiou experiments because they more nearly simulate the type of exposure to which the workmen are subjected (1). The in halation experiments were carried out in air-tight wooden compartments through which air, containing the volatilized compound being tested, was driven by electrical blowers. With this apparatus we were able to test simultaneously the effect of four compounds. Eighty rats were ex posed to each compound. The meth ods for determining the daily air concentration (mgms. per cu. m.) of the chlorinated hydrocarbon being used, as well as the rate of airflow have been described (1). Compound A (trichlornaphthalenes) was administered by inhalation to two groups of animals. In the first of these experiments, the rats were ex posed to low air concentrations (aver age 1.31 mams, per cu. m.) 16 hours daily for 134 days. In the second experiment the concentration was increased to an average of 10.97 mgms. per cu. m. This 1G hour daily exposure was continued for 102 days. Compound D (hexa- and pentaehlcrnaphthalcnes) was administered to three groups of rats. In the first experiment the exposure consisted of 16 hours daily for 134 days, with an average air concentration of 1.16 mgms. per cu. m. In a second experi ment the average air concentration was 1.44 mgms. per cu. m. This was maintained S hours daily for 143 days. In the final experiment, the average air concentration was in creased to 8.SS mgms. per cu. rn. and exposure, 16 hours daily, was con tinued over a period of 52 days. WATER PCB-SD0000030435 .0 * the nents late the \o in- A out ents , the ested, rers. ,C tO f four ex- icthly air - m.) jeing ^raow tnes) two nrst of :e ex'aver- G hours second was i 10.97 nr daily ; days. ..uchlorcrcd to : e first sted of with an i 1.16 experititration This i for 143 .- nt-, the was in- m. and as con- i \ ; i | ; ; ; : ! j ; j i j ; j j 1 i i ; , ' . i ! | ; < ! I ! ' ; ; i ! J i ; | t i Feb., 10SS] CHLORINATED HYDROCARBONS 99 Compound F (hexa- and pcntachlornaphthalcnes 90 per cent plus chlorinated diphenyl 10 per cent) was administered in low concentrations to two groups of rats. In the first experiment an average air concentra tion of 1.37 mgms. per cu. m. was maintained 10 hours daily for 134 days. In the second experiment, an average air concentration of 1.60 mgms. per cu. m. was inhaled S hours daily for 143 days. Compound G (chlorinated diphenyl) was administered to two groups of animals in low concentrations. An average concentration of 0.57 mgms. per cu. m. was employed 16 hours daily for 134 days in the first experi ment. In the second experiment (em ploying an average air concentration of 0.93 mgms. per cu. m.) the animals were exposed 8 hours daily for 143 days. Feeding experiments All compounds tested were employed in _ .the feeding experiments. A weighed amount of finely ground material was mixed with rat food. This mixture was placed daily in one food container to supply the rats in a given cage. Thus each animal had an equal opportunity to ingest the compound supplied. As the number of rats per cage was reduced, the dosage supplied was proportionately decreased. Most compounds were supplied in large and small amounts. Supplementary experiments were conducted on a smaller number of rats by feeding several of these com pounds in small known amounts by stomach tube or by injecting them subcutaneously as suspensions in gum acacia. Carbon tetrachloride and alcohol administration to experi mental animals Seemingly the incidence of acute yellow atrophy in workers exposed to chlorinated naphthalenes and chlo rinated diphenyl is very low. This single fact is of some importance in that it suggests that certain indi viduals may be more susceptible to the compounds or that in these in stances the liver damage may have been intensified by some other agent. With this in mind, groups of animals that had been exposed for varying periods of time to certain chlorinated naphthalene compounds and chlorin ated diphenyl were subsequently given a subiuthal dose* of carbon tetrachloride and ethyl alcohol by stomach tube (0.75 cc. of each per kgm.). This dosage did not result in a single ricaiii among liio 1G coultol animals and the degree of liver damage produced was quite constant and never very great. Administration of carbon tetrachloride and alcohol to rats exposed to the more highly chlorin ated naphthalene compounds and chlorinated diphenyl caused extensive liver damage and proved widely fatal. Therefore, the use of carbon tetra chloride and alcohol has been employed regularly on representative groups of rats from each inhalation experiment. Examination of tissues Animals were sacrificed for patho logical examination after varying peri ods of exposure. Complete autopsies were performed. Liver weights were recorded. In representative animals * One cubic centimeter of carbon tetra chloride plus 1.0 cc. ethyl nkonol per kilo gram when administered orally to normal white rats causes a U per cent mortality (2). *i j i l I ii i i i i i i i WATER_PCB-SD0000030436 100 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [vol. SO, no. 3 from each experiment, all organs excepting the central nervous system were examined microscopically. The livers of all animals were studied microscopically. Tissues were fixed in Zenker's fluid, 10 per cent formalde hyde solution, and absolute alcohol. The routine paraffin sections were stained witii eosin methyiene blue. Other stains employed were hematoxy lin and cosin, phosphotungsric acid hematoxylin (Mallory), Foot's modifi cation of Bielsehowsky's stain (reticu lum), Best's carmine stain, and Iron reaction with fcrrocyanide of potas sium (Mallory). In addition, frozen sections from certain liver specimens were stained with scarlet- red, Lugol's solution, and methyl violet. Results Compound A. Trichlornaphihalcnes (a) Exposure by inhalation to loxo concentrations (average 1.31 mgms. per cxi. mi.) 1C hours daily.--The SO rats subjected to these conditions showed no ill effects. Autopsies were per formed on groups of 3 to 8 animals after 37, 72, 105 and 134 days' exposure. There were no significant abnormalities in liver weights.* Macroscopically the majority of the livers appeared normal although an occa sional one was paler than normal. Rarely slight mottling was observed. Microscopically, the liver cells often appeared slightly enlarged, move gran ular than normal, and occasionally they were vacuolated. These slight abnormalities and the presence of a rare mitotic figure suggested that very slight injury' to the liver cells had occurred. There was no demon strable increase in the above changns after the first exposure period (37 days). Occasional livers of rats ex posed fnr the total period showed * Calculations pertaining to the weights of livers in these and subsequent animate are based on data in II. H. Donaldson's book "The Rat." 2nd ed. The Wistsr Institute, Philadelphia, 1924 (p. 211). PLATE I Fics. 1 axd 2. Camera iucida drawings showing the portal areas of the livers of two rats that were fed large doses of a mixture of tetra- and peiuachit.rnaph- thalenes and chlorinated diphenyi. The changes illustrated in figure 1 occurred within 4S days after exposure was begun. One should note the marked accumulations of hyaline globules in the cell cytoplasm, and the marked swelling of liver cells. In creased numbers of mitotic figures were also present. Similar changes are apparent in figure 2 which was made from the liver of a rat exposed 125 days. Degeneration in the central portions of the liver lobules is also present. Fics. 3 and 4. These drawings illustrate the marked swelling and fatty vacuolization of liver cells observed in all rats that were fed small doses of tetra- and pentachlornaphthalene?.. The changes shown in fig ure 3 were present after 2d days' exposure, those in figure 4 after 48 days' exposure. Note the mitotic figures in figure 3. The compound responsible for the changes con tained no chlorinated diphenyi. Otherwise it was similar to the compound responsible for the lesions illustrated in figures l and 2. Fins. 5 axd 0. Camera Iucida drawings of the portal areas of the livers of two rats fed moderate sized daily doses of a mixture of penta- and kexachiurnanhthalenes for 14 ana 29 days respectively. In the first of these figures one notes swelling of liver cells with narrowing and distortion of the sinu soids. The liver cells show an incieased granularity of the cytoplasm with a massing of basophilic granules near the nuclei. There is'also an excess of fat in smail and medium sized vacuoles. This was more prominent in the central portions of the liver lobules. An increased spacing be tween liver cells and between liver eel's and sinusoidal endothelium is present. After 29 days' exposure (fig. 6) the above changes are greatly increased. Numerous large intercellular spaces containing serous precipitate, strands of fibrin and leucocytes had developed between liver cells. These spaces were bounded by distorted liver cells. See also plate III, fig. 1. Camera Iucida drawings X 395. Ci r l \ r F k i. t V" i: r i ! t O V WATER PCB-SD0000030437 lls ac,rg Ull. ed. n _u- illy it a ery ' -,d i- iges '37 x- wOd "Uts >3 sioie nd 2. Ags ai3 ure 'or 1-4 st of ells nused issing uclei. ind sore the l becells snt. -ove ,'OUS serous ocytes hose ivcr PLATE I S' _ --v-r.w ' ^ . -i ' fv . t* - .--i ,f,'-->w*.t/ S' *.*7*.. v; i. - . -rm. _ 1 .X J fO/ *`i.-. / ' V** ' r **O>i' > {/ rJ -- * - O \9 - * oj <tto a. *<.0 . V3- - y* . .f* - -:T-1 v? * i. .. . i . -7 ' . V~'\ V >- ,.* .*ao .. '.--V' b* -. ^ ?'] - Av- .- .- X . >Ky .* V. r'r' T-> " 7* ^'"* w.-'n- .- ^'rV*0? /v.T.* y\*/- 4:** - ?>; .4^ ? ` - . rf-. V* ".*:Vi ^; " ..' . =.*.' J M. **'4 -,,'foVV^ \ <a , :... /IrS; * < .-.i *7 rrn` V* * `T//' 7 *';i * "- '-V A* ty V - ft-v; : .T'cr cc '>, _ -J ".r5i , .v I}- 1? % d yS .** ^ y'/ **> s`*#v ^-V/u*v,v : * PTJ y v-V/ ,;7 -/4 '2>- J (As t . >--i 77-, '/ .; . * ' : ' ` - ^ 50.*s* *<9 . . ?co.*i' ,";'* * .' tJ . J.'.M7-V- ,,< I' - - T v . \%a* * a.:W*>-r*' dn v *> -*5>J *2 . ^ `N >* -M-* ^ * 'V.. i.-.' ,v ;7-^."k :7.- , v e**.- AV -V. ^ ?' C s V . _ . >1 jr....,.'. ;v >" -."V** ..*> . V . * .; 7'.v74 ;/ 7 ;7 i ' '7 7 V . . ... r-- \ - & ^ - `ar-'.-T.-.->* Vr,' <* \ - ^. j-7fSv>'"A >V'-. ' V* J-">. L-t -H :- ... A- 7 *j.- * -. v . ^ , v : fa C ' si- t'r ?^rv*;ifv. ;* i i'.' -*V ...... ' fU*- '*. .^ * i 7, 5.-' V ? .; r- - /' 7-!'T; -v y' N.'\ fi 5 .,..2.^ Zc * - - ..._; ;' " -r- t*~ - 7 '. . - . -r *.* cr^7. - ;. JX V.* 1- ~ v _ ... - t *r O C -rr t :y4r.pt'(*..7'. : /' *.. Vv Ov: - *:'>. 'V.v. \'f .. *. . .. - ^ - 'i/T-iwl: ^r.. <0^ fI \ll- t l WATER_PCB-SD0000030438 102 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY pad. 20. no. 2 no tlefmite microscopic change. (See fig. 3, plate II.) All other organs were normal.* Further evidence that the liver tissue injury had been very slight was obtained from the carbon tetra chloride and alcohol test. Of 10 rats exposed for 144 days in the above maimer and then fed 0.75 cc. per kgsn. each of carbon tetrachloride and ethyl alcohol bv stomach tube, none died. At autopsy and on mi croscopic examination there was little or no evidence of liver injury greater than that produced by the same dose of carbon tetrachloride and alcohol in normal control rats. (See plate IV, figs. 1 and 2, and plate V, fig. 1.! The livers of S animals exposed to trichlornaphthalenes for 105 days and autopsied after a 2 month recovery period showed no definite pathological change. This is further evidence that this compound is only slightly toxic. These findings represent a marked In ai/DP of the experiments to follow were significant chances found in any organ other than the liver. contrast to the persistent hepatic lesions resulting from exposure to the compounds of higher chlorination. (b) Exposure by inhalation to high concentration (10.07 logins, per cu. in.) 10 hours daily.--In this experiment, the concentration of trichlomaphthaIrnes was approximately eight times that previously employed. Exposure was continued for 102 days. The 50 animals subjected to this concentra tion appeared normal. Groups of 3 to 17 rats were sacrificed after expo sure periods of 31, 4$, 50, 72 and 102 days. Most of the liver.- were pale yellow in color and .-lightly mottled. There was, however, no significant variation in liver weight. Micro scopic examination n-vealed ,-wollen liver e'dls with slightly increased granularity and vacuolization of the cytoplasm. Occasional degenerating and regenerating cells were observed. These change:-, which were slightly more marked in the central portion of the liver lobules, were present after the first mouth of expo.-ure. They PLATE H Fi<;s. 1. 2. axi) 3. Liver chances resultinc fr(u the administration of trieidornaphthuieaes are illustrated in thc-e camera Uteida drawings. in fiettres l and 2 are shown the most marked chances observed after prolonged flno and 130 days respec tively) fcedinc in daily doses of 3 gni. per 10 rats. The chances observed consisted of slicht to moderate swelling of liver cells, increased crauuiarity of the cytoplasm and finally, after prolonged exposure, fatty vacuolization of the majority of cells and complete fatty t.iecuieration of occasional cells. Ficure 3 is a drawime from a section of the liver of a rat expose..) by inhalation to low concentrations of trichlornaphthalenes for 100 days. No strikinc or constant changes were observed jn rats .> treated. Occasional cells are slishtlv swollen and show increased eranuicrity of the cyto plasm. A rare mitotic ficure was observed in occasional sections. Figs. 4, 5. and 0. Microscopic changes observed following the administration of a mixture of penta- and hexaehlornnphrbaleues arc illustrated in these three drawincs. Ficure 4 shows tk" dianecs observed after feediuc this preparation in 3 cm. daily doses per in ruts, for a period of 30 days. The liver cells are markedly swollen and vacuolated. There is also necrosis and deceneration of scattered celts, l.aree intercellular spaces are oli-erved. i.-ee plat" VI. fic.- 1 and 21. I .ess markon but similar eimnc"s are observed in tie; livers of rats fed smaller do.-es tbi- preparation 'see fic. >). {2ti days' exposure.) Ficure 0 illustrates tin- liver ceil ~t atict-s that oc curred after inhalation <*f low eoneentrations. of penta- am; hoxaehlornuplohulcnes for 75 days. Note the swelling of the liver cells, the increased vacuolization and pres ence of larcc numbers -A hyaline eiobules within ceils. Mitotic r.cures were fre quently observed. Camera lucida arawinzs V 395. WATER PCB-SD0000030439 ) F'patic - to the? ttt high ch. in.) | meat, ..phtlint times >osure 'he 50 lccutra of 3 expo und 102 VP pulp ntled. .uificant Micro.vollen reused i of the r vutins: ! vecl slightly 'ortion t after . TIipv ..nil of a natiiitbi/'c (iriuv- `isorveri 3 cm. or 30 :v swollen m-iTosi. 1 I.aree I !- (See iitf'i but tli livers urn ration "luure 0 nat oe- 'iiieutra^iitli;il:nes |t(ie liver j nl pres1 rlobuies "ere fl'Q- PLATE II tz&rr??'?- & "Jr*** ~.4f . :-r\c~ f0f-M *%;*T- : * .>* "T ,w v . v. * . * V * " ' `MJkb As'/Oi * - .r,--> 5 4*.<* -- , : /:;/-* vr>.-. * *' -y J *: ^3**, g^--. ~ * v-y.4^*^ :: .vfi*<. -V* ' ' J >. * A v. > ; `A.* .i- 4 C&. `i 4 ,:Y- ; , :< ` *!* f* -. ' ` 'K V -.J J(1` -Vf `'Cx* *A \ 1 - Y .> .-- ;v.t-r. ? s--` Rfr Sf.- i&y ' V--.* ..-e^" - .. s* ^1$ aV** - f .. y:v.' . * *. . `fi. e ." . .-.3. . i .-TV-. . ^ --^V- < * ^ .' ,, ^ _ V-2 g ^ .--V 2 % .*%.. S V ^ >.v :* .1 :-rr: C'"-" -;r l/*# C9 .> . ..I , . . ^ "'V-l . * . * - ,. o-. &. ^ ' ' -5'.V ^S-;V V. >"f3/- ' ' *C* S-. .? ; : ;.i 4 * % Ii AA _ ..vVV'-; i.--. '* r..* *x:.7v 9 . y- p'l-?/ -v * -: ^-*s tv "i <, t.-. vf _ v ^ v`- -A- . r 3 .yv * ~ < K 'i s "* \ rS-; --; *v y? .. -TV 4 y: *. : " v *f% ' ' . : .i /> n- -.. :^ : ' -%.* 5 Y ' A- ~.M . :/;r. l ,,c. ' f %-*< . v *. ,, ?> r . \ .''`V * -* *4 <Lv -u:'' *:, ,> v-> ` r*' - *C" ' :y. * ** . . -^ v0 >v - J' ;- . -r * ' - -;.. *.: ib-\ . --i. .y-* . . _ . .* -v --/ . s57 v,*. * r .' ._.'V'-' ' L;f. A V. ' V/V* y . :j'4. fh* 4 }- fa SA : WATER_PCB-SD0000030440 104 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY M- 20, no. 2 were somewhat increased with longer exposure. After 102 days the liver cells were more markedly swollen and contained large ami small fat vacuoles. Small hyaline droplets in the cell cytoplasm were occasionally observed. Mitotic figures, although never numer ous, were increased in number. N'o structural changes had occurred in the liver lobules. Carbon tetrachloride and ethyl al cohol were administered by stomach tube to 9 vats kept under these condi tions. At autopsy their livers were slightly to moderately enlarged. They were yellow in color and the majority of them were mottled. The micro scopic findings were variable. In one animal the observed necrosis was no greater than thar seen after the administration of a similar dose of carbon tetrachloride and alcohol to normal control rats. The liver-- of 4 animals showed increased fatty de generation ill the central portions of the lobules with small necrotic foci. In the 4 remaining animals extensive centrai.necrosis had occurred. In the most marked of these (fig. 2, plate V), the central one-half or two-thirds of each liver lolmie was necrotic and in occasional areas two or more adjacent lobules had undergone degeneration. Hemorrhage and extensive leucocytic infiltration had occurred in these necrotic areas. The less damaged liver cells at the periphery of the. lobules showed large numbers of mitotic figures, indicating accelerated regenerative activity. (e) Exposure by feeding.--Nine of the 10 rats fed r.riehlornaphrhalen.*s in do.-es of 3 gm. per day per 10 rats were sacrificed at varying periods of time from 9 to 136 days. The tenth rat died of a respiratory infection after 182 days' exposure. There was no .'iguifieaut abnormality in the weights of the livers of these animals. After 2 months' expo.-mv, microscopic exam ination of riie livers showed slight swelling of liver cells. This was accompanied by increased vacuoliza tion of the cytoplasm due to the accumulation of abnormally large PLATE II Figs. 1, 2. a.vd 3. Liver changes resultin': from the administration. by feeding and inhalation, of a mixture of penta- and hexai-hlorimphihalcne.- ; and chlor inated diphenyl (lt)f'j) are illustrated in these drawings. Figure I is a drawing made from a section stained with F.T.A.H. In addition to other chances it demon strates the presence of larsre and small intercellular spaces, interpreted as dilated bile capillaries. These spaces contained serous precipitate, fibrin, and occasional leucocytes. Note the sharply outlined boundaries of the spaces and the manner in which these spsees ramify between cells. This rat was exposed by the feeding of lar.ge daily doses for 12 days. .Similar but less marked changes resulted from the feeding of small doses over a period of 23 days (see fiic. 2). After 75 days' exposure by inhalation to low concentrations of this preparation, marked liver changes. con sisting of cellular swelling, vacuolization, and hyalinization were apparent (fig. 3/- Fxg.s. 4. 5. A..\\o 6. The microscopic chances resulting from expo-ure to chlor inated diphenyl arc illustrated in these figures. Figure 4 was made from the liver of a rat fed large daily doses for only tj days. Marked swelling of i-eHs and rapid regenerative activity are appur<-nt. Figure 5 illustrates tin- changes tiia: were uni formly produce.i by the feeding of small doses of this preparation. Note the ex tremely numerous hyaline inclusions in liver cells. This rat was exposed for 29 days. Very similar chalices were prodiu _-d by the administration of this compound hv inhalation methods (fig. ti). The rat from which this drawing was made had been exposed to low concentrations for a period of 11)7 days. Fig. i. Camera iuetda drawing X 95. Figs. 2 to 6. Camera lucida drawings X 395. WATER PCB-SD0000030441 V), of jU in acent ion. ytic these li 8<?tl ) the ns of -rated ie oi \s iu w rats ods of :*nth iftor as no I'sthts ! or dight was uolizato tho j large (fig- scopic :n rlilorin these ; : liver I only 0 -i. rapid Kisrurr `re uni- v 'mail | iie ex .*>ns in '1 for 29 rodured I Kiel hv i;.t from --.I lieen a period | X 95. ' -swings j1 PLATE III . *.; ' /.' - tr---- T.\ti *?sf*v* I: *& -o <>.- ?/;*? *./ v -* \ s'-i 'C - <f - .,r . r'* ' r . `ii f.V*vV ."< ,.* 1 t.!`-Or - - ^ . , "sV V.<X-;... r.iy'S'X J/ :. ., ;*/-* * >/ ^ A- 'sy*f*+l S' 'V' . .A>. V- - *** /*- ' ?' '-il 4.- ---- .V-X*! - . .** ^ r*:::- ffj > St X) * //. ., - -y . s ov.-/>'T/cir ./* " i/ l- AT' K" '-*`s v' }iV ^ AA `A'Ac ~S 1- yv V Ft.a*vTCA-'^';v^.?-i-'A.>iv/r..,* -T?v*eJt>'-V-`1T/.*/.*/^O,*.*i >v- -; 'vryc^'W V'Vvj ^- ": rXj^A -'T.y*SfV*,* ''-:r<,. ; }.rS<^ '/r * \ ' ** '*A --- . -. v - .,. , ; S-' vv*.f*<rr#r v.'s^.--r*V./%. .'-"f'<sy-.v* - .(*, .. -./<i0</ X S> < .1 7r- ^ '-V ti" " ' y. '<!> -v>i -v 'r-or <*u v :: ;:;-7 `S'-uiAXSJ: dv` U.` sA^V.'-b 5 >r- ' 5x.`>>d,-c?. - '~r- $ - /v v**# <; * i. A:C .-.v.r /.-: --- ?r ` : ?- A, *-'""'s- ir r.:s#^ L-':- *'/.. -- - : f *> .A - ' '<*. ^ * li-' ^iC-*.*** ' .;^D-'_vv,'-; --A3 `Va. v-*A' ~f' ,5" `i-v f* > V*r . * ' * J' r.%: '&} *-<: t I I i i *1 i1 t i [ f WATER_PCB-SD0000030442 10G JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [ml. 20, no. 2 amounts of fat. These changes in creased .-tightly with longer exposure. (See plate II, figs. 1 and 2.) There were no architectural changes in the liver. Because the effects resulting from large doses of triHiloniaphthalenes were slight, this compound was not fed in small amount's Compound B. Tetra- and pentachlornaphthalenes were present in increased numbers, indicating accelerated regenerative ac tivity. The microscopic changes re sulting from the feeding of tetra- and pentachlornaphthalencs are illustrated in plate I, figures 3 and 4. They are those of diffuse fatty infiltration and fatty degeneration. Compound C. Tetra- and Pe'nlachlornaphthoknes pins Chlorinated Diphenyl Exposure by feeding.--This prepara tion was employed only in feeding experiments. The daily food ration for 10 rats contained 0.5 gm. of this compound. All animals fell ill and either died or were sacrificed bv the sixty-third day. The livers were pale yellow in color, friable, and occasionally showed mot tling. There were no significant varia tions in weights. On microscopic examination the same changes were observed in all rats. Those consisted of moderate to marked swelling and rounding of the liver cells. The sinusoids were markedly narrowed. The majority of tlu* liver cells con tained large numbers of small fat vacuoles. This findina was most marked and first observed in the central portions of the lobules. In the least altered cells there was promi nent massing of basophilic granules near the nuclei. Occasional necrotic colls wore observed. Mitotic figures Exposure by fa-ding.--Feeding (3 gm. doses daily per 10 rats) was continued with but one 4 day inter ruption for 130 days. Ten rats were so treated. Seven rats were sacrificed at varying period* of 47 to 124 days. Three rats died after exposure periods of 122 to 130 days. In all animals the livers were enlarged 133 to 00 per cent). The average weight increase was 71 per cent. They were also friable, pale yellow in color, and somewhat mottled. On micra-copic examination, constant changes were observed.' Practically every liver cell was swollen and rounded. Their cytoplasm, contained large numbers of hyaline bodies. These were circular or oval in shape and varied in size from about half the size of a red blood corpuscle to twice the size of the nucleus of a liver cell. (See figs. 1 and 2, plate I.) In numer ous instances, many small hyaline bodies had fused, forming large circular masses as large or larger than a normal PLATE IV Fig. J. Photomicrograph X 45 illustrat ing the extent of the liver damage produced in 48 hours by 0.75 cc. per kxm. each of carbon tetrachloride and ethyl alcohol when administered by stomach tube to a normal control rat. The degenerative chances were limited to the central portion of the lobules. Fig. 2. Photomicrograph X 220 of a section of liver tissue from a rat exposed by inhalation methods to trichiornapiithaler.es in iow concentration for loll days. Foil cov ing this exposure u.75 cc. per kgm. each of carbon tetrachloride ami ethyl alcohol was administered by stomach tube. The animal survived and was sacrificed 17 days later. No significant liver changes are demonstrable. WATER_PCB-SD0000030443 PLATE IV WATER_PCB-SD0000030444 10S JOURNAL OK INDUSTRIAL HYGIENE AND TOXICOLOGY [rot. 20, no. 2 liver roll.- These bodies stained bril liantly with oosin dve. They wore often laminated and occasionally con tained small clear fat vacuoles in their central portions. Mitotic figures in liver cells were sufficiently numerous to indicate an increased rate of re generation (fig. 1, plate I'. In the livers of 2 rats exposed for 124 and 125 days respectively, there were, in addition to the above changes, large areas of complete liver cell degenera tion (fig. 2, plate I). For the most part this was limitpd to the central half or third of the liver lobules. Occasionally, however, there was com plete degeneration of all liver cells except for a narrow zone of cells around the portal areas. In such livers there was a heavy polymorphonuclear and mononuclear inflammatory cell infil tration in the necrotic areas where liver cells were being removed. The most conspicuous feature of the microscopic changes in the livers of these rats was the presence, of large numbers of circular hyaline droplets in the cytoplasm of the liver cells. Tins'material did not stain in a manner characteristic of amyloid and its staining properties were not like those of the hyaline observed in alcoholic cirrhosis. In frozen sections stained with Scharlaeh R, the staining reac tion of these bodies was inconstant. The larger vacuolated hyaline bodies contained small droplets of fat while the majority of the smaller droplets did not stain red. Although similar hyaline droplets were ohsorved in livers of rats exposed to various chlori nated naphthalenes, this type of degeneration occurred much earlier and to a much more marked degree in those rats that were exposed to preparations containing chlorinated diphenyl (figs. 1 and 2, plate I) or to chlorinated diphenyl alone (figs. 4, 5, and 6, plate III). Compound D. Penta- and hexachloraaphthalvncs (a) Exposure by inhalation to low concentrations (arcrayc 1.10 mows, per c.u. in.) 10 hours daily -- Kiahty rats living under these conditions appeared normal throughout thoex(>osurc period (134 days). Representative animals were sacrificed in groups of 3 to 15 animals after varying ex)x.uro periods as described in tilt* experiments em ploying tricbiomaplitlialeues. There were no significant alterations in liver weights. Macroseopicaily the major ity of the livers were light v'dlow in color and slightly mottled. After the initial exposure period of 37 days there wits evidence of slight injuvy to PLATE V Fic. 1. A photomicrograph X 120 show ing small areas of necrosis in central areas of the liver lobules. This rat had been exposed by inhalation tn low concentrations of triehloniaphthnlenes for 134 davs follow ing which carbon tetrachloride and ethyl alcohol were administered by stomach tube in doses of n.75 re. each per fcgm. The rat was sacrificed 5 days later. The extent of necrosis is similar to that resulting from carbon tetrachloride and alcohol alone. Fio. 2. In this pnotoinierograph X 45 are shown the most marked liver changes that were observe,l following the adminis tration ->f carbon tetrachloride and aleohoi '0.75 <!. of each per kgnt.) to rats which had been exposed hy inhalation methods to i:ich concentration- of triehlornaphthaienes. This rat- had been exposed to triehlornaphthaienes for 1 month. Very extensive necrosis was present in the central areas of each liver lobule. In occasional areas the major portion of several adjacent lobules were neurotic. This rat was aut'-.psied 4f hours after the administration of carbon tetrachloride. WATER PCB-SD0000030445 t I PLATE Y Fig. 1 i i ! i i ! ? ( i > t ' I ! ; t 1 ! i i ?i WATER PCB-SD0000030446 110 JOURNAL ()[ INDUSTRIAL HVGIli.XK AND TOXIUOLOOV (</. JO. no. 2 liver rolls which appeared swollen and more granular than normal. The cytoplasm of occasional cells con tained small acidophilic Hyaline droivlets and there was a moderate excess of fat in the form of tiny vacuoles (fig. 6, plate II). All microscopic abnormalities increased slightly during the second period (37 to 72 days') but no significant advance was detected lietween the "2nd and 134th day of exposure. The livers of rats exposed for 105 days and then removed from exposure for a period of 2 mouths still showed cellular changes similar to those ob served at the beginning of the recov ery' period. In a few of the specimens they appeared slightly loss marked, indicating that some repair had taken place.- In no instance was there evidence of increased damage. Carbon tetrachloride and ethyl alco hol administered to rats exposed in the above manner for 144 days proved highly fatal, out of 10 rats dying within the first 6 days--0 within 72 hours. At autopsy the livers of these rats were yellow and mottled and they were increased in weight (10 to 44 per cent). Microscopic examina tion showed widespread central necro sis similar to that observed in early acute yellow atrophy of man (see figs. 1 and 2, plate VII). The central one-half or two-thirds of each liver lobule showed complete liver cell necrosis and degeneration. Diffuse* extravasation of erythrocytes had occurred and numerous polymorpho nuclear ami mononuclear leucocytes had invaded the necrotic areas. In the narrow perilmrtal zones where the liver cells were less markedly damaged, there were largo numbers of mitotic figures (fig. 2, plate Y1I). (!>) Exposure by inhnlatiun to low conrrtitrations {average 1 ingota. per eu. ni.) S hourn doily.--Animals were subjected to these* conditions for 143 clays. Groups of animals sacrificed after 42, 77, 9S, 11!), and 143 days' exposure showed liver changes that were essentially like those seen in rats exposed 16 hours daily. The carbon tetraeliloride and alcohol test was uniformly fatal and produced liver changes that were identical to those already described and illustrated (figs. 1 and 2, plate VII).' (c) Expo&un- by inhalation to high cuncriilration.i (u itrage S.SS ntgm.s\ p<r cu. m.) IQ hours daily.--Peuta- and hexachlornapiuhalenes when adminis tered in this concentration proved highly toxic. F.igiitv animals wore PI.ATI-; VI Ftn. 1. Photomicrograph X 220 of the liver nf a rut exposed to peuta- and he>:aehiornaphlhnlenos hy tin* feeding of large daily doses fur Hi da vs The liver cells are swollen and markedly vacuolated. There is distortion of the liver ceil columns. In frozen section preparations stained with Seharlach R. there was evidence of marked fatty degeneration. In addition to these changes, numerous oval, round and irregu lar intercellular spaces are present through out the liver lohuies. Such spaces contain a granular serous precipitate, strands of Shrin and occasional leucocytes. These spaces, insofar as could in: determined, were duo to dilatation of the bile capillaries ami eanaliouii. Fin. 2. This photomicrograph X 215 when compared with roe ah<*vc figure dem onstrates the simiiaritv of tin iiver changes produced by tile same compound admin istered in a different manner. In this instance the rat inhaled high concentrations of peuta- and hexaohlornaphthalenes for 3S days. WATER PCB-SD0000030447 PLATE VI WATER PCB-SD0000030448 112 JOl'RXAL OF IXDCSTKIAL HYGIF.XK AXD TOXICOLOGY M. 20, r,n. i used, all of which lost weight and appetite. Four died bj* the end of the first month, 55 within 52 days. Most <jf these were markedly jaundiced. The remainder were sacrificed for pathological examination or suc cumbed to the effects of carbon tetrachloride and alcohol. Only S animals survived the 52 day exposure. The majority of livers were moder ately or markedly enlarged. They were all yellow and in addition the ma jority were markedly mottled. Occa sionally they wore granular and their cut surfaces showed minute spaces which became more prominent after fixation. After an S day exposure, the liver cells were markedly swollen and moderate fatty degeneration was present. This was most mark'd in the central area.- although all portions of the liver lobules were affected. The extent of liver injury rapidly increased so that after 30 days' expo sure, marked structuud as well as cellular changes were apparent. The microscopic appearances were identi cal to those to be described in rats fed large doses of pent a- anil liexarhlornaphthalenes (figs. 1 and 2, plate VI) and in rats fed similar do.-es of !)() percent penta- and hexachlornaplithalenes plus 10 per cent chlorinated diphenyl (see fig. 1, plate III and figs. 1 and 2, plate VIII). Huts removed from exposure be tween the third and fifth week con tinued to die, the longest survival time being 35 days. Microscopic examina tion of the livers of these animals revealed no evidence of recovery. Administration of carbon tetra chloride and alcohol again proved fatal to animals exposed in the above manner. The livers of .>ueh animals showed the characteristic massive necrosis previously described and illus trated. (cl) Exposure b>j fveiling.--When fed in doses of 3 gm. daily to a group of 10 rats, penta- and hexaehlornaphthalones produced marked cellular and structural changes in the liver in a short period of rime. All rats lost weight and appeared ill from the beginning. The longest survival time was 33 day.-. Although the liver edges were blunt and rounded, there were no significant weight varia tions. The livers were friable, yellow and mottled. Microscopically they showed marked swelling and vacuoliza tion of the cells. There was a bo complete degeneration of scattered cells. Occasional-mitotic figures were observed (see fig. 4, plate II). Suit able stains, revealed very marked fatty degeneration. Occasional cells con tained oval shaped or circular aeid- PLATF VII Fic. 1. A low power photnniieioirraph X 45 illustrating the marked d'-grro of degeneration of liver tissue resuitimr fre,m the administration of a small dose of -'arbon tetrachloride and ethyl alcohol (0.75 cc. of each per kirm.) to an animal that had been exposed previously i>v inhalation methods to low concentrations of penta- and hoxa- clilornaphthalrnes for 134 days. This animal died within 24 hours after the admin istration of carbon tetrachloride. Similar degrees of necrosis resulted from the administration of carbon tetrachloride and aleoh.il to rats which had been exposer! to a mixture of !kicI. penta- and hexaehiornaphthalcnes and 10'F rhlorrated diphenyl or to refined chlorinated diphenyl. Fir,. 2. A higher power photomicrograph X 225 of the liver illustrated in the above figure. WATER_PCB-SD0000030449 ^ j < i ) PLATE VII - i` i j Fig. 2 WATER PCB-SD0000030450 114 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY M. 20, no. 2 ophilic hyaline inclusions. In addition to these rvtologieal changes there were numerous varying sized intercellular spaces throughout the livers (see figs. 1 and 2, plate VI and fig. 1, plate III). These spaces were usually circular or oval in shape, although many of them ramified between ceils in the liver columns or between liver cells and the sinusoidal endothelium in such a wav as to suggest that they represented greatly dilated bile capil laries or eaualieuli (fig. 1, plate III). They contained serous precipitate, a lew strands of fibrin, and occasionally a few leucocytes. Rarely one de served one or two degenerating liver cells within these spaces. In some of the more markedly damaged livers, red blood corpuscles were also present. The above microscopic changes a|>pcared identical to those observed in the livers of rats subjected to high concentrations of this compound by inhalation methods (figs. 1 and 2, plate VI). They were also similar to the changes that resulted from the feeding of another penta- and liexaehlornaphthalene compound (figs. 5 and 6, plate I) and from the feeding of a mixture of penta- and hexachlornaphthalene (fiO per cent) and chlori nated diphenyl (10 per cent). When this preparation of penta- and hexachlornaphthalenos was fed in smaller amounts (0.5 gm. every second day to a group of 4 rats) the above described liver changes occurred less rapidly and to a lessor degree (see fig. 5, plate II). Compound E. Penta- and flcxachlornaphthalcncs Exposure by frediny.--Feeding in doses of 1 gm. daily for It) rats was continued with but one 4 day inter ruption. All 10 rats had died or had been sacrificed by the fifty-fifth day. These livers were friable and yellowish, and exhibited moderate to marked mottling. One animal was jaundiced. Of the 5 animals that were sacrificed, three showed enlargement of the liver (30 to 40 pier cent: the remaining livers wore normal in size. On micro scopic examination the livers of rats exposed for 2 weeks showed marked cellular chanties that appeared to be degenerative in nature. Although-the liver cell injury was somewhat greater in the central portions of tin* liver lobules, no portion of the liver was .-pared. The liver cells showed vary ing degrees of swelling, increased granularity and vacuolization of tiic cytoplasm. Marly injury was indi cated by a massing of basophilic granules and small rod-shaped struc tures near the cell nuclei. The* pe ripheral portions of such cells were acidophilic and vacuolated. Small irregular spaces were present between PLATE YH1 Fig. 1. A photomicrograph X 95 of the liver of a rat fed a mixture of penta- and hexachlnrnnphthalenes and IO1'- chlori nated diphenyl, in 3 sm. doses per 10 rat-uor 12 days. This rat survived for 23 days after feeding was stopped. A comparison of the liver of this rat with those of rats sacrificed earlier indicated that the liver ehang".? had progressed considerably after administra tion of the compound had lwen terminated. One should note the marked chance in the liver architecture. Tlte spaces like those previously illustrated (figs. 1 and 2. plate \ I) are very large and the remaining liver tissue is compressed. The liver cells show marked fatty degeneration. Fir,. 2. A higher power photomicrograph X 235 from another area in the liver dlustrated in the above figure. WATER PCB-SD0000030451 LI.J nk, PLATE VIII ; >v ; .V . * >.: ^ % V s?:-.-*.*-. . l .' ,---> .'^v>*. * , . 1 Y'S**'V *<*' ..xVt.V-'' '4'-'.-. -"*" --:r>- ' ' >-? -,< v . ' . { ' . ' . **' ** . "-' 4-,*:ii . :*> .. W3 ... : VTV. >. %, Ip . : --sS ! >W . !/ .'*:;. . V;*v. . "v^; .i V z. , -'VT- ', > S&ZSAS- * .iV?. T; v * ><** A . ,j'* .''* 5 N^V 'j I < . ' -i Tic. 1 Fiu. 2 WATER PCB-SD0000030452 lie JOURNAL OK INDUSTRIAL HYGIENE AND TOXICOLOGY (ro/. 20, no. 2 liver colls and hotween the columns of liver cells and the sinusoidal endo thelium (see fig. 5, plate I). Those were interpreted to he the early stages in the formation of large intercellular spaces (fig. 6, plate I) which subse quently developed and which were identical to the spaces described in the foregoing section. After longer exposure the liver cells showed wry marked swelling and fatty infiltration. Occasional degenerating and regener ating cells were observed. Compound F. 90 per cent Penta- and Ilexachlomaphlhalrncs and 10 per cent Chlorinated Diphenyl (a) Exposure by inhalation.--Inha lation experiments were carried out on two groups of rats. The first of these groups of SU animals were exposed to low concent rat ions (average 1.37 mgms. per cu. m.) 16 hours daily for 134 days. The second group, also comprised of SU rats, was exposed to low concentrations (average 1.66 mgms. per cu. m.) S hours daily for 143 days. No evidence of illness was noted in the living animals in cither group. In each experiment represent ative groups of animals wore sacrificed after exposure periods corresponding to those used in low concentration iuhalatiou experiments with triehlornaphthalcnes and peuta- and hexachloruaphthalcnes. Since there were no recognizable differences in the amount or type of liver change observed after similar exposure periods in these two groups of rats, the second group (exposed .$ hours daily) will not he discussed. Macroscopicallv the livers were usually light yellow. After pro longed exposure, this abnormality was more marked and in many instances morning was present. There were no very significant alterations in the weights of livers although the major ity won* slightly swollen (average of 20 per cent increase in weight). Microscopically the livers showed constant changes after 37 days of exposure. These increased somewhat during the second period of 36 days. Longer exposure produced little if any increase in liver damage. The ob served pathological changes consisted of swelling and ro.unding of liver e-lls accompanied by a definite increase in the prominence of the cytoplasmic granules. Hyaline droplets in the altered cytoplasm were a conspicuous feature (see fig. 3, plate III). Mitotic figures were present in abnormally large numbers. In an occasional liver, as was the case with similar exposure to both peuta- and hexaciilornapluhalencs alone and refined chlorinated diphenyl, there was a marked deposit of finely divided granular and re fract ile yellow or yellow brown pig ment. This was present within liver and Kupffer cells. Larger amounts were found in the cent rat portions of the liver lobules. It failed to stain in a manner characteristic of cither hemosiderin or hemofuscin. The livers of rats removed from exposure for 2 months after an expo sure period of 105 days were Miniiar to those of animals sacrificed at the end of 106 days' exposure. There was, however, no evidence of any increase in the pathological changes. The administration of carbon tetra chloride and alcohol was almost uni formly fatal. Macroscopic examina tion of livers of these rats revealed marked enlargement (approximately WATER PCB-SD0000030453 Feb., 193S] CHLORINATED HYDROCARBONS 117 SO per cent). Such livers were yellow and mottled. Again there was exten sive and widespread liver necrosis on microscopic examination (see figs. 1 and 2, plate VII). (b) Exposure by feeding.--When fed in large doses (3 gm. daily for 10 rats) this compound proved to be very toxic. All animals appeared ill and feeding of the compound was stopped after 12 days. Despite this the rats continued to die, the last dying on the 35th day. Subsequent microscopic examination revealed that the liver changes had continued to progress after feeding was stopped. Macroscopically, the majority of the livers were enlarged, the largest showing an increase in weight of 1 IS per cent; the average increase was approxi mately 40 per cent. All livers were yellow, friable, and many of them were markedly mottled. Subsequent micro scopic examination indicated that much of the mottling was due to hemorrhage into spaces within the liver tissue. In addition, the external and cut surfaces of the livers appeared slightly pitted or granular. The micro scopic changes observed in the livers of the animals autopsied after 10 to 14 days' exposure consisted of moder ate to marked swelling of liver cells accompanied by marked fatty vacu olization. Occasional liver cells showed additional signs of injury' such as nuclear degeneration and poly morphonuclear leucocytic invasion. Scattered circular or oval shaped inter cellular spaces (fig. 1, plate III) like those previously described in experi ments where penta- and hexachlorncphthalenes were employed (figs. 1 and 2, plate VI) were observed. In the less markedly damaged livers, such spaces contained serous precipi tate, strands of fibrin, and small numbers of leucocytes. In rats sur viving 18 to 35 days, these spaces were much larger and often had attained the size of a normal liver lobule (see figs. 1 and 2, plate VIII). In these more severely damaged specimens, extensive hemorrhage had occurred (fig. 2, pia.tc VIII). In sections stained with P.T.A.H. these spaces were seen to be bounded by flattened liver cells which showed sharp cell borders. Liver cells between such spaces were distorted, swollen, and showed marked fatty degeneration. In many' of these livers there was a slight increase in connective tissue between the .remaining cords of liver colls and around recognizable portal areas. Proliferative changes were occasionally observed in the bile ducts. Tins was indicated by increased num bers of ducts in certain areas and by' mitotic figures in the bile duct epi thelial cells. No recognizable dilata tion of medium sized or large bile ducts was present. In many sections the architecture of the liver was so completely altered that it was difficult to recognize the portal areas. In all sections there was evidence of fatty and hyaline degeneration of the re maining liver cells. Slight polymor phonuclear leucocytic infiltration was present. This was somewhat more marked in the periportal areas. Feeding of this mixture of pentaand hexachiomaphthalenes and chlo rinated diphenyl in smaller doses (0.5 gm. every' second day for 4 rats) resulted in liver damage that devel oped less rapidly. The rats were sacrificed after exposure periods of WATER PCB-SD0000030454 118 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY' [roL SO, no. 2 21, 34, 58, and 81 days. Their livers were yellow, increased in size and showed slight mottling. Microscopi cally, the chief abnormality observed was marked fatty vacuolization of liver cells. No intercellular spaces like those described above were present. Compound G. Chlorinated Diphenyl (a) Exposure by inhalation.--Expo sure by inhalation methods to low concentrations of chlorinated diphenyl was carried out on two groups of animals, each comprised of SO rats, in a manner identical to that de scribed in the foregoing sections. In the first group a 16 hour daily expo sure, with an average concentration of 0.57 mgms. per cu. m., was employed. The second group of animals was exposed 8 hours daily to an average concentration of 0.03 mgms. per cu. m. Neither group of rats appeared ill. Representative animals were sacrificed in groups of 3 to 21 rats, after varying periods of exposure. In the first experiment, the animals were sacri ficed after 37, 72, 105, and 134 days of exposure, in the second experiment after 42, 77, 9S, 119, and 143 days. The livers wore pale or slightly yellow and somewhat mottled. There were no constant variations in the weights of the livers. The microscopic find ings m the two groups indicated that the different exposures had resulted in the same degree of liver injury. The pigmentation of the liver and Kupffer cells was somewhat greater in animals exposed 8 hours daily. In both experimental groups the carbon tetrachloride and alcohol test was highly fatal and led to the same exten sive central necrosis of liver tissue that was observed in rats exposed to the toxic, chlorinated naphthalene compounds (figs. 1 and 2, plate VII). The microscopic changes resulting from exposure to low concentration of chlorinated diphenyl alone were simi lar to those resulting from inhalation of penta- and hexr.chlomaphrhalenes, plus 10 per cent chlorinated diphenyl. The most conspicuous change was hyaline degeneration (fig. G, piate III) although swelling of liver cells, in creased prominence of cytoplasmic granules and increased vacuolization were present. Mitotic figures were present in increased numbers. No microscopic evidence of recovery was found in the livers of rats exposed for 105 days and then removed from exposure for a period of 2 months. (1>) Exposure by fconing.--When fed in large daily doses (3 gm. for groups of 10 rats), chlorinated diphenyl proved highly toxic, reeding was discontinued after G days. Seven rats died within the first S days. The 3 remaining rats gained in weight after the feeding was stopped and were finally sncrificed. Tire last animal was sacrificed 63 days after feeding was begun. The majority of livers showed a moderate increase in weight. Micro scopically there was evidence that liver injur}' had occurred within the first few days. The cells were swollen, at times sufficiently to obscure the sinu soids. The cytoplasm of the liver cells was acidophilic in staining quality and contained small vacuoles. Small eosin-stained hyaline granules and globules were present. Mitotic figures were numerous (see fig. 4, plate III). There was little inflammatory cell infiltration and no structural changes were observed. 1I ii t ! ! I \ \ l t t WATER PCB-SD0000030455 Feb., 1*3$] CHLORINATED HYDROCARBONS 119 When smaller doses (0.5 gm. every -exposure, no pathological changes second day) of this compound were attributable to the compound injected, administered to 10 rats, the first death were demonstrable. Tho mixtures of occurred after 9 days. Two addi tetra- and pentachlomaphthalenes and tional deatlis occurred after 12 days of penta- and fcexaehloruaphthalencs and two more by the end of the fifth proved highly fatal and degeneration week. The remaining animals were of liver tissue was observed. From sacrificed between the 40th and the their findings Flinn and Jarvik con 190th day. The livers were enlarged, cluded that certain chlorinated naph the average increase in liver weight thalenes or impurities contained in being 33 per cent. They were pale them are capable of producing yellow and slightly mottled. The most out atrophy of the liver in the rabbit. It standing microscopic abnormality was is noteworthy, in the light of the the presence of large numbers of present experiments, that the com varying sized acidophilic hyaline glob pound of lowest chlorination (a mix ules in the cytoplasm (sec fig. 5, plate ture of tri- and tctrachlornaphtha- III). These hyaline globules, although lenes) had no apparent effect on the larger and more numerous, appeared well being of the animals and did not identical to those encountered in the produce demonstrable changes in the livers of rats exposed to other chlori liver although injected daily for a nated diphenyl preparations. The period of 2 mouths. liver cells were markedly swollen and The present experiments demon showed fatty vacuolization. There strate that chlorinated naphthalene was also increased regenerative activ compounds and chlorinated diphenyl ity. No structural changes were ob are capable of producing marked liver served in any of the livers. . damage in the white rat without Discussion demonstrable microscopic changes ap pearing in the other organs. Further Published reports concerned with more, the characteristics of the liver the possible systemic effects of chlori lesions resulting from comparable nated naphthalenes have been few in amounts of any given compound are numbers. Lehmann (3) in 2919 re the same, regardless of the method of ported that animals exposed to chlori administration (inhalation, feeding, or nated naphthalenes by inhalation or subcutaneous injection). feeding refused to eat and at death An attempt has been made through showed "peculiar" lesions in the liver. out this study to grade the toxicity of In 1936, Flinn and Jarvik (4) reported each of the several compounds tested the results observed following the in accordance with the severity of the daily subcutaneous injections into 30 liver damage produced. Although rabbits of enormous doses (30 mgrn.) some of these compounds appeared to of certain chlorinated naphthalene produce similar degrees of liver injury, compounds dissolved in paraffin oil. the toxicity of each compound seemed The rabbits receiving a mixture of tri- directly reiated to its degree of chlorin and tetrachlornaphthalenes lived and ation. Thus, a mixture of trichlor- when sacrificed, after a 2 months' naphthaJenes (chlorine content 49.4 ! WATER PCB-SD0000030456 120 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY [vol. SO, no. 2 per cent) proved to be the least toxic of any of the compounds tested, whereas chlorinated diphenyl (chlorine content 65.0 per cent) was highly toxic even in very low concentrations. Trichlomaphthalcnes, when com pared to other naphthalene compounds of higher degrees of chlorination, were relatively innocuous. Even alter pro longed feeding of large amounts of this material, the rats showed no evidence of ill health. After 2 months' exposure, microscopic examination of the livers revealed only slight to moderate degrees of fatty infiltration and degeneration of liver cells (plate II, figs. 1 and 2). These changes were slight when compared to those result ing from short exposures to compounds of higher chlorinaiion. Exposure by inhalation to low concentrations of triehlornaphthalenes produced liver cell alterations that were never more than minimal (see fig. 3, plate II). Such animals withstood additional liver injury from carbon tetrachloride and alcohol in a manner indistinguish able from that of the normal animal. Somewhat greater liver injury re sulted from inhalation when the air concentration of tliis compound was increased more than eight times that used hi the previous experiment but again the liver changes were slight compared to those produced by com parable exposure to the more highly chlorinated compounds. The mixture of tetra- and pentachlomaphthalenes employed in these experiments appeared to be consider ably more toxic than was the mixture of trichiomaphthaienes. Animals fed small doses of the former of these preparations fell ill ana died or were sacrificed by the G3rd day. Their livers showed extensive fatty innitra tion and fatty degeneration (figs. 3 and 4, plate I). Feeding of tetra- and pentachlor- naphtlialenes in combination with chlorinated diphenyl resulted in pro nounced liver changes. These livers had increased in weight (average 71 per cent). Microscopic examination revealed a peculiar type of hyaline degeneration involving practically every liver cell (see figs. 1 and 2, plate I). This type of cell degeneration was more marked and occurred earlier after exposure to preparations con taining chlorinated diphenyl than to any other compounds tested. Fur thermore, it was most marked in the livers of animals exposed to refined chlorinated diphenyl (figs. 4, 5, and 6, plate III). Comparable exposure of rats to penta- and hcxachlornaphthalenes (Compound D), penta- and hexachlor- uaphthalenes (Compound E), and penta- and hcxachlornaphthalenes (90 per cent) plus chlorinated diphenyl (10 per cent) (Compound F), resulted in liver changes having few detectable differences. The morphological changes were slightly greater in the livers of rats exposed to the last named compound. In these instances the hyalin degeneration of the cell cytoplasm was more marked. Al though rats inhaling low concentra tions of compounds D and F showed no demonstrable signs of ill .health, microscopic: examination of their livers revealed marked liver cell injury' (fig. 6, plate II, and fig. 3, plate III). These lesions were still demonstrable after a 2 months' .recovery period. Further evidence that liver damage had resulted from inhalation in low concentrations of these compounds was obtained from the carbon tetra- Feb. chlc wer rest (% tfcrtox: typ in 1,.: 1 : eh: la? I i1 i WATER PCB-SD0000030457 Feb., 1938} CHLORINATED HYDROCARBONS 121 chloride and alcohol test. Such tests were almost uniformly fatal and resulted la widespread liver necrosis (figs. 1 and 2, plate VII). These three compounds proved to be very toxic and resulted in a most unusual type of liver change when administered in large amouuts i>v feeding (see fig. 1, plate III, fig. 1, plate VI, and figs. 1 and 2, plate VIII). Identical changes were observed following inha lation in high concentrations of com pound D (see fig. 2, plate VI). In. such livers the most conspicuous feature' was the presence of large intercellular spaces. Examination of routine sections, supplemented by specially stained preparations indi cated that those spaces bad resulted from dilatation of bile eanalieuli. The reason for such dilatation was not apparent. There was no correspond - -J.M- i.U UiU, UiJilUlLXVJil SJ4 WJU UtAgW tL&Al* bile ducts at the periphery of the liver lobules and no inspissated bile was ever observed. The organically com bined chloride in the livers of these rats was no greater than that observed in the normal animal. Although seri ous liver damage results from exposure to high concentrations of these com pounds, the resulting changes are most unusual and do not resemble those of acute yellow atrophy on microscopic examination. Of the various chlorinated hydro carbons tested, chlorinated diphenyl gave evidence of being the most toxic. When administered by inhalation in very low concentrations (average 0.57 to 0.93 mgms. per cu. m.) liver cell changes were very pronounced after the first- exposure period. The most striking change was the hyaiinization of the cell cytoplasm (see fig. 0, plate III). Such cellular alterations were essentially unchanged after a 2 mouth recovery period. In these animals small suhlcthai doses of carbon tetra chloride and alcohol uniformly pro duced extensive liver necrosis and was highly fatal to them (figs. 1 and 2, plate VII). Chlorinated diphenyl fed in small uoses produced similar but more marked liver injury (see fig. 5, plate III). In largo doses this com pound was highly fatal. The liver changes in animals dying after short exposures were inconspicuous and consisted mainly of swelling of cells and active regeneration (see fig. 4, plate III). However, animals re moved from exposure before being fatally poisoned, subsequently devel oped hyaline degeneration of liver cells similar to that produced by pro longed administration of small doses of this compound. Tims the results of the present study, as well as certain field studies that have been made (1) suggest that the solution of the industrial hazard involved is dependent largely on a reduction of the air concentration of these compounds to a level that will not produce liver damage. The pres ent experiments indicate that lower air concentrations must be obtained in the case of the more highly chlori nated naphthalene compounds and chlorinated diphenyl than for trichlornaphthalenes if a safe environment for workmen is to be assured. Because of the pronounced toxic effect of small doses of carbon tetrachloride on the livers of animals already injured by exposure to chlorinated naphthalenes and chlorinated diphenyl, its use as a solvent for these compounds would appear to be very hazardous. Although this investigation was designed to determine what systemic WATER 122 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY M. 20, no. 2 effects, if tiny, would be produced in experimental animals by a group of chlorinated hydrocarbons, the patho logical changes produced have been of such a nature as to justify further comment. Exposure of rats to highly chlorinated naphthalene compounds and chlorinated diphenyl results in a type of liver injury differing consider ably from the liver lesions resulting from carbon tetrachloride (5, C) and chloroform poisoning (7). The changes resulting from exposure to the compounds employed in this study involve all portions of the liver lobule. This is true despite the fact that the initial morphological changes are more pronounced in the liver cells nearest the hepatic veins. The changes ob served arc degenerative in nature. The successive degenerative changes were cloudy swelling, fatty infiltration and fatty degeneration and finally, the complete disintegration of the coll. Degeneration characterized by the accumulations of large amounts of acidophilic hyaline material was also observed. This feature was much more marked in the livers of animals exposed to the comi>ounds containing chlorinated diphenyl. It is of interest that widespread degenerative changes of ail liver cells can exist for several months without producing any evi dence of ill health in the animals or without causing important structural changes in their livers. Such findings suggest that- the alterations take place slowly and that the scattered necrotic cells are efficiently removed and re placed by new cells which also become injured. There was no observable morphological evidence to indicate that the regenerating liver cells had acquired an increased resistance to the injurious effects of the compound being used (S). The fact that liver cell changes were still present after a 2 month period during which the ani mals were removed from exposure to the more highly chlorinated com pounds is further evidence that this type of injury is persistent and only slowly recovered from. The administration in high concen trations of two compounds of inter mediate degrees of chlorination re sulted in marked structural changes in addition to marked degenerative changes in liver cells. The most conspicuous change was the occurrence of progressively enlarging intercellular spaces. These were interpreted as markedly dilated bile canaliculi. We are unaware of such changes having been preriously described. From the above it is apparent that exposure to highly chlorinated naph thalenes and chlorinated diphenyl not only results in liver changes having marked differences from those caused by other well known toxic agents, but also that the characteristics of the liver changes produced can be mark edly altered quantitatively and quali tatively by varying the degree of exposure. For these reasons one wouid seem justified in suggesting that detailed studies on animals in which the types of liver injury observed in these experiments had been produced might further our understanding of liver ceil function. Sfmmary 1. Studies concerning the effect upon white rats of a group of chlorin ated naphthalene compounds and chlorinated diphenyl are reported. This investigation was undertaken because of the recent occurrence of 3 fatal eases of jaundice in men work WATER PCB Feb., 1S3S] CHLORINATED HYDROCARBONS 123 ing in industrial plants where these compounds were being used. The compounds tested were selected as representative of a certain range of chlorination and because of their relative industrial importance. 2. Four of the compounds tested were administered by inhalation. The air concentration and exposure pc-ricds were varied. All compounds were tested by feeding in large and small amounts. In addition three of them were tested on small numbers of ani mals by subcutaneous injections. The effects of any given compound did not appear to be influenced by the method of administration. However, marked quantitative and qualitative differences in the effects of each of several compounds resulted from vary ing the air concentrations in inhalation, experiments and the dosage employed in feeding experiments. 3. Macroscopic and microscopic examination of the tissues of rats exposed to these compounds indicated that the injurious effects are mani fested solely in the liver. Blood examination during life revealed no significant abnormalities. 4. The liver changes resulting from exposure to the chlorinated hydro carbons employed have been described and illustrated. As indicated by their ability to injure liver tissue the toxicity of these compounds increases with increasing degrees of chlorination. Thus a mixture of triclilornaphthalenes is relatively innocuous iu low concentrations and v.heu compared with compounds of higher chlorina tion, its toxic properties even in higher concentrations are relatively slight. Chlorinated diphenyl appears to bo the most injurious compound of all those tested. 5. Administration of small sublethal doses of carbon tetrachloride and ethyl alcohol to rats whose livers have already been injured by the com pounds under consideration is highly fatal and produces massive necrosis of the liver. 6. The significance of this study in relation to the industrial problem involved is discussed. 7. It is suggested that the types of liver injury observed in this study might be employed in experiments designed to study liver ceil function. BIBLIOGRAPHY 1. Drixeer, C. K., TVarp.en, M. F., and Beckett, G. A.: The problem of pos sible systemic effects from certain chlorinated hydrocarbons. This J., IS, 2S3 (1037). 2. Lamson, F. D.: Personal communication. 3. Lehmann. K. B.: Kur7.es Lcbrbuch dor Arbeit und Gewerbobygiene. S. Eirzcl, Leipzig, 1919 (p.251). 4. Flixx, F. B., and Jarvie, N. E.: Action of certain chlorinated naphthalenes on the liver. Free. Soc. Exp. Biol, and Med., 35, IIS (i03G). 5. Gardner., G. H., Grove, R. C., Gustaf son, R. K., Marie. E. D., Thompson, O,1. J., Wells, H. 5., ans Lamson, P. D.: Studies on the pathological histology of experimental carbon tetra chloride poisoning. Bull. Johns Hop kins Hosp., SC, 107 (1325). 6. Cameron, G. F... .and Karcnaratnf., IV. A. E.: Carbon tetrachloride cirrho sis in relation to liver regeneration. J. Path, and Bact., JJ. 1 (1S36). 7. Whipple, G. K., and Sperry, J. A.: Chloroform poisoning. Liver necrosis and repair. Bull. Johns Hopkins Hosp., iO. 27S (1009). 8. MacNidkr. W. A.: A study of the ac quired resistance of the fixed tissue cells morphologically altered through processes of repair. J. Pharm. and Exp. Therap., 50, 353 (193C). WATER NEV 130808 WATER PCB-SD0000030461 REPORT OK 4465. Inhalation Experiments Animal a. VJhite rats have been employed throughout. They permit the use of a large number of animals In a relatively smell inhalation installation. They thrive upon, a diet very similar to man, and in the case of these chlorinated compounds it is possible that diet my be very significant* Finally, their normal characteristics have been described so veil as to meke the detection of abnormalities both easy and certain* Method of Exposure. The inhalation experiments were carried out in four large air-tight wooden boxes, each capable of holding ten rat cages, size 22" x 22" x 14", in two tiers of five cages each. Rhen the experiment was not in progress the doors were opened wide and the oages kept in place (see Figure 1). At the center of one end of each box ~ the Inflow end -- air was introduced through a pipe 7 inches in diameter (see Figure 2). Each box was equipped with an individual electric blower which blew the air through several feet of 7-inch pipe before entering the end of the box. An orifice was placed in the pipe line, and the flow of air in cubic feet per minute could be read off directly frau calibrated flow meters. A damper plaoed at the entrance of the pipe into the box was adjusted to assure a uniform distribution of the stream of air to the two tiers of cages. At the opposite end of the box -- the outflow end -- the air from each box was exhausted through a 7-inch pipe fitted with a damper and connected to a large central exhaust fan (see Figure 3). About 4 Inches from the entrance of the 7-inoh pipe into the box, the fume a of the substance were introduced into the inflowing air. NEV 130809 WATER PCB-SD0000030462 ! Itoat view aad inflow end of two boxes wita rat cages ia place aad doors open* NEV 130810 WATER PCB-SD0000030463 Fig 2 Kleetrie heater for maintaining chlorinated diphenyl at a constant temperature in place at Inflow end of box NIV 130811 WATER PCB-SD0000030464 : NEV 130812 WATER PCB-SD0000030465 Hg# 4 lyres glass flasks* NEV 130813 WATER PCB-SD0000030466 Specially designed pyrex glass flasks (Figure 4), 7 3/4" long and with a diameter of 1 l/2", were used to hold the heated waxes* These flasks were made with a side ana and tube whloh extended to the bottom* The top of the flask was ground, and into this fitted a short tube 1 1/2" in length. This short tube was inserted into a large rubber stopper which fitted tightly into a hole cut out of the 7-inch pipe on the under side (see Figure ) The flask in turn was placed in an eleotric heater made to coyer it completely below the side arm and ground glass top* Rubber tubing connected the side arm with a compressed air reservoir and a gentle stream of air blown through the ftelted wax kept it in . motion and assured uniform heating. Into each flask was inserted & long stem centigrade thermometer which was kept in place and could be read at any time above the 7-inch pipe through which it passed (see Figure 2). Approximately 2D gm* of pulverized .nrw aftxffrxxng chlorinated diphenyl were piaoed in the VaJsbrlzlAg*-ih flask and melted in the electrio heater. Ifresh samples were used every other day tertrxVtrmimrig^^ munaaaany Aa ift&d BQ aBbfltacaguuteiflu^^ the collected sublimate was always removed from the upper part of the flask and a clean top used each day. No sanjple was ever used far more than two runs. The flask plus the contents was carefully weighed at the beginning of the run and at the end, and the loss in weight used to calculate the average amount in a oubic meter of air per minute as determined by a series of flow inter readings. The figures obtained were not absolute because of slight variations in the air flowing through the boxes and bscause of deposition of material on the thermome ters and on the inside surfaces of the box, but they checked well with dlreot determinations through air sanjplee. NEV 130814 WATER PCB-SD0000030467 Approximately one hour me allowed for the wax in the flasks to malt end come to a ooastant temperature. At that time the box doors were tightly closed, air bubbled through the flasks, and the blowers turned on. This was the beginning of the exposure period. By meens of the theostats on the fans and the dampers in the outflow pipes, the amount of air flowing through the boxes was adjusted and an attempt was made to keep the four boxes as uniform as possible -- usually between 165 and 175 cubic feet per minute*. Experiments Exposure to chlorinated diphenyl was begun an July 1, 1936 and ceased on November 18 of the same year* The average length of exposure was 16 hour8 daily for 6 days a week. Each morning at about 9:00 exposure ceased, and between this time and 4:00 p.m. the rats were cleaned, fed, weighed, etc. In order to secure uniformity of exposure the cages were shifted on a regular schedule, cdnce wax concentrations were somewhat higher at the Inflow end of the boxes. Preliminary runs showed that once properly adjusted, wax concentrations in the air remained very uniform from day to day, but to insure absolute safety readings of temperature, air-flow, etc. were made every night between 10:00 and 18:00 p.m. as well as on starting and stopping. At different times during the course of the experiment tests for free chloride were made but were uniformly negative and showed that under the temperatures used no decomposition occurred. TrtdJLl NEV 130815 Conditions Maintained in Inhalation Experiment from July 1 to Nov. 18 Material ' Chlorinated diphenyl Temperature Average concentration . of air in box Total length of exposure 165-175 Mg./ou.U. Range ft.'5.7, High 1*19 Low 0.23 hours 1696 Average daily exposure hours 16 WATER PCB-SD0000030468 _ Results of Inhalation Experiments Ho abnormalities were seen at any time in the living rats. Blood and urine examinations were made; there were no losses in weight, and one would have considered that the animals were perfectly sound. However, rats sacrificed after 6 weeks exposure showed slight liver damage, which advanced during the next two months. The changes consisted in slight to moderate swelling of the liver cells, increased granularity, and many mitotic figures. Hyaline inclusions were present and were invariably plentiful in animals receiving chlorinated diphenyl by inhalation or by mouth. t One is forced to oonclude from the experiments that an average concentration of 0.57 mg. per cubic meter Inhaled 16 hours daily produces definite slight changes in the liver and in this organ alone. These changes are resisted efficiently by the animals and cause no depreciation of health. The situation is not unlike that seen in factories where acute yellow atrophy of the liver has occurred. In the case of such instances the patients have been singled out in some way or other from ' large groups of fellow workmen who have been perfectly healthy. In considering the entire matter it seemed to us that the mssnxft chlorinated hydro-carbons,/if inhaled in sufficient concentration, might cause a slight degree of damage to the liver. This damage is resisted efficiently and causes no depreciation of health, but if the individual in* question happens to suffer seme ordinary disease of the liver the condition is superimposed upon a substratum of Injury. In accordance with this hypothesis we determined that a dose of 0.75 oe. of carbon tetrachloride plus 0.75 co. of ethyl alcohol per kilogram of rat was entirely non-toxic to normal animals. When, however, this dose was given to animals which had Inhaled chlorinated diphenyl as has been described in this experiment, the result was acute yellow atrophy NEV 130816 WATER PCB-SD0000030469 -5'i"" of the liver. It would seem therefore that Inhalation of a concentration of chlorinated diphenyl in the manner I have described is capable of producing a condition will oh may be dangerous to the individual inhaling it, though of Itself no visible harm will be done. Animals removed from exposure to chlorinated diphenyl after two months Inhalation and given two months for recovery still showed liver changes when killed and examined, so that it is evident the alterations produced by this hydro-carbon are persistent and individuals who have been oaused trouble by it should be removed completely and for a long time. The Inhalation of a somewhat higher concentration of ' chlorinated diphenyl over an 8 hour day The circumstances attending this experiment are given in Table 2. * Material ._ Table 2 ' Temperature ' Average concentration of air in box _________________ _ i___________________________ ______ _____ o 0 Mg,/cu.m. Baaga . Total length Average of exposure daily exposure Hours Hours Chlorinated diphenyl 153-174 0.93 High 3.23 Low 0.03 920 8' Again 80 rats were used in this experiment. None at any time showed the slightest evidenoe of illness. Microscopic examinations, however, showed a degree of change very comparable to that found in the 16 hour experiments which have been described and the carbon tetrachloride test was uniformly fatal. General Summary. From these experiments we are foroed to conclude that concentrations of chlorinated diphenyl in workrooms should not be allowed to rise above 0.5 mg. per pubio meter of air. This ccncentraticn is readily obtained by careful . NEV 130817 WATER PCB-SD0000030470 hooding* It la in our opinion oafs for workman who would not be exposed with the absolute steadiness characteristic of our experiments on rats and such a concentration limit would probably do away with the troublesome skin lesions which have been tha particular cause for worry in regard to these compounds in tbs past* In addition to the inhalation experiments many feeding experiments . this were done with/chlorinated diphenyl. The general changes in the liver were the same as those already described* Wo diarrhea or gastrointestinal aspects seemed to he present. NEV 130818 WATER PCB-SD0000030471 i REPCRT TO THE MONSANTO CHiMOAL CCUW by Oeoll K, Drinker, l',D. Dean and Professor of Physiology Harvard School of Public Health, Boston, lines, SeptCKbnr 35, 1950 NEV 118912 WATER PCB-SD0000030472  ( i 1 Experiments to determine the possible toxicity of tbo following sub8tano0s:j V*. 2 1. jhloroosane ... 2 2. Diphenyl phthalat 3 5. Chlorinated diphenyl #2268....................................................... 4 4. ijixture of chlorinated diphenyl and chlorinated liphenyl benzene #5460................................................. ... 7 II. Some genorai oonsideratlone: 1. Evidenoe for the deotruction of a mixture of pente and hexachlornaphthalenes (#1006) in the body ..... 9 2. Ihe effect of increasing the sodium chloride in the diot upon animals receiving toxio -doset; of #3.006 .... 30 5. Tie effect of high and low calcium intake on animals libeling high concentrations of #1006 ................... ... 10 4. Til effect of inject ions of xanthine on animals libeling high concentrations of #1006 ........................... . 11 NEV 118913 WATER PCB-SD0000030473 i K" I. yxramciAs to pbthb.skb the possima Tcociom of the' yoimim suivypAHnFs l. CHLORCOSANK On July :i4 1937, we wore informed by T. D. Stoith tlmt each 6 ounce bottle of a soft drink night contain 00 to 70 ngm. of chlorcosane, Shore was no pharmacological reason to bellove the compound would be irritating or toxic in any way, and 1 herefore it was decided to dose rats with it Tory heavily, the argument beinj; that if they survived heavy dosage without any evidenco of damage a verdict of no tozlolty might bo given and much time saved. If, on the other hand, tie large doses proved poisonous it would be necessary to reduce dosage until a non-toxic level wa6 reached. The first of these alternatives proved correct. Experiments. -- August 10, 1937. 0 edult white rate ranging in weight from lf*3 to 215 gm. wore each fod 70 mpn. of chlorcor.one in 1 eo. of olive oil dally by stoma ih tube. These animals shovied no evidenoe of damage in any way attributable t > chlorooaene. They were examined microscopically from the 3Dth to tbo 92nd da:. The only constant change observed was a possible slight increase in th > granularity of the colls of the liver cords, but the pathologist in oharge felt this to be within normal limits. This finding is doubly reassuring since many of the animals developed a chronic suppurative lung condition|very prevalent in Boston stock rats at tbo timo, But even with this added handicap there was no evidenco of damage and we thus conclude that ehloroosnne evon in enormous dosage is an inort material in the body and in all probability pnfsos through tbo intestiue unchanged. NEV 118914 WATER PCB-SD0000030474 i 8. DIPHENYL FHTHALATS On August 83, 1937, wo were informed by Dr. R. X. Kelly that diphenyl phthalate might be absorbed through the skin or might be Inhaled and swallowed during' spraying operations. After consultation it wae decided to food heavily, and if there were indications of toxicity to project other experiments. Experiments. -- September 80, 1937. 80 adult white rats ranging In weight from 848 to 3)1 gn. were each fed 0.5 mgm. of diphenyl phthalate suspended in 1 oc. of wate:' by stomach tube dally. The animals were from new stock and with one exception remained clear cf tha lung condition which existed in the ohlorcosane group. During the period of experiment, which lasted 98 duys, practically all the animals gained weight. Examination of 8 animals killed at Intervale curing thiB period of test resulted ae follows: The orgars were normal grossly, and on microscopic examination lho liver alone was of yossible interost. The changes in the liver varied from those in whieb tho ocIIb of the livor cords vusre almost normal, having only a slight svrolling and granularity of their cytoplasm, to those in wbioh these changes wore pronounco3, In the letter there was a moderate dogree of vacuolization Or* and a raro hyaLino body. These alterations it must be understood wore the result of very certain dosage, since tho compound was given by stomach tube and the animal t had no possible ray of avoiding it. Neither tho pathologist nor myself was able to consider them of enough moment..to couse us to make further exporlnonts. X am oonfi dent that with the ordinary precautions accompanying spray lecouering no poccihlo hern could be done, even if the concentration of diphonyl phthalate in the locouor nun far nbovo the 0 nor cent figure given us by U. E. itolly. . NEV U*915 WATER PCB-SD0000030475 S. 0HHMIKAT2D DIPIiaftL -- OOLB'OUHD #1260 This material was furnished by the Monsanto Chemioul Conpany. It wus edd to have a Ichlorine content of 68 per eent, the higheol ehlorlne figure of any eotnpound tested by ue during tho past three years. It had boon our impression that; toxicity and chlorine content followed one another closely, but this has not turned out to be the case for #1268.actdxiuxsixnxseiissxocitis ^axxwapssxtes Further discuscion of this point will follow the description of the experlmo itel observations and their results. Experiment;. -- august 2, 1937. Tho compound #1268 was administered by inhalation, tho techniquo being thet described in the paper entitled The Problcn of Possible Sye;cialo Effoots from Certain Chlorinotod Hydrocarbons, Journal of Industrial Hygiene, 1937, 19, 283. For the first observations 80 adult white rets were used.| They v;ere exposed ca follows; Group 1. --[ 80 onimels. Te;r,porfituro to which tho compound was boutod to Introduce fume into air line 140-190 C. Average concentration in air breathed by rata 0,53 jngu. per cubic meter. Average dally exposure 16 hours. Tho experiment was continued for 119 days, euiionls being oacrificed at intervale for pathological examination and for tho carbon tetrachloride and alcohol test (see paper previously citod). Tho conditions described above caused swelling end incroure of granularity of the liver cells. Hyaline Indusions were rare. Those changes worn quite uniform in oil eiln&ls examined after the Slat day, but wore not certainly progressive. Ths rats wars very haaltliy throughout tha period and thero was an 'almost uniform c: \in lnwoight. Tho carbon tctrachloride-ulcohol teBt was positive after Bi days, which indicates that though liver damage wob apparently slight sous degroe of h;..;v. had been duno tho organ. There wan no evidence of NEV 116916 WATER PCB-SD0000030476 I damage in any part of tbe body except the liver. At the end of 93 days 10 rata, apparently in excellent condition, wore removed from the experiment and oot eside for observation?; upon liver rocovery. These recovery' animals showed no clinical chances of any 6ort. When sacrificed 78 and 141 dairs after removal from exposure it was observed that the swelling of the liver sella had disappeared, but the granular and liyollno material remained in tie liver cells and had apparently become permanent. There was absolutely no progression of damage efter removal from exposure. These ch niges may be compared with those produced in rats by inhalation of chlorinated diphenyl #4465, administered in a similar manner in concentra tions of 0,57 to 0.93 rajm, per cubic rioter over sinllar periods of time. In the case of this compound the conspicuous difference was the fur greater Incidence of liyalinisation of tho liver cells, which in our opinion vine the characterlet it lesion caused by jjwwi ond mixtures containing it. Ono can therefore conclude that #1860 in low concentrations ic definitely lose toxic than #4465. . Group 8. -- After 119 days, tho low concentration of #1860 having proved but slightly joloonous, it was decided to increase the concentration in the air breathed by tie 54 ruts then remaining. Temperature and other conditions were 'wiintc ir.cd ns during the first 119 fioys, but by the ucu of 4 vapor inert; instead of 1 the avert) go concentration of #1860 in tho air wan brought to 6.83 iiigw. per cubic netor of air. Exposure to this very high concentration was continued for 07 days. . . Undor these extroiw conditions the animals again remained in perfect health. Tho carbon tetrachloride t.ri ulcoV^l tent. var. positive but there wan 4 . - absolutely no >ther indication of liver darings and no evidence of fliotuvborno to other argunls NEV 118917 WATER PCB-SD0000030477 i [` Vhm eiamined at autopay at the end of 87 deya -- the animals had experienced a total exposure of 806 days -- there were no gross evidences of abnormality in any part. On microscopic examination no organ showed changes except the liver. The number of cells containing hyaline and the amount of hyaline in t cello involved increased during this period of exposuro to the hlghor concentration of #1268. The hyaline was most abundant in the portal zone of the lobule with very little in the central area. After 42 days of exposure 10 rats were set aside and were sacrificed 75 days later. They wore in excellent condition when removed and did not change. Grossly, at autopsy, they showed nothing abnormal, and on microscopic examination the liver colls had lost tboir swelling but retainod increased granularity and hyaline inclusions. The experiments on inhalations of high concentrations of #1860 reenforce the oottclusio| that this compound is of low toxicity ot compared with #4465 or with chlorinated naphthalenes above triehlornapbthaloiie. Tho question as to why #1860, tho moot highly chlorinatod coinpound tested, proved but slightly harmful cannot bo answered with any definitenocs. It has been suggested that the toxicity of alL these chlorinated compounds, even though of varied composition, may depend on the obilily of the animal to decompose them eftor lodgmont in the tissues, and t:>at thin decomposition might be shovm by on increase in the chlorine in the urine is suitably conducted feeding nxjwrlrjontr.. In experiments upon thispoint we have ihown e definito Increase in the urinary chlorides when dogs and rats were fed mixture of penta and bexachloraaphihalanec (#1006), a compound highly injurioi to tho liver and containing C2.6 per cent of chlorine, but similar observations hub*o not been made with any of tho chlorinated diphenyls or allied compounds. It inay, however, bo that when about 65 per cent clilor inntiou is ruocbcii tho substances rorr-ad era ouito stable in tho body and no cuuuo a. minimum of dnimtgo. NEV 116918 WATER PCB-SD0000030478 -v- lo oonolnision( #1868, if handled with ordinary jireonutlonB ac to ventilation should bo entlrely harmleos to workmen, While it cannot bo given an absolutely elo'an bill as to health, it is preferable to #4460 and #i>4C0. 4. MIXTURE OF CHLORINATED DXPHKNY1 AND CHLORINATSSD Dmm BENZENE -- COMPOUND #0460 Ibis substance wus furnished by the Monsanto Chemical Conpniiy end v/e/s Bald to have a ehiopine content' of 60 per cent, being In thie respect bolcnv #4460 and far below #186&. Experiments. -- The inhalation toclmique nae used an in tho ear.o of '18t0 and #4465, August 2, 1937. 00 adult white rets wore tho subjects. Temperature to which the compound wes hoatod to introduce into the oir line 140-If6 C. Average concentration in eir breathod by rats 0,085 mgm. per cubic meter. Average dally exposure 16 hours. Tito'experiment lasted 119 days, a certain number of animals wore sacrificed for pathological examination, others wore used for tho oorbon tetrachloride end alcohol tost, and still others set tsiOo for observationii aa to recovery from poeslblo damage. In spite o:' tho fact that tho concentration in tho rir breathed by tho r n; averaged about '. /& that obtained at comparable tempers! tu a from '"lhOC, t. mu ;> r of theso onimolr became siejc and lent weight towards tho end of tho record m'. s th of exposure. T/lsn hilled, such individuals showed grotsc mottling of il.o liver but no changes in other organs. On microscopic examination swelling of the liver colic, increased granularity and hyaline inclusion:' x.oro notal in .:ii .. hilled as early is tho 16th day of exposure. Theso changes-obviously occurred rapidly. Kynlins doj>oslts wore never as numerous us with #1860. /Almal's NEV 118919 WATER PCB-SD0000030479 ~8~ removed for recovery after 51 and 119 days of exposuro did not Rain markedly In weight, and one died for no obvious cause, tfhen oxtuiiined at autopsy the swelling of tho' liver cells bad subsided, but abnormal grocularity and byallnlnatlon remained. Needless t say the carbon tetraoblorldo and alcohol test was positive whenever usod. In view of| the foot that 05460 in euoh low concentration proved so definitely toxlb, no higher concentrations wore tested. It seems imperative that whenever tils compound Is used In industry, Great care be taken to keop concentrations .n the air at an extremoly low level. No liberties can he taken with It, us with rl20P. I N6V 118920 WATER PCB-SD0000030480 II. 801.23 Q35C5t/lL CONSIDERATI01!S la adaitp.on to these teste of compounds, certain experiments were done which are of ^aterest to those manufacturing or using chlorinated hydrocarbons, Details as to these experiments are of course available but aro not included in this report since they are not of direct industrial interest. 1. [-EVI1XKNC3 FOR TK3 K331KUCTION OF A MXlVHIt OF PUN'fA AND J2)CA0IIL0IUlAHrniALlSaS (#1006) IN TUB BODY It has always been a question as to whether the chlorinated hydrocarbons which have boon examined by oursolves end by others do harm per se or whether toxicity depon] is on their breakdown in tho body with tho liberation of sorathing hu'iiful ;o the liver. A partial answer has boon obtalnod by feuding #1006 to rats and docs which wore on n low chloride diet with uniform excretion of cl derides in the urine. Y/hon these animals received tho ohloriiu.ted hj'iIroourbon (1006) the urinary ohlorides rose. This indicates that the body certainly hi:s power to detach chlorine from this compound, and it is prohublo that the same condition is true for allied toxic compounds. It would bo most interesting to see whether ingestion of 1260 results in eirJlar findings or whathor in tho case of this relatively non-toxic compound there is practically no splitting off of chlorine, Nolthor time nor our fincnolal resources permitted such tests. NEV 118921 WATER PCB-SD0000030481 -10- k q, THE EFFECT OF INCREASE; TIE SODIUM CIUOlillE IN THIS DIET UPOII AHIMAIS RBCSXVUKJ TOXIC DCSES OF #1006 . On the ground thut clilorides might be fundamentally associated with toxiolty, a froup of 15 rats was fed a low toxic doe of #1006 and coloured with a sirallur group on the sente dosage of #1006 plus a narked increr.no in ohlorlde Inti ko secured by giving 5 oc. per kilogram of body weight of 4 per oent KaCl solution daily. Ho differences wore found botween the two groups, and it nay be oonoluded that chlorine increase secured through the diet does not enhance toxicity. This erperlrK. was done in order to find out whether increase in chloride intake during hot weather might be harraful. A particilur phase of the problem,. the possible enhancement of typieel skin lesions by increasing ehlorido intake, cannot bo decided by experiments on fur-beorlnc animals with no sebaceous glands. All that can be seld at the moment is that increased chloride intake does not increase systemic toxicity. 3. THE EFFECT OF JHGli AHJ) DO'.; CALCJlEi Oil AKBALS lkiiALXKC iiio;: cG::C.r,i:tvj'ic;o c? Jiooc It is well known that a diet rich in calcium is rojrkedly effective in preventing tho acute yellow atrophy of tho liver produced by carbon tfttra- chloride (The ijrevfcjition and Treatment of Curbon Tetrachloride Intoxication. By J>. I), luasorj , K.D., a. S. Minot, Di.D., and B. II. Robbins, M.S., Journal of the African Medical Association, 1920, voliu.-e 90, pugo 34b). . To dir coves- whether calcium. in the diet would preset t^.inut Jive-r tktv.go NEV 1 1 0 9 2 2 WATER PCB-SD0000030482 -11- k from a toxii chlorinated hydrocarbon, 30 adult white rate were placed upon a diet of lean horeeroeat, starch end. lord, a combination adequate for maintenance but Tory lor in oalolun. Another group was given a diet consisting of dog chow, milk, lettuoe and eggs, with added calcium lnotate -- a ration very high in' ealoluia. Both groups were exposed simultaneously to inhalation of high concentra tions of #10)6, an average of 11,SI iqpn. per cubic meter for 16 hourn a day. After IS days, is high calcium diet rats were allvo and 7 of the low calcium grou]>. At intervals animals were killed for examination. In both groupB the liver was abnormal grossly and microscopically, and in both groups animals died from liver damage. It was impossible to consider that the high calcium diet wus in the leuBt degree protective, it may, therefore, be concluded that adding culclum to the diet of workers either in the form of extra milk or of calcium lactate will not prevont liver damage. 4. TOE EW15CT OF INJECTIONS OF XANTHINE Oil ANBiAlB INHALING HIGH COIICENIRATIOIS OF #1006 In 1937, R. 0. Neale published a brief paper (The rrotectlve Action of Certain Purina against liver Necrosis Produced by Carbon Tetrachloride and Chloroform. Science, 1937, voluno 66, pace 63), lie claimed that rats injected with itodlun xanthine became markedly renistent to onrbon tetrachloride. This suggested that xanthine might have similar protective power over liver damage from th< ohlorinnted hydrocarbons on examination in this laboratory. accordingly 36 adult white ratr. were caused to inhale #1006 in concentra tions cveroglnf 15 mgm. por cubic meter for 16 hours daily. Ono group of 0 rats was given 80 mgr., of xanthine oubeutanoously every other day nd 40 rig:... NEV 118923 WATER PCB-SD0000030483 1 f xanthine by stonaoh tube'on the alternate flays. Ifce second group of 18 rats had the same exposure to 1006 without xanthine treatment. No differences were noted and one oannot expeot any effioaoy from xanthine either In the prevention or treatment >f liver disease flue to this chlorinated hydrocarbon, and in all probability tie sans negative result would be enoountered in connection with allied toxio compounds. ' NEV H892* WATER PCB-SD0000030484 i as'S>j': v m. the liver can bet to toimale, but fevl t yellow atrophy &v, r- et tbe only one* 3sj! S3$&$>''% on on the chlollsif ^ -: saturated andW;- ' 'T "d qfcS&'L ufeetationa andw< --- "- group and la-t $|M-r v ` ie of the moat'*- T1'. THE JOURNAL OF INDUSTRIAL HYGIENE ** tWffiB ifgSp^: ne acute action^ f*?:#?-'" portance com^^fe'%'- rs AND TOXICOLOGY s damage to the*! :ially the Uvef/rjlf !^^T::Volume 21 MAY, 1939 Number 5 n. All glycola m*; 5. i, the ethers of |vg%-i lii-ife the eaten, and:"!!* K*iK~ FDRTHER OBSERVATIONS ON THE POSSIBLE SYSTEMIC TOX- ie goee up the f&| &:. .ICITY OF CERTAIN OF THE CHLORINATED HYDROCARBONS ion to the for- . WITH SUGGESTIONS FOR PERMISSIBLE CONCENTRA ercotic effect of ' f ethyl alcohol. ^ >n on Dioxen, 1 been shown to :*n'V Hfii-f- >; stion on both TIONS IN THE AIR OF WORKROOMS* _ Cecil K. Drinker Department of Physiology, Harvard School of Public Health, Boston, Mass. lisulfide, which ^;v ie said to pro- : / catcd clinical S ': , anything can fyjj., INCE 1937, several papers have The Effect of High and Low S appeared dealing with the sys temic effects of certain of the Calcium Intake on Animals In haling High Concentrations of a M: chlorinated hydrocarbons. These are Mixture of Penta and Hexa- practical imin a table the lest allowable airiouas volatile these figures fence given in He does not, ^present more e and experi- listed in the bibliography in the order of their appearance V(1X,, 2, 3, 4, *5/). ^ of these P*Pe ^eluded /- recommendations as to safe concentra- " Hons in the air of workrooms, that is, . concentrations that would cause no ill J;':.: effects to individuals who breathed t them throughout a working day. It also mentioned certain other observa- Hons, such as the effects of diet on toxicity, which were being made at tt*me hut which had not progressed '*ar enough to merit description. It is novr possible to report upon these ` matters and to list a fairly large num- . .. her of chlorinated hydrocarbons to- >Bother with permissible concentrations in air. V-*. - - - |>i : Received for publication February 10, CHLORNAPHTHALENES 62.6%). ........ (CHLORINE It is well known that a diet rich in calcium is markedly effective in pre venting the acute yellow atrophy of the liver produced by carbon tetra chloride (Lamson et al. (6)). To dis cover whether calcium in the diet would protect against liver damage from a toxic chlorinated hydrocarbon, 30 adult white rats, were placed upon. a diet of lean horsemeat, starch and lard, a combination adequate for main tenance but very low in calcium. Another group was given a diet con sisting of dog chow, milk, lettuce and eggs, with added calcium lactate,--a ration very high in calcium. Both groups were exposed simultaneously m 155 WB8 - DEPOSITION EXHIBIT 6 WATER PCB-SD0000030485 I kH 'A A iHj?i!i : ii mi m ri * r i];Sk: il] ? i 156 JOURNAL OP INDUSTRIAL HYGIENE AND TOXICOLOGY M. tl, no. ] to inhalation of high concentrations of a mixture of penta and hexachlornaphthalenes (chlorine 62.6%), an average of 11.21 mg. per cu.m, for 16 hours a day. After 16 days, 12 high calcium diet rats were alive and 7 of the low cal cium group. At intervals animals were killed for examination. In both groups the liver was abnormal grossly and microscopically, and in both groups animals died from liver dam age. It was impossible to consider that the high calcium diet was in the least degree protective. It may, therefore, be concluded that adding calcium to the diet of workers either in the form of extra milk or of calcium lactate will not prevent liver damage. The Effect of Injections of Xan thine on Animals Inhaling High Concentrations of a Mixture of Penta and Hexachlornaphtha- lenes (Chlorine 62.6%). In 1937, R. C. Neale (7) published a brief paper in which he claimed that rats injected with sodium xanthine became markedly resistant to carbon tetrachloride. This suggested that xanthine might have similar protective power over liver damage from the chlorinated hydrocarbons on examina tion in this laboratory. Accordingly 38 adult white rats were caused to inhale a mixture of penta and hexachlomaphthalenes (chlorine 62.6%) in concentrations averaging 15 mg. per cu.m, for 16 hours daily. One group of 20 rats was given 20 mg. of xanthine subcutaneously every other day and 40 mg. of xanthine by stomach tube on the alternate days. The second group of 18 rats had the same exposure to the compound without xanthine treatment. No differences were note|l and one cannot expect any efficacy! from xanthine either in the prevention^ or treatment of liver disease due tol this chlorinated hydrocarbon, and in] all probability the same negative! result would be encountered in coaj nection with allied toxic compounds^ .1 Evidence for the Destruction of ] Mixture of Penta and HexH ACHLORNAPHTHALENES (ChLORINiJ 62.6%) in the Body. ' It has always been a question as whether the chlorinated hydrocarbons] which have been examined by our* selves and by others do harm per 5 or whether toxicity depends on theirJ breakdown in the body with the^ liberation of Something harmful to the g liver. A partial answer has been! obtained by feeding the above com-*^ pound to rats and dogs which were on ^ a low chloride diet with uniform' excretion of chlorides in the urine") When these animals received the ' chlorinated hydrocarbon the urinary.1 chlorides rose. This indicates tbat.| the body certainly has power to de^ tach chlorine from this compound, and ,| it is probable that the same condition is true for allied toxic compounds. At. ' 31 The Effect of Increasing the K' Sodium Chloride in the Diet upon Animals Receiving Toxic ' Doses of a Mixture of Penta * and Hexachlornaphthaleneb (Chlorine 62.6%). On the ground that chlorides might'tj be fundamentally associated with tox icity, a group of 15 rats was fed a low toxic dose of this compound and com- pi pared with a similar group on the j same dosage plus a marked increase in ^ m WATER_PCB-SD0000030486 V OGY tool, ti, no. in TOXICITY OF CHLORINATED HYDROCARBONS 157 ferences were noted 'i xpect any efficacy'?' r in the prevention j rer disease due . 'chloride intake secured by giving 5 cc. per kg. of body weight of 4% sodium chloride solution daily.- *'N<r differ- ' ~enCes were found between the two sebaceous glands. All that can be said at the moment is that Increased chloride intake does not increase systemic toxicity. drocarbon, and uf?i e same negative^.countered in w TABLE 1 A 14List of Chlorinated Htdrocarbonb, with Chlorine Contents and Permissible ALimits (in mo./co.u.) tor the ib in Workrooms* * compounds w f ] Destruction of .k J. -:-.*'*/.* ? ' ' couromm CJXULXOXCONTENT ruuo* mLUCxT ' m>TA AND Hex-^W enes (Chlorine 5 ODY. ....... % j, Trichlornaphthalene plus a trace of tetrachlornaphthalene. Tested #./ *!. , ,-a'"v t n a question as to t~r. tfed hydrocarbons \ upon rats by inhalation and by feeding................................................ 49.9 2. Tetra and pentachlornaphtbalenea. Tested upon rats by inhalation and by feeding........................................................................................ S6.4 10.0 1.0 acamined by our- * 3. Penta and hexachlornaphtbalenea. Tested upon rats by inhalation and - R do harm per depends on their | ' body with the *L? s- v. ! rig harmful to the ^ by feeding, and upon dogs by feeding alone........................................ 62.6 4, Tetra and pentacblornaphtb&lenes plus refined chlorinated diphenyl, Tested upon rats by feeding--............................................................ 43.5 S. 90% penta and hexachlornaphtbalenea plus 10% chlorinated diphenyl 63.0 0.5 0.5 0.5 mswer has been : the above com- 'v Kgs which were on ' 7- it with uniform es in the urine. ils received the rbon the urinaiy s indicates that i_ ;v i', ias power to de- 1 : ' is compound, and -- ie same condition fr Tr i 6. Chlorinated diphenyl plus chlorinated diphenyl benzene. Tested upon rats by inhalation and by feeding......................................................... 65.0 0.5 7. Chlorinated diphenyl oxide. Tested upon rats by inhalation.............. 64.0 0.5 8. Chlorinated diphenyl oxide. Tested upon rats by inhalation.............. 57.0 0.5 9. Chlorinated diphenyl. Tested upon rats by inhalation........................ 50-55 0.5 10. Hexacblor diphenyl oxide plus 5% trichlornaphthnlcne. Tested upon rats by inhalation................................................................................... 50-55 0.5 11. Hexachlornaphthalene and crude chlorinated diphenyl. Tested upon rats by inhalation................................................................................... Un 0.5 known 12. Special chlorinated naphthalene. Tested upon rats by inhalation....... 50-56 0.5 13. Chlorinated diphenyl. Tested upon rats by inhalation........................ 68 10.0 14. Chlorinated diphenyl benzene. Tested upon rats by inhalation........... 60 0.5 IkCREASING THE : in the Diet ;*; eceiving Toxic TUBE OF PENTA /' NAPHTHALENES t chlorides might ociated with toxits was fed a low .pound and comr group on the arked increase in ` --C ( r ' * The analytical method and apparatus used routinely lor field determinations ia that described by Tebbens (This J., IS, 204 (1937)) and by Drinker e( al. (ibid., p. 283). groups, and it may be concluded that chlorine increase secured through the diet does not enhance toxicity. This experiment was done in order to find out whether increase in chloride intake during hot weather might be harmful. A particular phase of the problem, the possible enhancement of typical skin lesions by increasing chloride in take, cannot be decided by experi ments on fur-bearing animals with no The Systemic Effects of 14 Chlo rinated Hydrocarbons with Sug gested Limits for Concentra tions in the Air of Workrooms. The technic for inhalation and feeding experiments upon rats has been fully described (Drinker et al,. (I)). It has been uniform for all the compounds listed in table 1, and for each group of experiments 24 to 48 WATER_PCB-SD0000030487 I sfcess&a if. Ilf- pte m 01 4 158 JOURNAL OF INDUSTRIAL HYGIENE AND TOXICOLOGY M- tl, no; t py' animals have been sacrificed and toxicity because it is not broken down examined grossly and microscopically in the body, but that would seemSfeJi; so as to -gain .very complete post the probable explanation. Compound mortem reports. number 12 was prepared especially in The first 6 substances in table 1 order to have a substance for test have been dealt with in previous pub which should fall in chlorination be-? lished reports (cf. l and 3 in bibliog tween compounds 1 and 2.' It proved M raphy). . In connection with them it somewhat toxic and, in addition to was recommended that in the case of changes in the liver, caused eye ir-EST trichlomaphthalene and compounds of ritation and in one instance cata-|? lower chlorination a limit of 10 mg. per cu.m, be permitted, the idea being racts. The substance is not mercial use and evidently in com at the . Jgfe: ? ^Si- that the essential feature of the action temperatures used for volatilization of these compounds, namely damage gave off some sort of irritant, possibly to the liver, did not become prominent hydrochloric acid, ......... -'STfr/- until a chlorine content of 50% was With the exception of this isolated |g.S reached. So far as trichlomaphtha effect upon the eyes, we have seen no lene is concerned the limit of 10 mg. lesions from any of the 14 compounds r should be allowed to stand, since listed except in the liver. The char nothing has occurred which indicates acter of these lesions has been thor mk the necessity for changing it. The oughly described by Bennett et al. second compound, a mixture of tetra (3). These additional data and the and pentachlomaphthalenes is in com tolerances recommended deal_ spe mon industrial use. A tolerance of cifically with systemic effects since one 0.5 mg. per cu.m, was formerly pro can get no information as to chloracne posed for it, but on the basis of labora in the skin from observations upon tory and field experience it is fur-bearing animals. But it may be apparently entirely safe to increase this to 1.0 mg. per cu.m. The sixth compound has been listed previously as chlorinated diphenyl. It contained 65% of chlorine and proved very destructive to the liver. Later experiments with compound 13, pointed out that where industrial workers have been careful as to clean liness and where air concentrations have been kept within the limits sug gested, skin lesions apparently cease to occur. - which contained 68% of chlorine and which was also labelled chlorinated diphenyl, were a surprise to us since this second compound was almost non-toxic. On inquiry it was found that substance 6 was in reality a mixture of chlorinated diphenyl and chlorinated diphenyl benzene and that number 13 was actual chlorinated diphenyl. 'We have no information as to whether this last compound lacks Summary - Certain experiments- carried but upon large groups of white rats indi cate the following: ' 1. A high calcium diet has no pro- _ tective effect against the characteristic liver lesions caused by these com- pounds. . |j|s?r 2. Administration of xanthine does not prevent liver injury. W>' WATER PCB-SD0000030488 il. Si, no. S Seen down mid seem 'ompound >ecially in : for test ation be lt proved Idition to i d eye Jr-, ice cata-f- VV^ ; in com- at the'y&3 itilitation *`$3 , possibly i isolated , e seen no capounds ' rhe char- sen thortt et al. and the j. eai spe, since one ; 'hloracne : os upon may be ndustrial to dean- ,/f ntrations nits sug- ; Jy cease _ I ied out. . ats indi- | no pro- cteristic "j ( se com- ' k WATER PCB-SD0000030489 { : VaL 154, No. 17 CHLORACNE--MEIGS ET AL. 1417 ,-tents. Thyroid function may be depressed in a higher r percentage of cases if urethane therapy is maintained for longer period. This possibility is currently being in- 41gated. ~ SUMMARY (Tofwuernettyhapnaet.ieTnhtse were treated with rectal suppositories therapeutic effectiveness of rectaily ad ministered urethane is equivalent to that of orally admin istered urethane. Urethane administered rectaily is as effective as Fowler's solution in spacing irradiation ther apy. Undesirable side-effects were minimal. It is con cluded that rectal administration of urethane is preferable to oral or intravenous administration. 2065 Adelbert Rd. (6) (Dr. Weiiberjtr). CHLORACNE FROM AN UNUSUAL EXPOSURE TO AROCHLOR /. Wister Meigs, MJ3., Jack Jonathan Albom, MS) ." end Bernard L. Kartin, MJ>. New Haven, Conn. The first outbreak of acne-like lesions due to high This paper reports the development of lesions of chlor boiling chlorinated compounds in industry in this coun acne in seven workers employed in a chemical plant con try was noted by Schwartz1 in 1936. Subsequently, dur cerned with organic chemical production. ing World War II, various reports of acne-like lesions in workers exposed to certain chlorinated naphthalenes and diphenyls followed. Collier * reported 12 cases of chlor- HISTORY OF EXPOSURE A chemical company had for some months been using acne of the face in workers exposed to the fumes or dust molten salt at 330 F as a medium for supplying accu of chlorinated naphthalene and one fatal case due to rately controlled quantities of heat to a large jacketed acute yellow atrophy of the liver. Fifty-five cases of acneform dermatitis were reported by Kelley: in 200 persons exposed to chlorinated naphthalene (Halowax). Good and Pensky * described 32 cases in electricians, who reaction chamber. Because of the dangers of solidifica tion of the salt in the return line, as well as the corrosion problem, it was decided to use a chlorinated diphenyl (Arochlor) as the heat exchange material. The same handled the cold-finished product for the most part. In apparatus was used. This included an oil-fired furnace none of their cases was systemic involvement found. containing the heating coils, a steel pipe supply line to the hwartz's report concerned the involvement of elec- reaction chamber, a return line to a large sump pump, jidans, who installed and stripped wires in ships during and an outflow line from the sump to the furnace (see the war. The chlorinated naphthalene, which was im figure). The reservoir for the sump pump had a capacity pregnated into the asbestos and wrapped around the wire of about 400 gal. (about 1,680 liters). * as insulation, flaked off in the stripping process. In the 'cases described, two months elapsed before the appear ' ance of the chloracne of the face. There was no systemic involvement. Peck,a Cranch,7 and Greenburg * discuss Jthe chlorinated naphthalenes and chlorinated diphenyls, . The chemical product, designated as organic acid A, was manufactured with the use of molten salt for heat exchange from January to December, 1949. At that time, chlorinated diphenyl was substituted as a heat exchange material. It was soon apparent that under certain condi tions there was slight leakage of vapors from a number Jhe appearance of the dermatological lesions, their value of places, particularly around the cover of the sump, and ^in industry, and precautions in handling these chemicals. also from all gasketed connections in the system. Because .v The value of the chlorinated naphthalenes and di- of the known toxicity of these substances, the assistance Iphenyls in industry is due primarily to their resistance to j water and alkali, high insulating value (high dielectric From the departments of public health sad Internal medicine, Yale University School of Medicine. 'constant), thennoplasticity, chemical stability, and flame ^resistance, as described by various writers.* Associate Professor of Occupational Medicine, Department of Public Health (Dr. Meip). Assistant Clinical Professor of Dermatolop. Depart ment of internal Medicine (Dr. Albom), and Assistant Clinical Professor * * The characteristic lesions of chloracne are pinhead to 'pea-sized pale straw-colored cysts formed by the plug of Medicine. Department of internal Medicine (Dr. Kanin), Yale Uni versity School of Medicine. 1. Schwartz. L-: Dermatitis From Synthetic lobs and Warn, As. J. ging of the orifices of the sebaceous glands, resulting in 'retention of the secretion and in the keratinization of the Pub. Health 28:316 (June) 1936. 2. Collier, E.: Poisoning by Chioriaated Naphthalene, Lancet is 72 (lan. 16) 1943. . fining membrane. Comedones are present but are not a ^striking feature. In nearly every worker exposed suffi 3. Kelley, . F.: Acne From Synthetic Wu (Halowax), UroL A Cutan. Rev. 47:239 (April 1) 1943. 4. Good, C. 1U and Pensky. N.: Halowax Acne ("Cable Rash**): ciently to these chlorinated compounds for a few months these lesions will develop. The exposure may be either to Cutaneous Eruption in Marine Electricians Due to Certain Chlorinated Naphthalenes and Diphenyls, Arch. Dermal 4 Syph. 48:231 (Sept.) 1943. fumes from the hot material or to the solid material on 3. Schwartz, L.: Outbreak of Halowax Acne ("Cable Rash'*) Among Electricians, i. A. M. A. 122:131 (May 13) 1943. '.continued contact. Repeated or continuous contact is " 'sential. Lesions have not been reported after short or 6. Peck, S. M.: Dermatitis from Cutting Oils, Solvents, and Dielectrics, Including Chloracne, J. A. M. A. 12S: 190 (May 20) 1944. 7. Cranch. A. G.: Chlorinated Compounds: Precautions In Handling, -drequent exposures. Vesiculoerythematous eruptions, f ' *s seen in acute eczematous contact-type dermatitis and lodusL Med. 13:110 (Jan.) 1944. I. Greenburg, L.: Chlorinated Naphthalenes and Diphenyls, Indusc. Med. 12:320 (Aug.) 1943. | ..simple erythematous eruptions with pruritus, have also 9. () Mayers, M. R., and Silverberg, M. G.: Skin Conditions Result ing from Exposure to Certain Chlorinated Hydrocarbons. I. Indust. Hyg. 4 Toxicol. 20: 244 (March) 1931. (b) Schwartz.* (c) Greenburg* WATER PCB-SD0000030490 i 1411 CHLORACNE--MEIGS ET AU J-AAI-A*, April 24, IH4 of the Bureau of Industrial Hygiene, Connecticut State counts and unnalys'u were normal in all instances. An Department of Health, was sought and received. A field _ had normal blood pressures with no other Hini/-^ ^ study under conditions of obvious vapor leakage was said ' dcnce of any chlorinated diphenyl toxicity. to have shown negligible air concentrations of the chlori After the recognition of these cases of chloracne, an nated diphenyls in the actual breathing zones of the but one of the gasketed joints in the heat exchange sys workers. In this study, which was four months prior to tem, including the cover of the sump, were welded to the dermatological findings reported here, the air con gether. A hand hole 6 in. (15 cm.) in diameter was left centration of chlorinated diphenyls was reported by the with a gasketed cover so that the system could be drained bureau to be 0.1 mg. per cubic meter of air. The rec or filled when necessary. After that time no vapors were ommended maximum allowable concentration is 1.0 mg. visible, and the odor of chlorinated diphenyls was barely percubicmeter. The figure shows that most of the leakage detectable in the immediate vicinity of the sump pump. was at points outside the building but under a roof. No Continued careful observation of workers has revealed one worked regularly at the points of leakage. Neverthe no new cases of chloracne. less, repeated attempts were made to control vapor leak age, without complete success. This operation continued for 19 months without incident or recognition of skin or other manifestations of exposure to chlorinated diphen yls. Each employee had a complete physical examination by an internist prior to working in this environment. COMMENT The unusual feature of this outbreak of dermatitis was the long period of exposure before any cases were recog nized. The sudden recognition of seven cases after expo sure up to 19 months was due to the especially careful examination of the skin Of all exposed employees after discovery of the first case. Of 14 men exposed or poten tially exposed to the vapors of chlorinated diphenyls, 7 presented clinical evidence of chloracne. There was not a very good correlation between the apparent degree of exposure and the development of signs of disease. For example, a foreman, an assistant foreman, and a plant superintendent whose duties would appear to have ex posed them only incidentally to the toxic agent had mild to moderate signs. The mean length of exposure of those in whom signs developed was 14.3 months and of those who did not show signs was 11.4 months, but there was considerable overlap, with chloracne developing in one worker after only 5 months in contrast to another who showed nosigns even after 19 months' exposure to vapors. Since the manifestations were exclusively on exposed areas of the skin, it appears that the vapors were de posited directly on exposed skin and did not go through the clothing. The nature of each factor determining the Schematic drawini of the heat exehante tyvtem ia which the chlorinated diphenyl was uaed tod the points at which workers were exposed to fumes. appearance or nonappearance of lesions is not dear. Skin pigmentation may be a factor. Three of the workers were Negroes, and none of them had chloracne. No correlation with perspiration could be made, but all cases were dis An operator making organic acid A ("operator," fig covered toward the end of the summer. ure) was sent to one of us because of acute contact der Prevention consisted of controlling the leakage of matitis of the face. In addition to the contact-type of vapors. The fact that tests of the air, even in the presence dermatitis, there were noted pinhead-sized straw-colored of vapors, showed only negligible amounts of chlorinated cysts and comedones on both cheeks and the forehead. hydrocarbons indicates that this type of intermittent but A diagnosis of chloracne was made, and the source of fairly long continued "mild" exposure is not innocuous. exposure was determined. Examination of other workers in this working environment uncovered six additional The low concentration of the chlorinated diphenyl in the air might account for the fact that lesions developed u> cases. In all the face was involved especially the cheeks only 50% of those involved. circumorbitaily, the forehead, ears, and in one case the SUMMARY l mastoid region. All employees were examined carefully Seven cases of mild to moderate chloracne of the face by an internist. The seven employees in whom chloracne and head occurred among 14 chemical operators ex : had developed had liver function tests performed. Tests posed from 5 to 19 months intermittently to small con included direct and total bilirubin determinations and 24 centrations of the vapors of a chlorinated diphenyl (Aro- and 48 hour cephalin flocculation, thymol turbidity, and chlor). Leakage of these vapors from a heat exchange alkaline phosphatase determinations. Six of the subjects system occurred chiefly outdoors, but chloracne was o had completely normal test results. One employee had served among men working inside the adjacent building- borderline cephalin flocculation and thymol turbidity. Inall cases the condition cleared up after treattneo- Thirteen months later repeated liver function tests Control of vapors by welding all joints in the beat e II showed an unchanged cephalin flocculation and im proved thymol turbidity. Results of complete blood cell change system prevented recurrences. 310 Cedar St. (Dr. Meigs). WATER_PCB-SD0000030491 COPY Monsanto Chemical Company St. Louis -4, Missoubi April 28, 1934 COPY I Dr. J. Wiater Meigs 310 Cedar Street ' Mew Haven, Connecticut ' Dear Dr. Meigs: , Your article in the current number of the JAMA is certainly confusing to me. We have had men exposed to concentrations of ohlorinated diphenyl four or five times what you have recorded for periods of five to ten years and have had no chloracne. Could you tell me which Aroclor was used? I notice you have referred to the product they manufactured as organlo acid A. Are you at liberty to tell me what that product was? If you are not, does it con tain any chlorinated compound or is any chlorinated compound used as an intermediate? Is organic acid A the only product this plant manufactures? And if not, . are any other chlorinated compounds, such as penfcachlorophenol, 2,4,5-T, 2,4-D, or DDT manufactured? Very truly yours. REKiMPL R. Emmet Kelly, M.D Medical Director 1 DEPOSITION i EXHIBIT I^ NEV 160086 WATER PCB-SD0000030492 i SECTION OF OCCUPATIONAL HEALTH ' V , 4- YALE UNIVERSITY SCHOOL OF MEDICINE DEPARTMENT OF PUBLIC HEALTH jio CEDAR STREET NEW HAVEN u, CONNECTICUT ,. H. Emmet Kelly, M.S. ,: Medical Director i . Monsanto Choaieal Coapany | it. Louis 4 Missouri Sby T, 1954 ' ' . \ * ' `_ ! Dsar Dootor Csllyt i Thanlc* for your lottor of April 28th. Our observations ; of ohlaraono (mostly quit* mild) wore onfusing to aa too. That ! was on* roaaon for publishing tho data, aino* there is a tendenoy ' aacng all of us to assumethat certain conditions oan bo characterised : as "safe" on tho basis of onTironasntal studios alono. Rons of us . . horo (including tho State Health Department Survey people) have any | good answer to why the air samples were so low. We assume that i somehow other conditions may have existed for a few weeks -or months . prior to the recognition of ehloraoso. . Organic aoid A did not oontain chlorine, nor did anything else la the particular building. However, the product may have played a part, although by itself it aots.only as an irritant (it is paokaged as a powder). The Arcshior was from one of the large suppliers and was a combination of high boiling ohlcrinatod hydrocarbons. 8laos the repairs on the circulating system there hare been no more oases - over a three year period. My conclusion is that chlorinated hydrooarbons should ultimately be judged aa safe or hazardous in relation to observations of workers themselves. Study of the environment would seem at best to be only a guide/. One of our purposes was to stimulate interest in oooupatlonal medieine. . X hope the article will not cause a rash of claims. ' It seemed to us that the diagnosis of ehloraenc was so spsoifio thst any competent dermatologist oould make it. Therefore, disputed oases should not bo frequent. On the other hand, if even mild ehloracne is occurring, management representatives would want their doctors to be alert to it, at. it would seem to be a preventable condition. DEPOSITION EXHIBIT _a_ \ Finally, X an not suggesting that the n.a.e, for Aroolor has been set too high. Xt will take years of observation and experience to determine this. Our "outbreak* may have been an isolated experience not likely to be duplicated. *n that oosneotion, it would be helpful and informative if you could publish your experience on exposures over long periods to these substances. We need many more reports to hasw a basis for sound judgsment. With best rsgards to you and Sltmar Wheeler. NEV 160085 Sinoerely yours, JEMinf* . /. Winter r.sige,'M.D. /tsor.Ssfctt iVc '"tor of WATER PCB-SD0000030493 1474 S. Vandhventbr Avb. St. Louis 10, Mo. Moresfcer 10, 1953 CERTIFICATE OF ANALYSIS oa;; SUBJECT - The Amsts Oral Toxieity (LDg9) of Aroelor i2$U Ptor Rata. TEST CONDUCTED FOR - / Monsanto Chemical Company, St. Louia, Missouri. Monsanto Project No.i SA-15 EJPLRIMEI.TAL PROCEDURE - . `S The LDeo was determined by the introduotion-ofgseaaured single doass of undiluted Aroolor 1251* into the stomaoha of Sprague Dawley strain albino rats by an ana of a rubber catheter attached to a hypodermio syringe. Animals weighing 17$ to 250 grams ware used in the teat. After the approximate Minimum Lethal Dose was found, groups of animals were fed at levels designed to blanket the toxicity range thereby supplying dsta for calculation of the L50q which was done aocording to a modification of the method of Bliss (Quart. J. Pham, and Pharwaool., 11, 1938). The data based on results obtained from a total of 1*1 rats are found in Table I. ' . 4 ' Selected animals from those receiving a lethal dose were autopsied and maorosoopio examination made of the vlsoera. 2 DEPOSITION EXHIBIT _JO_ NEV 017288 Thu a a confidential report which may he wed when requeued by pbyticiim and health oificiiU, but it not to be used in any form of advertiainK without written pennUoon. CONSULTANTS TO THE FOOD, CHEMICAL AND PHARMACEUTICAL INDUSTRIES WATER PCB-SD0000030494 . I. TOi Nonaanto Char' "al Co^>aay St. Louie, tt jovrl , Soiantifie Associates Cartifissta of Analysis -- Pays 2 (11-10-53) TABLS I TH5 LDBo OF ARQCLOR 1254 FCR RATS ilmal Mo.- Sex 1- Mala 2- Male 3- Mala 4- Female 5- Female 6- Mala 7- Female 0- Female 9- Female 10- Male 11- Mela 12- Mala 13- Female 14- Female 15- Female 16- Mala 17- Mala 18- Mala 19- Mala 20- Female 21- Female 22- Female 23- Male 24- Mala 25- Male 26- Mala /' 27- Female 26- Female/' 29- 111/ 30- Female 31- Fertile . 32- Female 33- Mala \ 34- Mala 35- Mala 36- Female 37- Female 38- Female 39- Male 40- Mala 41- Male Weight Orame -- "1 - 225 115 240 185 195 220 200 175 190 230 250 215 205 190 180 210 230 240 _ 225' Dose KU/Kg. 2.50 3*00 /X \ 3.75 Fata_ Survived Died A Survived Survived \ \ Survived . \ Died .A' \ Died \ Survived Survived Survived ` Died ' Didd Survived Survived Survived Died Survived Survived Died Survived Died Died Survived Survived Died Died Survived Died Died Tied Survived Died Died Survived Died Survived Died Died Died Died Died I NEV 017289 WATER PCB-SD0000030495 TO i Nmsanto Cbm. .al Company ' St. Lodit MLsMQrl Soisntifio Assosiatss Csrtiflsats of Analysis -- Pag J (11~I0~$3) I DISCUSSION - Tho LDkm mu oaloulatsd to bo 3.10 sdllilitara par kilogram with uppar and loMir limits of 2.98 to 3.2U millllltsra par kilogram. Surriwal tins ranged Area 1 to 6 days with ths majority of daatba occurring in 2 to 3 days. Thsrs saa modarats diarrhea, lethargy, and poor appstito in all of ths animals. At autopsy ths liver varied from a straw oolor to dark rdd in diffsrsnt viaoara. Ths intestinal treat was irritatsd with muoup praasnt in several instances. - \A SUKKART - Ths LDg0 of Aroolor 1251* for rats kilogram. Fiduoial limits wars ~ d\to fep 3.10 millilitors par 21pmil^Liiltsra par kilogram. // f/ //%.\ V ( \ \ sci^tifF:'JC ASSOCIATES \ BYi FRED M. YOUNGER NEV 017290 WATER PCB-SD0000030496 c^>cusiii^ic cz?J-**0ciate4- 1474 S.Vandbvbntba Av. St. Loin* 10, Mo. Beoeiriber 11, 1?5>3 CERTIFICATE OF ANALYSIS KHKI - ^ The Arsis Oral Toxicity (1S^9) ef Areola^ ltl^Wr Bats. fXST OOXBOCttD m - - Hwttt* Gbmisal Orapasy, ft. Leals, MihuI, Jfeaasate Prefect Ss.i BA-19 Mi&state Staple fs.i let Me. 1M zxrammL psookdoss - the Aovte Oral Texisity (U^g) m fend by the latraduction ef muv*4 Slagle dotes ef nsdilnted Arooler lato the atoaaebe ef fpragee Cawley strala albl&e rats by saas ef a rubber oathster attached to a hypederslo syringe. After the texlelty raage was fenad, groups of aalaels sere dosed at lerals designed to blanket the toxlelty raage thereby ruyylyiag data for eel- eolation ef the ID,, which was done according to a nadlfleatlen of tbs atethod of Bliss (Quart. J. Pham, aad Pharnaeel,, 11, 1931), Beloeted aalaala froa those roeeielag a lethal dese sere astopsled aad saareaeepie oxaalaatlon sad* of the Tlsoora. The data acre gluon la fable I. DEPOSITION EXHIBIT 4t Tbb b w**"*bl raport wfcfcb my b* und wiwn nquotod by pbyaOMa tad health oScbb. but b oot to b, und la toy fora wl Klvtrtuiii* without wripw pmbitoa. NEV 017291 rnwiULTANT* TO TH> 'OOP, CHIMICAL AND t H A % M A C BUTIC A L INDUSTltlS WATER PCB-SD0000030497 It* Leals, Me*. eri Scientific Aasoci*. -8 Certifl c*t'of Analysis Fa,,< 2 (12-11-53) TABLE 1 THK ACVTF. ORjU. TOXICITY (IJ^) OK AKWL-^R li'iii: FOR RATS ilmal No.- Sex 1- Female 2- Female 3- Feral e i- Ka.le 5- Kale 6- Female 7- Kale 8- Kale 9- Kale 10- Female 11- Female 12- Kale 13- Female 11*- Kale IS- Kale 16- Female 17- Kale 18- Male 19- Female 20- Female 21- Female 22- Male 23- Kale 2U- Male 25- Female 26- Female 27- Female 20- Kale 29- Kale 30- Male 31- Male 32- Female 33- Female 3l*- Female 35- Male 36- Kale 37- Kale 38- Male 39- Female 1*0- Female 1*1- Female 1*2- Female 1*3- Female 1*1*- Female .j , f*rnjns 185 170 190 230 215 200 210 235 21*0 195 165 21*5 220 215 235 100 190 225 210 230 175 220 2l*0 21*5 200 185 190 230 220 250 195 210 180 175 220 230 200 21*0 185 200 175 180 190 175 Lone Ml./Kg. ?.?i 3.7? U.2? 1*75 Fate Survived Survived Survived Died Died Survl ved Bled Survl red Died Survived Survived Survived Died Died Survived Survived Survived Died Survived Survived Died Survived Died Died Survived Survived Survived Died Survlvod Survived Died Died Died Died Died Survived Survived Died Died Died Survived Died Died Died NEV 017292 k WATER PCB-SD0000030498 Ttt fteaaante Chead... ,4. Coapany . . >. 8t, Louis, W.5 iri ' Soientlflo tepociates' Certificate of Analyst* -- Pa** 3 (12-11-53) I DISCUSSION Th* LD,0 i calculated to be 25 nillillters p*r kiloera with fiducial liait* of Iu0?*ii36 sd Hilltars par kilograa. Survival tins for tboaa aniaals reDelving a lathal doao varied froa 12--51* hours. Lethargy developed in tha Majority of animal* a fsw hours after dosing. There was also soderate diarrhea. At autopsy, the liver was dull red with a greenish hue while the spleen was abnormally dark. The kidneys appeared normal by aaoroeeeplo exasdnatiea. 8UXKAKI - The LDq0 of Aroclor 12l*2 was found to be it.15 aillilitsr* per kilogram with lever and upper Halts of l02-lj.36 adlllllters per kilo gram. SCIENTIFIC A5S0CIAT1S Bit FRED M. TOONQSR NEV 017293 WATER_PCB-SD0000030499 I WATER PCB-SD0000030500 Sequence of Tables X. The Mortality, Length of Survival, and Changes in Weight of Animals Exposed to the Vapor of Aroclor 1242 in Air. Experiment No* 1. 2. The Mortality, Length of Survival, and Changes in \7eight of Animals Exposed to the Vapor of Aroolor 1242 in Air* Experiment No* 2* 3* The Mortality, Length of Survival, and Changes in Weight of Animals Exposed to the Vapor of Aroclor 1254 in Air. Experiment No. 1* 4. The Mortality, Length of Survival, and Changes in Weight of Animals Exposed to Conditioned Air Containing No Vapor of Aroclor 1242 or Aroolor 1254. Experiment No. 1. 5. Summary of the Data on the Mortality Among Groups of Ex perimental and Control Animals. 6a The Average Changes in Weight of Experimental and Control' Animals. 7. The Relationships of the Weights of Certain Organs to the Body Weights of Animals Exposed to the Vapor of Aroclor 1242 or Aroolor 1254. . .t . 3. Summary of the Data on tffe'Re1ationship of the Weight of the Liver or the Kidneys to the Body Weight of Animals Exposed to the Vapor of Aroolor 1242 or Aroclor 1254. 9. The "Prothrombin Time" (Method of Kato) of the Blood of Animals Following Their Exposure to the Vapor of Aroolor 1242 or Aroolor 1254 in Air. . . 10. The "Prothrombin Time" (Method of Kato) of the Blood of - Animals Following Their Confinement in Cages Supplied with Conditioned Air Containing No Vapor of Aroolor 1242 or Aroolor 1254. 11. The Effeot of the Exposure of Cats end R&hits to the Vapor of Aroolor 1242 or Aroclor 1254 in Air for 7 Hours per Day on 5 Hays per Week Over a Period of Several Months, Upon the Apparent Prothrombin Content (Method of Kato) of Their Blood. NEV 016553 WATER PCB-SD0000030501 I ii 12. The Effects of the Intermittent Inhalation of the Vapor of Arodor 1242 or Aroclor 1254, or of Conditioned Air, upon the Numbers of Erythrocytes and leucocytes and upon the Concentration of Hemoglobin in the Peripheral Blood, of Animals. 13. The Average Numbers of Erythrocytes and Leucocytes and the Average Concentration of Hemoglobin in the Peripheral Blood of Experimental and Control Animals. 14. Application of the "t" Test to the Hematological Data Ob tained in Observations on Exposed and Control Animals. NEV 01655^ 1 WATER PCB-SD0000030502 iil Sequence of Figures As Equipment for Volatilizing Aroclor, Chamber and Equipment for Combustion and Colleotion of Samples Bs Front View of 6CQ-Liter Chamber. C* Humidifier for Furnace Sampling Towers. D. Sailing Towers. E. Standard Curve for the Determination of Aroclor 1242 and Aroclor 12p4. 1. Weight of Cat A-446 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air ' 6.6 Micrograss per Liter (Experiment No. 1). 2. Weights of Individual Guinea Pigs B-331 to B-336 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - 6.6 Micrograms per Liter (Experi ment No. 1) 3. Average and Range of Weight of Guinea Pigs B-331 to B-336 _ Recorded at Semi-weekly Intervals During Exposure to the ' Vapor, of Aroclor 1242 in Air - 6.6 Micrograms per Liter' (Experiment No. 1), 4. Average and Range of Yfeight of Mice B-251 to B-260 Re corded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - 6.6 Miorograms per Liter (Experiment No. 1). 5* Weights of Individual Rats L-436 to L-440 Recorded at Semi-weekly. Intervals During Exposure to the Vapor of Aroclor 1242 in Air - 6.6 Micrograms per Liter (Experi ment No. 1)e 6. Weights of Individual Rats L-44l to L-445 Recorded at Semi-weekly Intervals During Ejj> osure to the Vapor of Aroclor 1242 in Air - 6,6 Micrograms per Liter (Experi ment No. 1). NEV 016555 WATER PCB-SD0000030503 I l lv 7 Average and Range of V/eight of Rats L-436 to L-445 Re corded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - $.6 Mierograns per Liter (Experiment No# l) . d. Weights of Individual Rabbits 1-374 to 1-377 Reoorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - f$.6 Micrograms per Liter 1Ex periment No. 1). 9. Average and Range of Weight of Rabbits 1-374 to 1-377 Recorded at Semi-weekly Intervals During Exposure to the Varcr of Aroclor 1242 in Air - Mierograns per Liter (Experiment No. 1). 10. Weight of Cat A-445 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air 6.{ Uicrogrens per Liter (Experiment No. 2). 11. Weights of Individual Guinea Pigs B-325 to B-32g Re corded at Seni-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - 6.S Micrograms per Liter (Experiment Ho. 2). 12. Weights of Individual Guinea, Pigs B-329, B-330 and B-419 Recorded at Semi-weekly.Intervals During Exposure to the Vapor of Aroclor 1242. in Air - 6.{$ Micrograms _ per Liter"(Experiment No. 2). 13. Average and Range of Weigh* of Guinea Pigs B-325 to B-330 and B-419 Recorded at Semi-weekly Intervals Dur ing Exposure to the Vapor of Aroclor 1242 in Air - 6,6 Micrograms per Liter (Experiment No. 2) 14. Average and Range of Weight of Mice B-24l to B-250, \.\ B-376 and B4o4 Recorded at Semi-weekly Intervals Dur"" ing Exposure to the Vapor of Aroclor 1242 in Air - 6,3 Micrograms per Liter (Experiment No. 2). 15. Weights of Individual Rats L-526 to L-530 Recorded at Semi-weekly intervals During Exposure to the Vapor of Aroolor 1242 in Air - 6.B Micrograms per Liter (Experi ment No. 2) NEV 016556 WATER PCB-SD0000030504 t 16. Weights of Individual Rats L-531 to L-535 and L-^6 Re corded at Semi-weekly Intervals During Exposure to the Vapor of Aroolor 1242 in Air - 6.(j Micrograms per Liter (Experiment Ho. 2) 17. Average and Range of Weight of Rats L-526 to L-535 and L-^36 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroolor 1242 in Air - 6(S Micrograms per Liter (Experiment No. 2) 16. Weights of Individual Rabbits 1-370 to 1-373 and I-964 Reoordsd at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1242 in Air - 6, El Miorograms per Liter (Experiment No, 2), 19. Average and Range of Weight of Rabbits 1-370 to 1-373 and 1-964 Recorded at Semi-weekly Intervals During Ex posure to th9 Vanor of Aroclor 1242 in Air - 6t$ Micro grams per Liter lExperiiant No. 2). ` 20. Weight of Cat A-444 Recorded at Semi-weekly Intervals During Exposure to the Vanor of Aroclor 1254 in Air 5.4 Micrograms per Liter "(Experiment No. 1). . 21. Weights of Individual Guinea Pigs B-319 to B-322 Re corded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254 in Air - 5*4 Micrograms per Liter (Experiment No, 1). 22. 'Weights of Individual Guinea Pigs B-323, B-324, B-415 and B-4l6 Recorded at Semi-weekly Intervals During Ex posure to the Vapor of Aroclor 1254 in Air - 5*4 Micro grams per Liter (Experiment No. 1). 23. Average and Range of Weight of Guinea Pigs B-319 to B-324, B-415 and B-4l Reoorded at Semi-weekly Inter vals During Exposure to the Vapor of Aroolor 1254 in Air 5*4 Miorograms per Liter (Experiment No. 1). 24. Average and Range of Weight of Mice B-231 to B-240 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroolor 1254 in Air - 54. ^icrograma per . Liter (Experiment No. 1). SSSS NEV 016557 I WATER PCB-SD0000030505 25. Weights of Individual Rats L-299 and L-517 to L-520 Re corded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254 in Air -5*4 Micrograns per Liter (Experiment No* 1), . 26. Weights of Individual Rats L-521 to L-525 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254- in Air - 5*4- Miorograms per Liter (Experi ment No. 1}* _ 27* Average and Range of Weight of Rats L-299 and L-517 to L-525 Reoorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254- in Air - 5*4- Micrograms per Liter (Experiment No. l). 2>. Weights of Individual Rabbits I-362 to 1-364- Reoorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254- in Air - 5.4- Micrograos per Liter (Experi ment No. 1). ' 29. Weights of Individual Rabbits 1-453, 1-567, 1-716 and1-667 Recorded at Semi-weekly Intervals During Exposure to the Vapor of Aroclor 1254 in Air - 5*4 Micrograms per Liter (Experiment No. 1). 50. Average and Range of Weight of Rabbits I-362 to 1-364,. I-l|r53, I-567.1-713 and; I-667 Recorded at Sami-weekly *... . Intervals During Exposure to the Vapor, of Aroclor 1254 in Air - 5.4 Micrograms per Liter (Experiment No. 1) 31. Weight of Oontrol Oat A-435 Recorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Air. . 32. .Weights of Individual Control Guinea Pigs B-313 to B-316 .. Recorded-et. Semi-weekly Intervals .During the Period of ' Coafifiement in Conditioned Air** - v.: _. 33* Weights of Individual Control Guinea Pigs B-317> B-316, B-402 and B-576 Reoorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Air. 34. Average and Range of Weight of Oontrol Guinea Pigs B-313 to B-316, B-402 and B-576 Recorded at Semi-weekly In tervals During the Period of Confinement in Conditioned Air. NEV 016556 WATER PCB-SD0000030506 vii 35* Average and Range of Weight of Control Mice B-221 to B-2J0, B--499* B-531* B-532 and B-533 Recorded at Semiv/eekly Intervals During the Period of Confinement in Conditioned Air* 36, Weighta of Individual Control Rats L-507 to L-512 Re corded at Semi-weekly Intervals During the Periods of Confinement in Conditioned Air* 37* Weights of Individual Control Rats L-513 to L-516 and L-743 Reoorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Air. 3d. Average and Range of Weight of Control Rats L-507 to L-5l6' had L-746 Reoorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Air. 39. Weights of Individual Control Rabbits 1-365 6114 1-366 Reoorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Alr.v 40. V/eight3 of Individual Control Rabbits 1-366. I-369 and 1-575 Recorded at Semi-weekly Intervals During the Period of Confinement in Conditioned Air. Ij-l. Average and. Range or Weight of Control Rabbits I-365 to I-369 and 1-975 Dud. ng the Period of Confinement , in Conditioned Air. - NEV 016559 WATER PCB-SD0000030507 I The Toxicity of the Vapor of Aroclora 124-2 and 1254- Soope The purpose of this investigation was to determine the effects upon animals of various species, of moderately pro longed intermittent exposure to air bearing the vapor of either Aroclor 124-2 or Aroolor 1254-. This report deals with the re sults obtained in experiments in which, for periods ranging up to 4- months, animals were subjected to respiratory exposure to air containing the Aroclors in concentrations that approached saturation and so were greater than any likely to be encoun tered in industry* The animals failed, to exhibit any signs of intoxication during or after the exposure to which they were subjected, and the numbers of deaths which oocurred among the exposed animals v/ere oomparable to those which resulted from incidental causes in the control groups. Gross and microscopio examination of the tissues of the animals of both the exposed and the control groups disclosed the existence of certain abnormalities-which did not differ significantly in -their severity or frequency of occurrence among the test end control groups* Other experiments which involve more pro longed exposure on the part of animals to lower concentrations of these materials in air, are now in progress* I- NEV 016560 WATER PCB-SD0000030508 Summary of Results 1. No signs of intoxication ware observed in any of the ' numbers of a group of animals, all of which survived through out a period of 24 days, on eaoh of 1J of vfcich they were sub jected to the inhalation of air bearing 6.6 micrograms of Aroclor 124-2 per liter (0.$3 ppm) for 7 hours. The viscera of representative animals that were examined microscopically were normal. 2. The incidence of mortality among the animals of all species, excepting the rabbits, in 2 groups that were sub- . jected (7 hours on each of 62 or 63 days over a period of 120 or 121 day3) to the inhalation of air bearing, respectively, Aroclor 124-2 in the concentration of 6.63 .micrograms per liter (0.66 ppm), and Aroclor 1254- in that of 5*40 micrograms per liter (0.4l ppm), was no greater than that among a similarly constituted control group. Two of 5 rabbits died lb1lowing exposure to the vapor of Aroolor 124-2, as did 3 of 7 rabbits exposed to the vapor of Aroolor 1254, whereas only 1 of 5 of the control rabbits died. The survivors, with the exception of the guinea pigs exposed to the vapor of 1254, grew equally as well as did the controls. The average weights of the livers and kidneys (relative to body weights) of the animals exposed to Aroolor 1242, did not differ significantly from those of the ' ' NEV 016561 I WATER PCB-SD0000030509 i unexposed group, but the livers of the rats that vere exposed to Aroolor 1254- vere enlarged. The prothrombin content of the blood of the animals exposed to either Aroolor, as measured by the method of Kato, remained with in normal limits. The deaths that oocurred among the test and control ani mals were the apparent result of an appreciable incidence of pneumonia. The lesions of certain of the animals were those of a frank pneumonia; in others such lesions were not fully de veloped. Degenerative changes in the visoera were usually found, in varying degrees of severity, in association with the pneumonia, but in certain animals, test and control alike, the degenerative changes were more evident than the pneumonia. The similarity of the lesions in test and control animals, and the .lack of characteristic evidence of chemical, pneumonitis, lead to the reasonable,but not altogether certain conclusion that all of these fatalities resulted from interourrent disease among the animals, and not from the effects of their exposure to the Aroolors. For praotical purposes, this conclusion is subject to the critique of further experiments involving more prolonged exposure of animals to somewhat lower concentrations. Properties of the Experimental Materials Aroolor 124-2 is a light straw-colored, mobile liquid with the composition of a chlorinated biphenyl. A letter from NEV 016562 WATER PCB-SD0000030510 *'****# i - 4- P.G. Benigus, dated August 30, 1953 states that it contains 42.0 +0.5 per cent of chlorine, whioh corresponds to the presence of 3 chlorine atoms. The positions of the chlorine atoms within the molecule have not been determined. Monsanto Application Bulletin No* P-115, gives its specific gravity at 25*725 as 1*373 to 1*366, and states that it has a distilla tion range of 325 to 36oC, a refractive index (D-line at 20C) of 1.^27 to 1*629, a Seybolt Universal viscosity of 100P of ^0 to 93 seconds, and a flash point (Cleveland Open Cup) of 176 - l6oC. Its vapor pressure is about 4 mm at 150C and about 30 mn at 200C. Although insoluble in water and glyeer- ine, it is soluble in rest organic substanoes. At 745 mm of mercury and 25C, the concentration of 1 mg of the vapor of Aroclor 1242 per liter of air is equivalent to 96.9 ppm by . volume. ' " Aroclor 1254, which corresponds to pentachlorobiphenyl, is a light straw-colored, viscous liquid. The positions of the . chlorine atoms have not been established. The molecular weight of pentachlorobiphenyl is 326.445, of which 54.3 per cent is chlorine. Benigus (ibid) reported that Aroclor 1254 contained 54.0 +, 0.5 per cent chlorine. Bulletin No. P-115, states that Aroclor 1254 has a speoifio gravity at 25/25C of 1*533 to 1.543, a distillation range of 365 to 390C, a refraotive index (D-line at 200) of 1.639 to 1.641, and a Saybolt Universal NEW 016363 . .. a A i I _ WATER PCB-SD0000030511 k -5 visoosity at 100* F of 1,^00 to 2,500 seconds. The vapor pres sure at 150*0 is about 1*3 mm and at 200*0 is about 9 mm. It is soluble in most orgenio substanoes, but it is insoluble in water and glycerine. At 745 mm of mercury and 250 the oonoentration of 1 mg of Aroelor 1254 per liter is equivalent to 76.5 ppm by volume. Experimental Methods Exposure to the Vapor. In a preliminary experiment with the vapor of Aroelor 1242, a group of animals, consisting of a cat, 6 guinea pigs, 10 mice, 4 rabbits and 10 rats, were oon- fined:for 7 hours on each of 5 days-per week in a;rectangular ........ plywood ohamber (volume 600 liters), of which the inner metal lining was coated with a baked ohemioally resistant plastic, through which was passed a stream of air laden with Aroelor 1242 in a known concentration. The stream of air, conditioned with respect to temperature (75 + 30F inside the ohamber), dust and humidity, entered the chamber through an "Anemostat," located at the oenter of its top, at the rate of 500 liters per minute, as measured by an inclined manometer attaohed to a venturimeter (Figure A). The air was withdrawn by suction from the chamber through an exit tube located on the rear wall near the floo, equidistant from the sides. ' MEV 016364 -- WATER PCB-SD0000030512 Aroclor 124-2 was volatilized from a heated glass well main tained at 132 to 13gC. The air passed over the surface of the liquid before entering the chamber (Figure B) - In a second similar experiment with the same Aroolor, the temperature of the liquid was kept at 100 to 105C. Uhile the seoond experiment with Aroolor 124-2 was in progress an experiment involving the vapor of Aroolor 1254- was carried out in a second chamber similar to the first except for the absence of a plastic inner lining. The rate at whioh air flowed over the liquid Aroclor 1254-, which was maintained at 130 - 1350, was 400 liters per minute. ' In order to accustom the animals to the experimental pro cedure, they were kept in their respective chambers for 7 hours on each of 4- consecutive days during the week prior to the in- troduction of the Aroolor vapor into the chamber. For purposes of control, a third group of animals was confined throughout a like period before and during the period of the exposure to the Aroolors in another ohamber supplied only with conditioned air. Method for the Determination of Aroclor 124-2 or Aroclor 1254- in Air. These materials were determined quantitatively by virtue of the fact that, on thermal decomposition, they yield hydroohlorio acid which, with silver nitrate, forms a suspension, the density of which could be measured by means of the Beokman spectrophotometer at 500 sjue ' MEV 016565 WATER PCB-SD0000030513 On each, day, 2 samples of air from each respiratory ehamber were collected by passing air at the rate of 1 liter per minute for 15 minutes through a Willson oombustion furnace, end then through 2 midget bubblers, in series, each containing 10 ml of 0*1 N sodium hydroxide. A quartz tube (13 mm outside diameter, 7 mm inside diameter and 16.25" in length) contained several strips of folded platinum foil. The middle of the tube vras wrapped with a heating unit. The heating unit of the Willson furnace (Figufce C) consisted of 20 feet of B. and S. gauge 22 nichrone wire (1 ohm per foot), and was covered v;ith asbestos cement. The semple of air was humidified by a cotton wiok ' saturated with water placed 0.5 inch upstream from the quartz tube. With the furnace maintained at ^5c the Aroclor was decomposed to free chlorine, which reacted with the moisture to form HOI; this vras absorbed in the 0.1 N sodium hydroxide in the midget bubblers (Figure D) equipped with fritted-glass bubblers (Mine Safety Applianoe Company No* 43^67). The 0.1 N sodium hydroxide was prepared by dissolving 4.0 g of pellets (ACS specifications 0.01 per oent 01) in about 50 ml of double distilled water. To this was added 10 g of arsenio trioxide (chloride-free) that had been dissolved in water and filtered through Whatman No. 42 filter paper. The oombined solutions were diluted to 1000 milliliters. NEV 016566 - WATER PCB-SD0000030514 The sample was transferred to a 25 ml graduated cylinder to which 1 drop of a solution of phenolphthalein was added (1 g of phenolphthalein dissolved in 100 ml of CP methanol). The solution was neutralized with 3N nitrio acid (190 ml CP concentrated nitrio acid diluted to 1000 ml with double-distilled water) and diluted to 23 ml with double-distilled water. One ml of 3N nitric acid was added to obtain a pH of 1. After adding 1 ml of a solution of silver nitrate (3 grams of AgNOg diluted to 1000 ml with double-distilled water)., the suspension was mixed by inversion. After standing 3 minutes, the trans mission of wave-length 500 m,u was determined in a 50 mm cell by means of a Beokman spectrophotometer which was set against a corresponding cell containing a reagent blank. . The amount- of -Arcelor was determined from a- standardized curve obtained In a'similar"manner from'known quantities of sodium chloride. A curve relating the transmission at 500 mp. of a suspension resulting from sodium chloride over the range of 5*0 to 120 micrograms'per 25 ml offinal suspension is shown in Figure E. On ther babis-bf 4j2.0 per cent of chlorine"in' Aroolor 1242 and 54.0 per cent in Aroolor 1254, 1 microgram of sodium chloride is equivalent to 1.4442 micrograms of Aroolor 1242 or to 1.102& miorograms of Aroclor 1254. hev 0ie!>67 WATER PCB-SD0000030515 -9- Experimental Results Aroolor 1242, Experiment No, 1. One eat, 6 guinea pigs, 10 mice, 4 rabbits and 10 rats survived when subjected for 7 hours on each of 17 days over the period of 24 days to the inhalation of air bearing the vapor of Aroclor 1242 in the concentration of ($.6 micrograms per liter (0.S3 ppm, Tables 1 and 5) No signs of intoxication were observed. The weights of the individual animals (oat, guinea pigs, rats and rabbits) have been plotted in Figures 1, 2, 5, 6 end {$. The averages end the ranges of the weights of guinea pigs, mice, rats and rabbits have been plotted at semi-weelcly inter vals in Figures 3 4, 7 and 9 respectively. Despite slight fluctuations in their weight during the period of exposure, all of the animals, except the guinea pigs, gained in weight during the period of exposure (Table 6) The average gains made by the various species, expressed as per centage of the initial weights were: cats, ($.1; mice, 10.3; rabbits, 7*7! and rats, 2.1 T' he loss in weight on the part of the guinea pigs averaged 2.0 per cent; most of them were gaining weight (Figure 2) at the termination of the period of exposure* All of the animals were examined post-mortem, and since gross examination of the visoera did not reveal any signifi cant alterations, the tissues of only representative animals NEW 016568 , 3 3 l _ \ WATER PCB-SD0000030516 - 10 - (1 oat, 4 guinea pigs, 6 mice, 4 rabbits and 6 rats) were sectioned and examined miorosoopioally. No abnormalities , were found in tbs visoera of these animals. Aroolor 1242. Experiment No. 2 (6.35 'f) /l) The pertinent data relating to the fate of the individual animals in the Ex , perlment No 2 are presented in Table 2 and summarized in Table 5. This group oonsisted originally of 1 cat, 6 guinea pigs, 10 mioe, 4 rabbits and 10 rats, but a few additional animals vjere used to replace those which died. One cat, 4 guinea pigs, 6 mice, 2 rabbits and 6 rats survived throughout their exposure, for 7 hours on each of 32 days over a'period of 120 days, to air bearing the vapor of Aroclor 1242 in the concentration of 6.33 micrograms per liter (0.66 ppm). Certain animals (2 guinea pigs, 1 mouse, 1 rabbit and 1 rat) were killed after 65 to 63 ' periods of exposure for examination. In addition, 1 rat and 2 mioe survived throughout 23, 4l and 5$ periods of exposure, re spectively. Seven fatalities occurred as follows: 1 guinea pig after 15 periods of exposure; 3 alee after 24, 4l and 70 periods of exposure, respectively; 2 rabbits after 15 and 64 periods of exposure, respectively; and 1 rat after 56 periods of exposure. Except in the case of the rabbits, the incidence of mortal ity among the controls was equally as great (or greater, Tables 4 and 5) as that among the experimental animals. One oat, 3 NEV 016569 r -rrg WATER PCB-SD0000030517 11 mioe, 3 guinea pigs, 3 rabbits and 3 rats survived, following their confinement, for 7 hours on eaoh of 34 days over a period of 122 days, in a chamber in whioh the air, conditioned with respect to dust, humidity and temperature, contained no vapor of either Aroclor. One guinea pig, 1 mouse and 1 rat v?ere killed after 66 (or 63) periods of exposure. Other animals introduced later in the experiment survived throughout the following numbers of periods of exposure, respectively; 2 guinea pigs, 20 and 77; 3 nice, 14, 14 and 19; 1 rabbit, 19; and i rat, 44.Eleven fatalities occurred at stated times among the oontrol animals, as follows: 2 guinea pigs, after 7 and 6o periods, re spectively; 7 sice after 5 59 63, 69, 70, 72 and 32 periods, respectively; and 1 rat and 1 rabbit after 42 and 65 periods, respectively; `" " ' No uniform or suggestive signs of intoxication were ob served among either the experimental or control groups. The weights of the individual animals (oat, guinea pigs, rats and rabbits) exposed to Aroclor 1242 are shown in Figures 10, 11, 12, 15s 16 and 13. The average weight, and the great est and least v/eights attained by any individual within the groups of guinea pigs, mioe, rats and rabbits, are plotted in Figures 13, 14, 17 and 19, respectively. Comparable data on the oontrols are given in Figures 31 to 4l. NEV 016570 WATER PCB-SD0000030518 - 12 - The surviving experimental animals grew equally as well (or better) as the controls (Table 6) During the period of exposure to Aroclor 1242 (6.3 lO/l), the average gains, ex pressed as percentages of the initial weights, by the various animals v.ere as follows: oat, 22.2; guinea pigs, 56*4; mice 21.2; rabbits, 505 &&& rats, 17*6* The oomparable findings on the controls were: cat, 6,6; guinea pigs, 56.6; mice, 17*4; rabbits, 44.5; and rats, l6.6. The weights of the livers and kidneys and the relation ships of their weights to the body weights (expressed as grams per 100 srcr-s of body weight) of the individual animals that survived following exposure to the vapor of Aroclor 1242, are shown in Table 7, whioh also gives the corresponding data on the oontrols. Comparison by the "F' test of the variances of the ratios of the liver or kidneys to the body weights of rats, guinea pigs and rabbits exposed to the vapor of Aroolor 1242 (6.*3 y7i), with those of the controls, revealed no significant differences (P *0.05) except in the case of the livers of the rats. Application of the "t" test to the mean values (Table 6) revealed no significant differences between the test and control groups of guinea pigs, rats and rabbits. The apparent prothrombin content of the blood, as measured by the method of Kato (Am. J. Clin. Path., 10:147 1940), of NEV 016571 WATER PCB-SD0000030519 - 13 - oertain animals that had been ejposed to the vapor of Aroolor 1242* are given in Table 9, Comparable findings on the con trols are presented in Table 10. In Table 12 the peroentile relationships of the clotting power have been obtained arbi trarily by dividing 100 times the average clotting time of the blood of the control animals by the clotting time of the blood of the experimental animals on the same day. No diminution in the clotting power of the blood, as a measure of the impairment of the function of the liver, was induced by the exposure of the animals to the vapor of Aroclor 1242 when present in the air in the concentration of 6.3 micrograms per liter. Determinations, of the numbers of erythrocytes and leucooytes and of the hemoglobin content in the peripheral blood of indi vidual guinea pigs and rabbits' subjeoted to.inhalation of the vapor of Aroolor 1242 in the concentration of 6,3 micrograms per liter, are given in Table 12, which also includes comparable results on control animals. These values have been summarized in Table 13. Application of the "t" test to differences in the mean values (Table 14) for the experimental and control animals yielded border-line evidenoe of significant differences in the numbers of leucooytes end in the concentration in the hemoglobin in the blood of the guinea pigs, but these were small and of opposite sign (the leuoooytes in the blood of the test animals NEV 016572 WATER PCB-SD0000030520 i - 14 - were low in number while the hemoglobin content was high), as well as being subject to individual variation, and they oannot be regarded as of physiological significance. Of the 7 animals' that died during the seoond experiment with Aroclor 1242, all but 2 mice were examined post-mortem. The tissues of 2 mice had undergone significant changes, post-mortem, and were not examined microscopically; gross evidence to determine the cause of death was not available. A third mouse (B-244) was examined and found to have a malignant lymphoma that involved the liver, spleen, kidneys and heart, and therefore its death was attri buted to natural cause. The cause of the death of the guinea V- pig was uncertain, but the 2 rabbits and the rat were found to have died of pneumonia, and not, apparently, of chemical pneu monitis. Generally, the survivors among "the~ various speoies had normal visoera. However, 2 guinea pigs and 1 rat had degenera tive lesions in the liver and kidneys, 1 guinea pig had aoute lobular pneumonia, and 1-rat had interstitial pneumonia, Eleven of the animals (2 guinea pigs, 7 mice, 1 rabbit and 1 rat), that had been confined in conditioned air only, died. The viscera of 6 of these (1 guinea pig, 4 mloe and 1 rabbit) were examined microscopically. Each of the animals examined had died with pneumonia, in association with degenerative NEV 016573 WATER PCB-SD0000030521 - 15 - changes of varying severity in the brain, liver and kidneys* Animals of all speoiea that survived were examined and found to have normal visoera, but for scattered pneumonio lesions in certain of the guinea pigs and mice, and for one case of cellu litis in a rabbit. Aroclor 1254-, Experiment No. 1 (5,40 .//l) The pertinent data relating to the change in weight and the fate of each in dividual animal during and following prolonged intermittent exposure to the vapor of Aroclor 1254- are given in Table 3 and are summarized in Table 5 The original group consisted of the same numbers and types of animals employed in the other experi ments . One cat, 3 guinea pigs, 4- mice, 2 rabbits, and 9 rats survived throughout their exposure, for 7 hours on each of {$3 days over a period of 121 days, to air containing Aroclor 1254 in the concentration of''5.40 micrograms per'liter1 (0.4-1 ppm). Certain other animals (3 guinea pigs, 6 mice, 1 rabbit and 1 rat) were killed for examination after 66 or more (up to 74-) periods of exposure. One rabbit, introduced after the experi- ' Tl . - ment had been under way for some time, survived, having under gone 33 periods of exposure. Five animals died during the experiment, 2 guinea pigs after 12 and 17 periods of exposure, respectively, and 3 rabbits after 15, 15 sad 20 periods of ex posure. Exoept for the rabbits, the incidence of mortality among the exposed and control groups was comparable (Tables .4- and 5) No signs of intoxioation were noted among the experimental animals. nev 016574 WATER PCB-SD0000030522 - 16 - The weights of the individual animals (oat, guinea pigs, rats and rabbits) are given in Figures 20, 21, 22, 25, 26, zt and 29i while the average weights of the guinea pigs, mice, rats and rabbits are shown in Figures 23, 24, 27 and 30, respectively. Except in the case of the guinea pigs and rats, the animals that survived throughout the period of exposure to the vapor of /troclor 1254 grew equally as well as the oontrols (Table 6). During the period of exposure the gains made by the animals of various species, expressed as percentages of. their initial weights, were as follows: cat, 22.1; guinea pigs, 40.7; mioe, 23.9; rabbits, 397> &md rats, 6,9, The lesser gain by the ' experimental rats nay be explained by the fact that they were initially somewhat heavier. The corresponding values for the control animals have been given previously and are also shown in Table 6. The weights of the livers and kidneys, and the ratios of these to the body weights, of the animals that survived through out the exposure core shewn in Table 7. Application of the "t" test to the differences in the mean values which characterized the experimental and control animals (Table 6) shows that the weights of the livers of the exposed rats were significantly greater than those of the controls, the ratio of the liver to the body weight of the former being 5*34 g per 100 g of body weight, that of the latter being 4.15'g/100 grams. NEV 016575 WATER PCB-SD0000030523 I - IT - The results on the apparent prothrombin content of the blood of rabbits and of a oat, following exposure to the vapor of Aroclor 1254 (54o *//l) are given in Table 9* Little or no diminution in the clotting power was induced by exposure to the vapor of Aroclor 1254- (Table 11) The numbers of tbs cellular elements and the concentra tion of the hemoglobin in the peripheral blood of the individual animalb subjected to inhalation of the vapor of Aroclor 1254 are shown in Table 12 and are summarized in Table lj. The slight apparent increases in the hemoglobin of the guinea pigs and in the numbers of erythrocytes of the rabbits, vhile statisti cally significant (Table 14), cannot be taken seriously as ex pressions of the effects of the inhalation of Aroclor 1254. Tha.-viscera-of.4 -of the~5 animals (2-guinea'pigs and 3-. rabbits) that died after varying periods of exposure to the vapor of Aroclor 1254, exhibited diffuse degenerative changes in the brain, liver and kidneys, in association with pneumonia? Animal's -that survived., following exposure-to the Aroclor,-were examined post-mortem, "and the viscera from representative ani mals were examined microscopically. The viscera of the oat and guinea pigs were normal. All of the mioe, exoept one, had normal viscera; the exception (B-237) bad slight degenerative lesions in the liver. A rabbit killed 1 day after the 66th NEV 016576 t WATER PCB-SD0000030524 I - lg - period of exposure, had degeneration of the liver and kidneys, and pulmonary hyperemia ana edema* Otherwise, the rabbits that were examined had essentially normal viscera* All of the rats that were examined had degenerative changes in the liver, and in 5 of the 10, these changes were found to be associated with pneumonia of low grade. The Aroclors did not appear to act as primary pulmonary irritants. The pneumonia that was observed in both the experi mental and control animals had the characteristics of an in fectious rather than a chemical process. The toxio degenera tive changes which were found in the liver end kidneys of all species of animals, in both experimental and control groups, while not associated with pneumonia in all instances, may well have owed their presence to the intercurrent disease which was characterized, ohiefly, inmost instances as pneumonia. Disoussion The probability of the ooourrenoe of the concentrations of vapor maintained in these experiments in actual industrial prac tice is remote, since these values approach those of atmosphere saturated with the Aroolors. (The significance of particulate matter, in the form of oondensed droplets or of products of ther mal decomposition, has not been involved in these experiments*) NEV 016577 1 WATER PCB-SD0000030525 - 19 - . Since the animals v/hich were killed for examination after 65 to periods of exposure were found to have degenerative lesions in their viscera which oould not, with certainty, be attributed to extra-experimental causes, these experiments were terminated and others have been initiated at about 1/3 to 1/4 the concentration. Frcn The Kettering laboratory in the Department of Pre ventive Medicine and Industrial Health, College of Medicine. University of Cincinnati, Cincinnati,. Ohio. ' Project Nc. 47* Experimental 7;ork and ?.sport by: Joseph F. Treon, Ph.D* Frank P. Cleveland, M.D. John Cappel, B.S. Frederic E. Shaffer, M.S. Ralph V/. Atchley, B.S. John P. Torbeolc, B.S. Approved EobTrTTJTiaTioe Bireotor ' June 22, 1955 [ r NEV 016576 WATER PCB-SD0000030526 a oN o\ o o HH -<-0 r3 X M WXXXXXXXXX X X X X X X X *'1 "^3 O O poooooooool 000000 a K3BS3SXSSS Q Q O o Q 0)0 p p 900000000 0 p p P P p >3 Ip o* o' ct o* o' ct C0PfQPtOP^tP^tUPOP^DPUPQttPUQPUQ 3)03tSC3BJBJtaCBtfltfl| H H H H H Hct 3S33i!S 9fl) 0P 00 0P 09 0P MH 1 1 1 1 1 1 1 1 1 1 r U 'o U U -0[tr- II VMVM rCu7NrUo1iUu1Vru/MtJu1f<uJ1rururuVroJlfl(sc3rtj|' -O-J,OM3 03-0 CVUl-pTUM fU M 'J'-Pi a tfl w w ta co 11111 kNUMUNVMVMUM (sMVMUMUMVMVM, OvJ'-PAX fU M FNM O OOOOOOOO C3*| OOOOOO ru OHcoOCpsfiLOtmu^OHo OiouhONONh OrhuOcrtu-OiouOrouo0ruitlne0sv^oxs--rp3urcCo^Oa<u-nrv^unnd.COD' -- H3 SC a ct rt-H ++ ++ I I + CD\Oj kn o ctum cool CD -f="OK3vjl HUM O 0-0 O crvcd O4="4^(30p--Pi + +lt I till I + 1111 + unuiwun o--P"-p-UN OUNVX H H ru fUUM Hi CEN>r\M OVMUM 03030 OuCCkOVJl CD0-0 fU ru 01 CO BBWWOOrotOWM tn co w cn w co es1 sees < H i < H < s 0 1 <<<<<<<<<< ooeooftasaa a aaaaa aaa a .<H<<H<* <I<*<H<<<H<<H fia affl fafl affl fafl fafl |52222i H- W H S{2{5 515 {532*2 HHI HHHHHHHHHH HHHHHH Ho Hto a 0, OHaDHaOHaQHaOHaOHaDHaOHaOHaOHa (H bia H oa HH sa oa H 1a H oa *a HVM W O r aa_ P. H HUN TO HUH H H H 0^000 H i-* HUMVM H H 0 DO -0-0 r* O aaaS'apaaaa a a a,t ap p<< a< u IPPP<<P<<PPIDP fivj< p< p< <<<< |p p p a pppp p> fi p << M it p< p p< p p << P p < p a B <4 B _ *0 h3 ffl O O *3 aM kH* 3 (3 o HU XT 3 ffl W P a * X 3 Hft fOfl **d3 Cff3l P 3 ffl H ft P H Pp3 & ffl o affl < H *3 Ho..t O P H O CD J3* t-1 P fflj Po O 1 a**33 H H O 0H ct 3" ct 3H O' H Po SJ p P: ct ct H O O 3 P*=d ^ O a< H H S' fod cuo > o M *3 H H p H ct 3 O ffl CO o a3 c ct S1 H o fCl o aI o o *f8fl 3 -b f2fl1 H 3 o S* 3ct 'g 3 SaS O HI P V 3a o 0 a H O3 t 1 CB T "5 o NEV 016579 WATER_PCB-SD0000030527 XICD o o ON 3 o | 3 !3 IP ct -* CD O V04 3 H-0 XHXMHXWWXX \h m oooooooooo PO ppt|8M>Bl|P tr* t* t< t f t* r* t< f f !IIIIIIIII B? ts"o P O O ft P? *c o , ** O o Hi P. O 3 _ *3 O' H* <9 P fT" oH s OOOOOOOOOO U4Ttuforotoforororofo rofcrro VKxvx.p-'j' no ovnui rob ro 4r*ooo o o O-O-P'O-Ojo'Jl _M 39* XH<t p.? ft H ++++++++++ 4TO ^T04="O ro O V'-P-!-' b ro : *.*,, *'.* . out ro onooro'O' ru-P'-csixNO + + I + + + l I+ + H- + vjhoj o H'J* o o\o o ro ONH ^ 09^.4 onF\X.F*CD 0>KD CD. M o fT3D TD at M 3J {3" O OC 3 0 P W O 3 31 H.9J-M 3 ' 3-H> W(-4 0$ fHt.U<B9 *.03 p 30 009 M ft M H> pup .tJCT O _ o p* ,, O P ^ * w t-b H> tt is- a*1 -5 CfltnMWM CO 09 09 09 M (to to nzznzztnnnzt <<<<<<<4<<<< V- <<<<<<<<<<4< ooooooooso 0 o 0.0,p.p.O.p.0PiO<0|O.P* H> t- HH H00H(H9H0H0H0H0H00H0H HH P0 PPPPPPPPPP a. P> HHHHHHH HVWVO ro*-* -4-4^3-J 0-4 O O P P 0.0.0.010.01000 P a 0 p p p p m p p pi4 4 B (B 4 <4 <4 *3 <4 4 <4 a a 4J oaottaaon p tt 'dH H" < O ku t*3 1 p <co iff H* P ft Htr 3 0 >o 3 *3 n lO Htf MOO ED I 0.3 jS t 1 *T3 0 *02 tU WEV 016580 WATER PCB-SD0000030528 ( K]p \ k fr-CD 03 CD pturoru NOH P"ONO ICUD-t.ojv0ru3 OaN\r0u9 ttuovPo croollr0o3 xxx XXXX xxxxxxx oooo oooo bbbbbob Table 2 as2g S2X300O O9 Q op oooo au aa au au aaaa oooocccccecli UUUU3D33333 ooooooooooo B p 0 p P ID P p D 21 rtP & CD tfl t S3 till ru ro ro ro "-P"-p--F" n ai o) a kg. rtu rio 33 P *3 P 3) P *? P 33 P 33 3) F* t- H-Ln 09 05 05 05 (R35I _Cf *P ao 3P 03-0 ONJ1 -fcrvx ru i-j co a ts w tu tu iI,mp1-v1vxxvrioxvrIoxvrxovrxIovrIxo ko OVD 03-0 CTOI ct P o 0 3" %o a p oooo oooo P fUfUH VO P OVD oooo o o o o o o o I fo pooro orooru -Ovx^r-O kw-(="Vxvwvx vx-Cr-l to tme\txo'-0O4^=-"CcBviOoCv-rij-o ^o p p ct .W05 P 3 C/3 a |n tf p ct + + + + s + I + +++ + | 5Ct p o P P PVX VJI-fr-Q P vrxo-p-rpuvoni -o-P" vn -O'-n CD-o o pp-j cvavx Ccvx o ov unvxv/i o VOVX-O 0-0 OvO H 33 cr P 9 P HP ro 3 P3 X 3 P ct O *0 C5 P 3 ffl 0 P3 a a* H ct IB P p Pup -SC9 a P !g 1 pa n . 3) >\ 72.0 p p VO I* I I VJ1 _ to oo till i i i i i 00 H " ?h3 p &xp2. O O 3 ct ag 30po3 3P O CO CO CO O C O coj MKUKMbM s.________ ! srssfl sissr P < < < p* < p. < < < < < < p. < PPP ffsl f<fl f<fl P< P< '< < < < < < ffl, 0 0 0 0 9 O ppp P> P Qt Qt Qt 0* 0* P* ton1 WWH! . 3HHHXlP&H P P P P P P P OPPPPOPPPPi HHHHHPP s ft3 ffl 0,3 << LP P P P P ffldftn < p p| P P p P P oo ft p 9 0* ^p-HnnHWH pvn -fcr- u a Xrtfippxtr. p. 0 *-4* lao*ovj 0*0 p HHXHH vji ui pp p p p ,p p P p P *4 HPV4P4 ua &P P 4 7E 3 % a tr* O3 P3 4 p a !P <05 H9S 333 \ft PP 3 Q ffl CP <9 P3 C NEV 016581 IK.. WATER PCB-SD0000030529 C o n c e n tra tio n Duration of Exposure O' OCND H .ofSn rSo 5ro3torocirKunoam5CmDroCrDoCvjrt XXXXXXXXXXX ooobbobobbo OOCTN> oHn OonvOpn"fCoOCioT pH*n0j3i 1C0DCfOD XXXXX XXX X -0--3--3--3--3 -3--3-3-3 ooooo oboo a 3 V0 %U Number gPjPvsiasaoaBsOjaPjsBoOsjaBsBc wcfcTftctctcfrt-ctrt-ft rJoL-'_n\<jiy1nv1jt o1n\1_n1on1un1on1or ry )_i ovroo oro-1o0 aro Sp)0pM5p3Wp 3p3 Oto-''oHO**' oOH*-' oHo>''eOHr-' a0uBcO0cuwO0caa0cBs& ct- ct ct ct ct N-f0FlO"Oo--HiHn3O--MwlN3O--Mh1X3O--oM1N3 F0oPt3TMv'o0-in330l0i^no33l.vf8nOr3- Id e n tifica tio n In itia l .P e g )______ W eight OOOOOOOOOOO j9owo-tootorvomoraCoH'vvo^ro'yhrjoororooorvohorv.c;\ -Ororo7<eooro0vorCtonoOtoOos--crroo3 POOroTOOroOOOtoOOOro \ + * + + ++1 + + ++ + + + + + 1 + + + ov"+c"-r>-r-o3 vOoM4t-=L*-f^Oc=i-oHsCH-nDf-*O-O3 vHvjri vho fo-op*tOooHnVoOn pvnroornooonx * ru axjn ropoq- 501 0 *0 5i to *>t- H0 JO 3 J- of + ++ 4=-*rvN 1 Oocnro ++ M : : on 1 1 00 PI nerict ieantlagWeeiogfht h3 aPoM' . lo _ 1 03 aa ro t t Change:in Weight ` Expressed as 1 A fter P e rio d Exposure Fate Lve ni vgathl F in a l Period o f Exposure S urvived K ille d 15 days S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d K ille d K ille d KK S urvived K ille d 17 days Died S u rv iv e d ii llllee dd K1 idllaeyd 1 day Died During 15th p e r i io d o f exposure Survived K ille d 1 day S u rv iv e d S u rv iv e d KK iillllee dd Survived K ille d 1 day Died 1 day Survived K ille d 1 day Survived K ille d 8 days S urvived K ille d 15 days S u rv iv e d S u rv iv e d S u rv iv e d j 1I j K K K ille ille ille d d d . o f Sur A fter 15 days 1 day I. 1 f l a y ____ day 1 day 1 day 15 days 15 days 1 day NEV 016582 WATER PCB-SD0000030530 31 % P I 03--3 cvqd--a-o a>-o--j 2>-3 OvM 3H OvftJ XXMXXXXXX 4MHX 0--3--3--3 ooooooooo 3000 VMOVCWDUCJDVCXdiMCD XXXX -J-- 3-3-SI-3 oooq o o c* u%10 H -- sssaasaess _uOO_u O3p Ocp Ocn Oca OcuOcqOcu . &u M O3 M O3 M s p p p pi 03 Cfl 03 WtB 03 03 03 tfl ro w fu fo fu i\j iu w w a M a M M33 3> M vo cd--3 cr*-n^~~\x fy (_i kmcmccs essfflia P ^piV-H* VXi amxnm.pw-v>ro oooooo.o.oo OOOO loyHoroUHoMoHorooWoWo fifing MvO Ovo Ov0-p"O Kj fo--3 cap- MMMM 30PO34 P30 P43 a a a ml MMMM noinip tnaJtDtn tiii VMVMVMVN Po ro ro t-> to m ovo oo o q to V2M^-oP'-oP -PTo4 OOM s a 3ct 3M 3&a *Ma3 4 cPt OM 3 ss 3M fVKa_'-CHpP-'CHpt* ' ct M + + ++ +4-0-+f+ + I VO mv/i ovo M--3vJi O' frv-n vm ro MVX H H OVJ4 O OVsJl Op F*-3p-o I ++ P"-3-P"ro) M 0-34=1 Mvnvjj-J VJ1 ** I IIII > + M, II VOP1 o fo 0tt ^ 7 S' 0MM3 M3 Bi( H 33 tf 3mcMpt 33o WSr3P a3a cp M CPt MM M3 SC5 5* fO M (I ; 33 >MPPO3#PWH Ou 3O P3cf & O3 m_____* HWIHW WtotoH MMMMt-2.2.2.2. < < < p. < < < < < < < < &< d< < Q> Q> P> Ci (3i p, Q! Q, pj QiQipt w co to cn| o> o.4 < <s <<o <O &<5 , 3.P.P. WI-- !I MI Ml 8.1 Ml v* iI nu5ho. nhu2& hn.pa..hn&..h.nM#p.,h.Mvp...*M.lnMfMlnMPMinMM&'P4i 'HHHHHHH IWMHtt* wH* OP* *Mb5 MMa MM MM MM O*-IP P. Qt D Pi o vHn HMovMo' l o. to 8,js8`&g'*g>g`p0, Oi 0 I d Sr P P. Oil VuJ sjv,S3r ? " co oi hg o m< a *33P M4 034 HP ct fdo a33 >o UC 3MeMt *-t> <3 00 0) ffi P, 3 | I S?P tor* 4 3* Io *3 a 3 f t3 3ct M *4 OP' M O2! P M ,tn Oi iCl 3* P 1 gt M NEV 016543 WATER_PCB-SD0000030531 (hours) %l 0 P" O O H3 03 03 COCD 00 03 0303On O: *>4 l\)HH 03030' 00 \J4\J4V>4\>4\>4V34V34V34 004 .34 OVJlsmj4^>4 O' 34 XXXXXXXXXX X X X X M X X X -0 oooboobooo obboboo O Mouse B-2k0 50 53 50 53 50 S3 50 50 50 50 50 50 50 50 50 50 50 PPPP000PP0 0 0 0 0 0 - 0 0 tr* t*ftftcfrtftttftrtrtct t-> t* t tr* tr1 t* t* t* i i iiiititi Oo'' H" ft Ocr' t-- ft Ocr' 1-- rt Cof' >-* tt Oo'' Prt Oo*' Hrt- Ocr' tt vVtjvJi<Ij-rP*\nVjrvXorfirvuOjri<HosrjOoi'jVti-vO>jiHCj3iv-hJjivVwoO -H00J33W0-H1J3\-HJCJ*1E0-XH4pJJ~4^vrW0%1>>l\>OJH144n0HfOV14J Number Id e n tifica tio n oooooooooo ro ro ro ro ro to ro f0oo-Pfroo"rrocy\nxfo\u>i 4rho\>-r4oo4rwo=^fi--u3*v'wJ>'i -wwP- t-4-0 0N04V0 0 tovn Ov>-F'-P`\>* axe 0301 0.022 ^-5 3 H> X* H ft RW P wp* P ft H + + + + i + + i i + 4- | t 1 + + + + \ o+jtvH>4 kj HH W4r*0 coo H M W-fr'H OV-4 h h-P~4=~ct 0O4MO004H#^\CN<O^0r\-j04 -4 0*0404^4 H CC --ro0 04 of of + iii + VOM O03HtMMH 1 1 1 I 1 H till* II Crv 0*0 Duration of Exposure During A fte r | I t of ig h ei antlag'"ee PI nerict Expressed as '"e ig h t in 1 Change o3. o' a 04 I *0 P W C o n c e n tra tio n i Period Period Exposure Exposure t Pate S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d Died Died Died S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d S u rv iv e d K ille d KKiillllee dd 2 days 3 days K1 idllaeyd 1 dav K ille d 1 day KKKKK iiiiilllllllllleeeee ddddd KK iillllee dd K ille d 15 days K ille d 15 days 1 day 1l 6 day days 1 day 1 day 1 day 1 day 1 day 15 days 15 days * 0 H < S* H t--< H 0 (3. H 0o *0 a >o # 1 nH 5 H ft a *5 0 a w p a p*l jS a4t 1 - NV 016584 I I WATER PCB-SD0000030532 ! vf i i i I t I i I i | I H| CO t& I CD 3 nc ilo1 S' o] 1o3 oo %\ -* w 1P*3 O' Se3t S' a * I CO a t o C- NV 016585 WATER PCB-SD0000030533 H XX -4-3 oo X XX XX -4-4-4-4-4 o oo oo XXXX i o o o o oooooo XXX -4-3-4 bob s wasjasoso pp ppp p pp fte* e* e* t* e* e* O' O' 00 U1IVH t VOInvItrnIVo>oVI lvoIn htt* HH vn to H'O--V OOCOWOMI o c3J50 PPPP cr g* o1 o'! S3SS33 oc op oc o6 oc o jOate'ffO|J'|O|>Jt1t tt tt tt tt tt tt CO tt tt tt tt o et c+ et c* a a a a to a HtHiHllH.Linviivni ^ir-ito tio VIVIVIVI Vxvkvmvovi to C'ttC'Oe'-,Vi to i-*vo ovo VO 03<MVI asses 0oo 8tt 8tt 8tt tt tt tt at1o atio atio to to to CO-3 ov oo wh* ova VH to ooood to row to rd t-> to hvj cl-- MvntoHVCfVH VX toVHVI o o o o o o * vi vn m o] oooobo vn ovonKcax h to to to ro toCDto-Ft OVXV4VXVIVM o o o ooo K ttou w h i--2$ P3 X* H c* P5E ++ I ++++ to to VI H *-*V4 Ov*3 4=-V0VD to-pfto o\<r> .p-Hvo to cpvn HO vi o 71 .r . Ovj v>4 M 11 V0CC\0-R >C0 Sit IS < < p< < 4 < H" H < a <a '<<<<' `so a, aP> ppppj + + + + i+i + Hvn-tr-.E" to CD VI p-OOcrl Ovp"^S" oo I 0-4 OVH f-3 CVVM O-P" i c o' o o o I o o ooo oo jrsOMWW a w w a w w p. < < < < < < p < < H* P p`- P P <t<t <O tt O' <1 < tt tt P< P* P p p p p p I IV _p4-So" o vnvo o III OOO P PP OOO pi pi pi IfP O 3 fa a cot W ott --Ho H3-4tt C O' K0* 3 3 O OP a_3___P____ c* H O 3 o a 3* o WP 3 ft 3 0*0 03 0 30 3O et tt H J-6*. 0 .?n o o p aj o O P HHtt Ct 3 P a H* 0 1 *a p o to $$0 zv* 3 HMMI- ooo 3 0 0 0 0 *o n pppp vxvx o *tt*tt* tovnvnvnvT 0 pappaj odXPaMnP XiPiMPu 3 I-- O o> !j5 JJ pjvx K* H* I* HMH, POMP HHtlh PP i t3*Htt*lt-t*Mtt H tt tPt HO 4P4 HH 900 PPP SVIVl I w P P p Pi Pi tt ppxoa o PH M I-*iXVXVX i vnvn I ct p p PPP 3*0a.vptvpi IP P P Wtt p4 Itt tt cc 3 t-- O o . VXVI O -4 tt O PPP It VpI VPJ fHfl tt tt tt *3 o *i H O P> P S a ssw <88 O H H33 P et M3 oePteO_rwoa >o *3 *3 et oa 3 tt tt < NEV 016566 WATER_PCB-SD0000030534 (1) Ct hoempourtei gd confined infrao .in m time l t ogrocpopn of in itia of contr ditioned oa l lisr exposure o f exper and the o rig in a l o n ly , on each o f k -FE"oOnNCDp-CfDr XXXX oooo 50 SC S3 5J PPPP ct ct ct ct wi wi witTi ' Fhhh CDO^Jl-F* oo o oa WWIUH HOOM-VHVVJOi X- H* mf** p-sv 1 rt- H a * V IV H rv\j4~rv a aa vOOt-p-CD o oo I o OO o MPO03MPW H-lflS ctotjia t* 1 8 Hp cptM t* Pop OsgOOT p _ flH O t-b O P H- Ottffl oO t**1ct O *0 P> s C/l w w w ccce 1 *0 H H < < <s < (-* H- H- H <<< < poop aaap* tfl fc* tv/ nnn w HMHH 9HH0 H0 H0 p. p. p* p# VH MVWVX PPPP Vu4lo(wu| *o ,O WOW HP (t ct M K PW etO O o >o O etO 1 n -* '"FS ** o OO CO I NEV 016587 im ental anima group o f exp fo u r days du ls to the vapor o erim ental animals ring the previous f A ro c were veek. lo r j WATER PCB-SD0000030535 h o>Soo-o ru h* . ^O^OO^OO 3tf<S^SP<33 40<ltt000 a O o n v g ca ''.SSSGSSSe Sf f < moo<ooooo co<33o33333 epoa 3 a *3 *3 i <M4 <4 <<a4P PP4PP t- * ou^ttuauu <I<1 o 3 00O9O^f00Aft0 3 3 3 X O X X XX (matt) *o tj t) *a XopooXoo sstt'auxoB'OMu PPMoa^ppoap ((I CD *o CD p O Q>P>d pt O CD OOCaQiOOOOQtOO X X O 3 a. 3 3 33 *o o. o o OO 30000300 cacao 3333 33 a O 3 O H MHO HH p-p. sv$ o *) 34m OH 3 F* o o 3<^vx t-*-o rovj ctvctn 3 3M VX* MVJ4 --Jvji << tv> -P" OO OP-JAPHO O 3 3 H P -C-p p 3 3 M M-P"P Vi< V< P V4^- 3 P. w o 3 g>ON . P p P pi Co On 4rC<H> 3p,-o >3 aam *_P PVO | <a u fkvj J 0 0>0> COP P *< V4 p* ca ca P M Po u *a<!- < a3 3 a 3 ca ca ca a3 . oa oct ott O F- ft < H- P< VMi) F a <<4 IKP fat 3 FP H w O 2! P let ca aO* 3 oHi **} P ft aP pM *3 F> 3H ffi ca ca ,S ioH* O' 3 O t r F* w S' ct I CD p. l A Hi V $? ct p o 3 5 3ct P & Ss I O 3 I ca Hi X % 3 3 ct P H P & a o 3 ct 3 O H & Table 5 R NEW 016568 WATER PCB-SD0000030536 aia 2 O o P ct P & H* o C *61- p ct c* o* t>9 H* 71 a ct a p tt H* o 04 os H SB SO oa P p H* p ct cr o ct CB o* H- 3 H* 7> a <4 ca p CB H o F H o as H c/> ` o t* co d o ru o o fO H ro sjl a r9u H F O F ro CO -0 H o 04 o O ro cx ko o ro Ov ro o a O' ro -0 rO ++++ o M o o o S*4 o ro 04 *-* \r* o F Sr-on 4 O sn c T o a o % d o a Q. rt o OB N O + 4- + 4> M s-H ro sP O' o on H CN ro F to rj ro o o H o H ro 04 t-* o Mro fr- ro N O ro 0H4 ro cr -roj o oro o o sN as sn I H Hct a a o ro o o ru sO o 04 o o aO' ko Hro* d i - o ro i 1i o o o ra o o o o t-> 04 sO ~0 o *- 2F1 1 1i H O' -a as M P H OS t o 04 4o o ro c as 04 G o H ro. F Fo ro a a 04 iS H > --*4* a1* <5 tito eta s-U(*P Ct PC4 H d 4a F4 O o s & as F 044 O o o a F o a Op 1 4-yP-cr' a ct p 04 04 a o d o3 c?tLpoaq If 53 t* ct ct O d a o + 4- 4- 1 o O4 O o o ro o o o so J1 h* oro o CD 4o ro 0to4 a p* W M Si O > ct n p W J *4 o itho 4*135 a a 35 --p ab* top os -- p TO Ct O TO i ffi OS S' + + 4 t 4* S h n? m t* a o ka o> ro -u SO ro 44 a (* t* h *a tr < TOctooPasSP a a-HtbOttOOBd M -0 H a H o a ct p H a ct t# U TtO* to TpO H -- oH aw TPfOfl" & P. ct a CoOo 1111 1I11 1111 19I1 Owa N H 4g H 5* * a < Ct ft**t* I a Ja TO ct 4 -- a* H* P ct p H TO Sw' 4 M X wo a fhfl> >< ss s* o S TO o 1 o 35 a H&tfa wt* a ct p H a p to 5 ct p a a ** ct ct O 9 - ifo> a h- i* If o 1 STtSS S',&S*S 3 w- *3to Ct O TO pa 4H *6 3 bSdH3* O > t* h> a *a tr < 1 ^.ctoopt 4l# tTH-hm o o a 1 ct a a u 4*to p H ct w TO tt TO t-> P TO tCtL tcrr O 05 c < i* < o *3 tt > Ht9| P H CB t3 sr p ct a P> > aa4 P (g CO p Hto NEV 016589 is I WATER PCB-SD0000030537 .I rticrl OL -T.& g UP o|PH a *b& tS To p bW ON w fgO o a a (0 I 3 I I j l\3 m ^Ht> _O 3 < ** a a toft tfi f-*tf H- P CtPR *-* o s5 a I0 B OMO8P. N oW tv o I m K-,1V a?<rH a *o o 1a. H<T''o< '3 *3 a 3* 3 3 ct P pn ct P o act o o3 9. r o Jfp H o> .sr? o W wS pHS ?fc55a ru M_.-iUg o a ON M r ro P* jin O' on a 3 ct Io a O -H *0 M. .0 3 O M 3 OP kKt. On iON a 3 vo; * i O iSo-o l.a a 3 *3 3 < n ct O 3 is P o 3*aoOP033 ftp 33 a B an P H ft t-o---C-P----sp it 3o o, o P op* o 3 a ct o a> 3 o ro O3 oM ro o I-* oo a3 -S 'H > & M > E 3 a3 tv .ro ,cPCh- 3h- ^a Kn TO CP <1-3 P---3*a p 1 ctpn HO *4 OKM IV o n, oI oc3 p _ 3a 3 wo o boo H a- ka - 3 1-4 a 3* 3 a 3 ct P SPOT O ct P O 3ct aoct J3_______ T* is KM o ,k---ae- J- g= ^ o& P4 >!* HO 3 aap o p ypi W an 3 3 3 K,rHno "3c*t 03 a OPP oo mas fs ? e?oH Km Hi a 3 ns 3" s tv fo etO O P 3 =1 P 3*t**t 3 3 ct p 3 u an P " h c4t OonP1 on` h ro a pt --o i 3 O' H O ON r 33 O tv NfcV 016590 WATER PCB-SD0000030538 l aa PP OO'' oOl >{ct cti I Ml ]$o' a O & OJH- o e a g h a Q & .1 KMMo 3 P ,i M 3 .KfOo P ii ro p C3 Mro 3 p M $*" H OQ F) m S' Ml w *3" CD ro ON ON CroD CD ro CroD I!'Ml t,eoa\ iVJV ON ON ro CD ft ft 53 ro M o F" K> MM Ml t=- m ft M M M C~fO v>> CO aM > ro( oo1 H -af a O H*Q M H US ro ON co M pq a? a a 4 I 44 M oft H* 8O O u oM OM1 &ON t ON K ro o ct P 1 fccmMVmi*1$3 m co On co m .00 03 ru M cro iSf a- ct o ^ a O4 P ,* a h ct ct ft ct e1 * s* o a H p ct H* 3O to V u o - ct 3* o 3 M O - o 3 ct P Q IR p 3 ca cot ct ? U I Hb i NEV 016591 WATER PCB-SD0000030539  \I i R at L-529 Rat A verage i G u in e a P ig _ B -319 G u in e a P ig ' B-320 G u in e a P ig Id e n tifi c a tio n N um ber p P n SO 50 W tri 50 50 50 ffl c* Hi CD & * et PPP 0 PPP ct ct ct ct ct ct ct 13 3 oi to p H > f tr* P 1 a s/i Ul t-f 1 tr* i f P l *J1 's/I sJI s/1 V HO') oo p 04 04 04 to to to O' O) H a O' 1 -0 On CD CD a M ON nX O) <X 04 P" oi on H to O as to O * o o ro H a \ *04 04 3 H st_on P" to 04 (V to w as ON -o ro to 0 to o * nX o o> s P9 * o to H O CD to . O ON -4 Ji P * P " P - O CD to to P ~ ON sO VJ1 sX sX 3 o tJ o 04 (?) ro \D (A > H* ct a tl i-1 O' o o CD O' ON O 5> P* H* o ON t-> ON O CD H 04 o o CD 1 f* i O' s>4 1 s/1 to a a a a cr a a CT P 04 to to to to to to to s/1 51 f res tl S O HO1 pop > o o 1 to to H to to a to -4 -4 to O' 43 to ro o w to o sji to P to sD 04 O M H to P" to ao o *- ro c--O o jo t- p. ~ls H* H H H H H H ON M -4 |*P O cc H M to CD ro s/l o o s/l o O p sT sH jq 3 H* O H O p 04 O' s/l 04 p p P p s/1 73 Or--' 04 Ln sO o sTl a O' s/1 tX H sX 04 sJI H to H D to > H* ct p >4 9 S??S? H* 3 t* ct 00H tt H H to to H M H H H to a *X 04 to a O' a a -4 a s/1 p O o H o H jn H JO o O H o O o o o O a -4 04 O' o O' o o 43 -4 43 p O' O'., -t4o sO- sO -4 O' -4 O sO O' sO to a tX 4 Ct O Cs W o(-*. O T O t= P et 'tCVrt oQWiPCO?C,t ot* HO o t* H t* O o p t* < o P & *4 a t3 (a Ct Hm -3 to p CD w i I i f i '' NEW 016592 f WATER_PCB-SD0000030540 -J ,i /l , , ' 1. \ l l ( / 1 1 i l , . 1 opft >1 Op04l acpt apct pCt pct acpt a Cpt a Cpt a Cpt a cpt acpt o<s P t-* jrno Ppi I {O P jpn1 0to4 OP1 l rruo ptP1oi p pptio> o ppI i p Opi l Ma ppI i ^t-3> ssPriOOo 9 9< ** pp a p p ap % a Op' 5Op3' t+ Hc*t Het* Hp* -Maa0it 0Hi4 P P 0PMpi44 0Mi4 rPo WPOd ) PJ_*P9 <> pCD p 9 7H9P1 Hx ro 21i0s 0. __P 3 O' P ct KOI* *< 0 H P I Pa Mc* 0c4 0e4x a 0e4r 0a4 0a4 0c4c 0a4 PP 0e4x CT p 0044 0a4 0a4 pp -p0 & ro P ro O04 PO OOn* ro lro 0tfou4 p0to1i to tao troTo ro naJ tMo ottoo rP 0O4l 0-o4 rao roo PH* a pi pM* pH -oprooi cc oto> H as Pa ji o otCHst Pro r--1 t- 0M4 p p oH Hto -v p o Hp 0P4 pto o -p0i OpoN Hp c pa P pIVi to o c: H CT to to CT >c o o p o o o v) o tt m w" rr on rr pi -J pH 04 to f\J to p P P" oH seOx pH ko p pp Luni p p o oo pa 0sO4 Ol 04 Oro' pO' a P pa H -0o4 M ro ro 4(O p H 04 0 to to p p 0 ro oH i <o 0 p p | p p i p P o (O a sO PI ro p p P o a p sj\ p o p o ffia ap ra *11, O P O' woo Pu 4 ao *> O H> o o H O ^ oW *1 tel P CL P orno ct P- 1P1 IS *ia o ro 2PS oftCO4Ra3H-cWit te a p p PctpOi SgctQ - MM O O*-> P P ct oPo *- a 4 p I a tj2 o 01 aCT O0 -0 o o O 0 a sC PI Q ex p5 o o M* -a -oPnpPs po04ii o v> vOOj o -0 pi pi o -Pa3- o 0p4 P o a vW o ooooo PJ er p 0044 p 0*4 p p 04 -po oa -j oo d a -4 PJ a P> as ct o 3$ W o * a p >* B Hrt ra M prop 'S'o s Po <4- p, ct <4M O M4 8 NEV 016593 WATER_PCB-SD0000030541 I k OH C pi 03 & ii fp- 7 Kw 37 aM g1 P| 3 W& 03 K^g , o C| 3H*! 3 ffli P ON Oro P HP ^n* H $s c# *3 J 71 ofsi P9 3 nji 4* (R cn 3S 09 H 20 P< P _ ft 3 O' H- ft Of 33 H H I m m CEKO $ CD CO ON CD o M M ifOJsrOoivMse1 CTiJi ON CD N* M oCO fo -0 CSS ON CD ON ftdUbc=So' M1V1 o.ro \(=~ ro K0 ON C D Kn oK-i CD w roiru[ M rvs cr-o G.Moin P" -O ON P" N3 I CD Lac4Ec- E OkN F~ H=* ON On o a 3 33 H* O' O p. 3 Uo oH 4 4 03 CD O fa-Hn>**P** g o o rn 3 ft o sg ft O 3 S3 O3 P H* CM H- ft p* tup<r hVostfo ft p-i ft 4 X' O O >-4 Ho 'll o PD <71a ojoJoj rufof akji 04-0 oo CD .ON fe CD M CD CD O On M ^ m H> et fe tt P<X H> |709iJ0 et pi ct XO OH HH O O 3 P o' M H i 03 *2 NEV 016594 WATER_PCB-SD0000030542 w pct w cpt SJ cPt p ct Pct SJ cpt >< CD H J1r* -0 Pa p1Jl oH- p1jrt P sp<jl H P JP1 i H jPn1 Pro a o O' CT a P a P a P a P 8 *t3 a a c *8 o<D' OP ro 0P4 r<oT a rH^o3 -or>oo rroo o oron IO PJS ta a^> ott H-p. HJM 5 O o - jn sO ro sO sO fM\J sO OH IO a H o o & o1 p O l P sM vJI 04 04 H O P ru p O' si a o- -s0O 04 H M ro M H ro ro ro O H Ol SO H P ro gcto^a OH. O KI fHfi *cPt tn p ts h* ftr o 3 o ct P* ct v)t( O O H H) O O O O o O O o o -ao0 -o PS s/1 SO sO -o0 p sO SO Jl ku 1* P a saO oHa!- wpct P H* 5 P. ** P O' ;h p x P n sn NEV 016595 WATER PCB-SD0000030543 (g*s . 4- jx g ) (gu Tc) I ct HruHroHruHruHroProHroHroKroKroHruHroHroHioHfuHfufHorHoal B v4n^v^J4iv^n-vpn`v-Pnv'-Fnv`-nPv_-nFrv`-nP.*Cro"'.rno".Tpo:v-frri-v.pn-v4n=.-fTco".Sro1'r.otr Hi w' w to rspBscceEsssfSBEscsed? ,,aooo&l,a330fflooo^, a |W *< **4 *1 *4 v* m sPCoTCp4twP-cstPcoroC(-sPotspoa,cHo*apcotops`oeHospcotPcsroCtfocPestocptraHoo-pcat W F0'SO ct p H0ct'SOp 0H**3CD ct p 0*3O ct P O' 3 Hct Qp 0Hct*-3p O F* O a a a a ha a ** H* H H* t* H* __ cs__ __ e_____ to__ __ E__ __ E__ -P"ono-P"cvo-P"onovj4 cr-vo-fr-ONOot owo| $ ct p Oa ct v4o-*-p~tu 0ro0o4vdoHi-ptu Hc->xwi . h NovNn HroNroNvoHh -->a o4 >< a* P Hi tR t-*C? vn CD0VMO4 O COOvH ONO vCnDOVO 030 OvfOfr&2 cl- Co5 so 04--* P Ct 3 H 04 04 04 04 \d cvrovo ro rovnvn ro o ovtuvnsn ro o onM .> 0vd4 G0>'J0r4-0v4c cCvVeOt4v-h0 CMDOoVj Jr4o-Op"-P0--v4>4 eMo\p40r4o^O-FOnrb' q'OH- < O ffl HRQ.3 $ t-t O 3CJJ CfOt V M3 3ct>*,C5H P*! ?' CD I ro o *o oro4="-P"0 o o o o 0 044=^04-p-.pl| s? octc3n s Wp. 01-0 004 CDOvVn H-P"C-0-P'a>-JOlOIO4 O o h r-,oi^^-1--o4 O6>voO---o4 o4 h-P*-P" >0 030104 OcNaCpTfi .teroW,,PSI4tRr* oCHtJ ct Pi (tD-X <<1 ct o --o *-t>! o *n 0 0 000004-4 004 4-*OH 1 t 1 1 1 i vwvnp-.p-.-4vn oovwvnprv: OhM-F-*~vhO--H4VrJo1CpD"HfO OOVvOD Q6C,Kr0o0a4 04 hvo--4--4 ro ro o ruol h 4-4 * OO \j v vVi-v/V Vah l/w I I I I I I 0 0000I 0 0 0 0 0| 0 0| vjiv/isjivj* OvJ'^nvJ' O vJ^ O OOOOv*n O O O H ro O Oo ,K4n0 NEV 016596 * WATER PCB-SD0000030544 HHH _J->_0 VO V3VJ r\J h ro Kj t-7 p" NON-XWU'v.IlrN-^W~o>.f"vXJ^ Vm*. vJivnvNvnvnvnvnvjv P"-P" \MVMV*V4V>4 S? ft hhhhmhm -OVO-J ONCD 030 O O OVJ OVJ1 o ooooooo cvovp-vnvji-ovo OVJl O Ovjl O o o oo o oo o MH HHHHHH vn -0 V/lVW VJINJI -J VO O O OVJI O O OVH oooooooo .- H H tit-1 H H <JN^X-P\>4 CQNO * * * ovjivj^vjt VJ1 oooooo o f >ct s?Ol *d 3 H* Ha o t H -0 3 ct o ct o ct Soi H* 3 (B ro sF> >g 3 50 O % Po H O' H* M 1 o' O 04 H* 3 O ct - O r Hu H 3 W- . '.sO. .-"tu - * rMo ` i ON CO V>4 H 1 04 -0 H w - is o 3 2O5 No ct a>, cr 3 o o H a. -ip & ft cf O o <-! ft `JS*o h'hhhhhh OnOnP'.P-Cm ON 03 -0-0 O OVJIVJIVJI vjivb o o o o o H 1 04o 04 I ( NfcV 016597 WATER_PCB-SD0000030545 Table ao> * o HO Ct c+ et * *1 O o -< 3 ro VJ1 P i VJ* l KO n & 9 fV) NEV 016598 WATER_PCB-SD0000030546 HHH M H M W M ONOVONO J\J H HNJ* I ct^n ro' . O. MVJ M n/tVjtV/t'v^tVj'tCp'vjtCtt' P"-P'4=" VXVXVJ4V>4V^ l |S a> hhhmhhnn C DvOCO#-~J COCTnOO O0V^J13O^UfIVu--HoV-WH oOOO o p ct > b~ kn 3 <tBr 0 1 O ft 5 o 6 H.H H HH HHH vjl OnCn NoJ1u-JiNoJ>\oJV0oNvMn Oo Oo H 1 OJ O' HHHHHHHH ONO1 ONNM-p-VX On On ovn w ovjivj'vji o OONJIOOOOO V H H H H H HH H H vji4r--jvj!v>i.prvn on On ** O lu-j-^j'jnvjujl O O Oknvjikn o o o o o H 1 S>J O' c SJ o* o' H H* 1 Ct 04 ca a<x o o ct ct H* s oc H* 3 CD os CD O C 2 p< U r-- oo aH*1 HHH HH H -Jnhvji.P'OnQO o o o-j rovn O O O'-nv.r o H 1 OJ O' <L> r\>l ifl Table 10 * l S P S3 * o ro ONTI H 1 < H HH H H HHH H VJ1" ON ONNH ON ON OnO CDHUINW fO H On o-P"hnx o axnvov* >< o P n o SL |S ta>r NEV 016599 WATER_PCB-SD0000030547 Table 11 (1 ) D e te rm in a tio n made 15 days a f t e r 82nd p e rio d o f exposure. 0 p- 0 M t-3 CD O O < 3 CD CD 3 >w P3 H O 3) O ffi O O O ft 33 + O O' M *"* CO IV s-n ct p0 c3 CD w 03w 3X s d ct H* 0 O' 3 u CD 3 3 3 > O CD a 3 w0 p *-t> 3 CD O 30 p ct 3 ct 3 03 * H3 U 3p 0 a 3 ct CD 0- 03 3M O MP 3 P iO & Vi O M* H* 3 O ft 3 0) c a ct 3 0. ct a> tt P ft- - 3 vi 0 ft u CD n< 0 00ft & CD 3 s ro CD P x0s 3 0 0 3 0 3 > ' 0< 3 wO p3 O ct |- a, 0*3 p 3 O 3 M 13 fU 0 H- 3 CO M O O P< NEV 016600 WATER PCB-SD0000030548 Table 11 - Page 2 ( 2 ) D e te rm in a tio n made 1I4. days a f t e r 8 3 rd p e rio d o f exposure OH O O > S3 PP '3H H c OHcHtr-] na X M0C3ccHat*t H O HCO t0* H- P Pw o*dX 0t-o MP O(Ctt M PPs Is Oa 0 n Mte OO P- O H* M- O O P P- ct am>pT' 0h> pH-pcrg0 g Hc>t Ho*- tO00-*0 h P3 OP PPH* PH* OM0O VHH)> f3fl 3 0 P.3 O ss o > 0 0o H 3 0pOCO. pH&sp>* Op*C0H0t DHa>* 3 pa09 00pCt p 0P 0H S3 P P O P HCO HCpO PO P. M vwn F" Pct*cXJ OPT O0 a 1 p. VJl k 4 TJ PI CO O OpH>* PMn- 300C4O S SV* cr 0 (T 0 K a (a 1 0 H- O 1 P cr os *>* 0 ct p H 3p 0H0 P p. *p6 gSe1S W S3 P *Xd h> Ha* OU PCt p t* 0 Hoh O3 ct CPO O3 n hO* H hP-o35 0Qi H* p .0 NEW 016601 ft WATER PCB-SD0000030549  5'-- VTl f= 0 3H- & 33 aa 99 H*t-) B H- OQ m Wa 1 vx NX tO ro -J O' M O V $ 31 V3 X VX NO $ N VJ1 04 H > M >. VJ1 -P 0! & 9 * H<n to 1 vx StJoI SVO M ^ V VJ1 Gr vx 4r .. I d e t nt io N um ber P> if n i c a SD V Jr oa oh- o. ha ct a vji a 5 o to a a p. P ct VX a a o a ct o H -b HH > (O OSD a wwh a mso o> m vSksOT a vxnxnx M t-> 4-* H* SD > ro osd > ro osd tJO ` 1 >a 'ru'ruHi" o 'ru'ro'e 3ap P ct ct Os a H SO CD a ro co-o ct a a N NJ1 3 vaowT <s GiCjiGr H-a o a o . VX a vxnxvx vxvxcx 9 xh % ao 9H I a ct to o ro > 9 oO H o a 1H* M H* *-- vn vjis-n ~ -P" OCDCs M M i- M H NJI .cr ir-o\^ H4 O CO sji ro4="CD so H* M H 0 0(0 * onx-F* VnS. B*a HO OCT HO H* gcor (- H- 3 . i O s CD NX 3 to tosjl osOsvJi %% so oaw -J vx ro on o ooo ft ftVJ1 -ft OS OsOsNJI * % Si > -0 -0~0 CD % CD O Osnpr pr hho vx H -p--P"Qo --J CDODVH -9 S89o O OOO o ooo ao gpo W* mm wtt SO3* S' VJ CD 09H os os rosx 4r- vx ooo NX OOO v ft ft ft-* wOs -0 VJ1 M HHH ro HOa H % 1* OS (OSXSX h o ovn H4 H H ro os sji ro o (O i-* s0/1-o0vSnD tr* t* CD O 1O o -J ooo CD OOO O 1M?2 a H+a 0 act 9a M3- J ct ct a 3 4 ct 3W 1H 9 ct H 3o O a ct 3" a <: p od a o a ao> a H* O a ,ro oa ao> o H Oa ,ro Kn NEV 016602 WATER_PCB-SD0000030550 I j \ i \ r I &!.2 aM s o p a PI oi MQ, e-la o C et 3 p i M Ol VMl M cq 03 C3 0 03 p 1 a; ji T* ttl to I VrMo, VM, vd VO Bs?s*g mvn -F sol Mo M \sJ\ \ P VVmMte vn VM v\n F vc ro S\ $S N N. VVMT vn VM so vo] vn VM F VO gCO oo>fp eto* W TWO, VJ1 O On ft MJ or w *o >1 H* H 53 ct > < ro o T >D ro ovo > <: O T .ro ovo P 03 Q VJ1 vm wo SOP P ct ct its a oT 3 I s ro r To o M 3 ro. ON MMM IOVM M -J-P"OS tc M P- l, x vMxvMn Mro 4H=- ^MTVMMvMn VM VMVM ro V O ro l-j oapro m iro m a - S a*' ro I *".O- v 03 \M 3 0 ,p Iga> ro ON VO ONvO o * vnvo-o VO o VO 0<P, ooo vo ko voovpr- VJ1 vovovo vo vovovo VO -a o V --JVM VO CD .p-~JVM o ooo VM VO --4 KpjvoovMo a> ooo 1-4 own ro toVO VO ooo f5 cfl o Mp o &* CO . cor or p H MM CD oo feOVO VD SSf=~F~ OOO ooo sr toCD M VO O CD ro O CDOv vn --vo vn OVMVO ooo ooo Oov o VVOOVMO --o4 ooo VMM CDMa * OV --4 vCn3vMnVoM ooo te* goo o 3<4 1 Ct mO w NEV 016603 WATER PCB-SD0000030551 p i VO OS P- OHPo-'' 0w HI VX V*JX -Jl to VX 1 o. |o Krt- M fOfi' 3O *H-*> *1 H* 0 V-JX P 1 H sol S1 u o *t--* VJl VX 3 3 2O SO 3 o o Vv7x1 M o to ST vn oF" 3 MM ro to >Ms \ M sn vn VJ1 -F- N VJ1 S V7J vx VX M VO to \\ H M \ VJ1 VJ1 VX vx \o M X\ H M Vs VJ1 >vn vx -P ><4 to JOjOSO oi top W 3 toP VO a o*Mct OH ** S? V IIOHB# O 's O0 to 4 H* viv to vx I 3 0 gS'S? P 00 Ct 1**1 030 S< O to > oo1oH* Os ro vx h h H OsOs , HM [TO VX H p" : H os i- so MM H ro h ro p-'-F'O tt ^8 MO 0(3 OH O I OV CD VJ4 4 to i to tS. M tf o vvo 5 vn -P" a op Ov Mi to B VO O vn assn < * IJtON *-^V%J5 f K SODV~O] oo osvnsn *** OSOSO vox-oF~oo tr 1 u p 4 k t H vtno SooO v*n to03to0CD o3 llo to oCOD o ON H ONp"H OVJ1VJ1 ooo OosVnO OO os Kx vvnxsoxstno kx OOO NEV 016604 I WATER PCB-SD0000030552 0* <n a CO i VIM vm ru VrMo ru VO O VO R^ vm| vm R VJ! -f=- o w u I VM IU I-1 VO y VO vm 4=- 0 Hp a a P *o H <R td VIM1U O VO VVJM1 VO 4? 3o gpi 2a a ct 3 <D p _cr HO Hrt- aan H 3H < CD p W I VVHMO VO ru \ VO VO VM 4= m3>O39 t* P, roO Ho H *i aa p Q* P rt- u t O rt HJ a vo \ru y VM ru hvo m ru ru ro ru iu i* IUHVO sa0pa cP*ca+ art Ho 1 3 vjivrf 1 vm<34 35 H t>VO O V* ko H VO 4="ovov H HHH vm vn-pr ODCDCv ko VHMVHOVHO poo HH IU VM IU Wm p-o| OCT oH o g <r- If a s H* 3rt ao o o H 0 *d Hru vo 4=1 VO 3 l p o* H a IU n p fga r.. . r- 0v .ru 8?-oj VM-0lO ko So oov- k>M3 O'O'VO ko VOVOVO Wo HHr SO VM OV-J "o4VoOVoJ1 ,00 t-*vO 0v hJ ooo VOVOVO co -m3 ota vovo-o CM CD & aa C? WsJ Wm od ca o Po 3M pu. art oa I w >p V4 CO VO o vn oO Hru Hovcrv * H 0M>M Ovovn OOO IU os VOVO M o vovo o ooo tr< ro ,ru h^ocn ko VM VO o ru1 ru.ta3o ao3 O VM a VOVO o o ooo 00 vnd oo] 1Ta tt NEV 016605 WATER_PCB-SD0000030553 nsszsssscssr oo H3> 3 H- O 3 H> TaO 33 TO TO 3 c+ 3 TO H- ct H 04 CTs| 04 P 0I4 on| 3) H* m to to P 3J H* OP td P O' O 3 H* H 3J 3 H H* O 05 P t tfl 1 -p- p- VN H CO VJI VO ro VJI VJ4 \Ji V4 p 04 04 VO to o> 04 04 -P S N to OI 04 Sr p VO to Sr > VJ4 04 Sf 04 04 sy;T P . . - TO 3 **1 . fa 3 & H- t| SCI O H> 3 TO TO o H w TO U pip ct 3M TO O ct H> ff d h* 1& > a< ro h* 3 O <B 3 0TO Op P ct ct ct TO TO H- 3 ogo 3 H 1-4 1 > 3 OTO H* % M MHH aW H MVN 0H4 0H4 4M=- ro ovovp- cv H-p-ru ff tn aS? JM H H H [p- 4TVXVJ4 3 o to ON ip- onFo fa kj4 #* tO 050 on OH O 3 o' ro VJ4 P" VJl fe 3 3J ,P &TO KJ1 *<rv 0104 0\ o -pNO-sJ ON -4 OH -4 O O O ^*3W OV 04 ON ON NO V4aO*O n Klan VJIVJI a *a aOV 4Oa v4aG'4taJ' sgu ro NO O CD CO -P%>4 ovrooi HVN ooo oo oo ~JNO o H* to CO to o HOOOI ON 04 HO OOO 'g? 3 os o 1 fl TO TO -- TO p 5o TO "c P TtrO* to 0--4 < < H 04 HHH vm ^rooi vn-pvp" 004 O ooo ov o o M-P\>4 OOOOOO 0oo4 OO ooONooQ hto*rroo M4 vHnoHNHp- f3aTO 3TOO VO VO oo H 100504 CN ooo o ii o oo -4 OOO 3 cr uO TO NEV 016606 WATER PCB-SD0000030554 P p cr cr H- ww pp Ocr' h O' O' t* w p o' cr H* . ct ct ct ct so p M a cr ha* ct 5fS 3 O' ctS H* ft O H- ffl 3 H 1 cr M 1 --4 H 1 vn 02 M ac H t N4J=1" M1 4 VS a M o p 1 a -"0 vs VO <I VJl VS -P > < M H* 4 ro hvo p (a "JvSvxrv? o 'V'xjVgJ^lr'Xu, VVJSIVSVVJS1 H H HH O MOO VJ1 O OnO O O 3Q O 3 o. M H H MH M H M O o o o vo \ m "r*soS VJl -fr V, S ro ro vs O v VJl vn VS l Nro 5\5vn sro ON \ vSX, VsVJl 'JV1s vji .J1VS nS.. Vs vS X VJl vs H X Id X VJl P WMP tJ Sbsup O 4 P. M Ct VC O M 4 0 'x ca o tr1 P p op. P ct 4 p O x O O Ct l-l VJ tn u 0o 4H* 1 3 ct HH ro O \ro \ vs vs o \\ VV>J4I VJl vs VH>f H H CD ON > tJt* hh BP OctPct NO 4 ro ovo ct O O N H* 4 H CD n rovs h VS O CD 03 3P-04 X1 VJl vji vji vn vs . vs vs vs W wo vHn o (-* ro xi t-H H ro rovs 4="-PN=" \3 HO on OH O a cr H H 3 > 4O o H O4 H VJOU 4=vn CS -T a b le 12 Page 6 % * IB roo *3 ooo ro vsro vcsd vonvonvon %O> N ON V ON VJl VS o Oo ssnr*On O n *OnOn ON vo vn vs oafr 4 0 3' o 3" ovovs C4 o o ooo 3 p O | Po X34 & ct ca CD wU VO %Ho VS O vCnD Oo 3* CD M H* H dB %I-* ntO OVS %ON on 4o o O 0030 o 1vn o n o o ovn ooo 3S st a . NEW 016607 I WATER PCB-SD0000030555 Id e n tific a tio n G uNi nuemabePr i g B - 3 1 3 OFa ti er ss t o f and Last P e rio d o f E xp o su re G u in e a P ig B -3 1 4 o0 && a8 P ( *t> 4) HM l <w tn to t VN 01 MH -0 C a & 8 0 "0 H* 00 to 1 H vn vOSO M H SO ro so \ S\ N\ \ vji -F wa O^s v>i V.VJI VnSN OJ h VN r> SJ1 -F an "" SJ1 ^> SN VJI -F a <] m SN VJI \ VN VJI F oa V SJ V* > < a mm 4 TO MsO a ^o'^ru' VJ]'s>4'0 V>4 SN > St > a0 < < < Sop a HH O HH <B MM p tt <t g ro hvo 4 to MSO 4 lO H SO ct to ffl s ro'^'ru 5 > 8 H4 03O Q SN'svSN O IOSOI VNWN 3 Ph VJI SN VJI VN VN wji\ VJI VN VJI VN VN '^.vn's, VJI SN VJI VN VN H HHH F vnv'-v.M * vn O o+r- f fF? to . toOsCD H or h 1H H 1 -pro 1 ro 0 M HHH F shsnvn * ro 4=TCDpr. HO ON OH - 3 o' - HM 3 os CTSCSSn ta H sn snsovo H VN VoOoOSo1 >ji snsnsn W *** os snsn-4 H -F*HSO SN O O O J1 V0VJ1VH to u % p- to-'J ro 04 SN COSO 04 0 a 0 sji sn ossn * ** * oo cao-p" H -0 COSO 04 OOO W o'" "S Kg* Sw 0 gPo up *4 pi ct tfl O wtt rf i H v OOl ON vn Fp-0 o snsno o ooo M sn CD M M M. -o-opr V O OSO OHI ip- *sMo*-Mjr-*Mfo ro co r~* os snsnsn 0 sn osn -a ooo O OOO Hro F* HHH HSJ1H H O (O oa snsnsn 04 OOO S* a is 00 *8 U CD U i NEV 016608 WATER PCB-SD0000030556 -w 8 H ct M 1 VXI cr> Cn W P O' o* H ct M 1 V* O' VJl I a p n H cn W i VJl -0 oS I-* \o H VO h HH ro N > ^ > ^ \ s, H VX V. s VX - >. N VJl N S VJl 4VJl VX VJl VX VJl VJl t=" vx Id e n tifie s tio n N um ber G u in e a P ig B -lp 0 2 H VO ro -s M Vx \ 'S. VJl VJ VX VX 0 a H a p p *Tj H <n to 1 vx H a HITS** B 1 p* H (t vo oh a o u O H to ta C 0.P ct *'> VJ .4 O ct O ^ Vx * > < HH 3 fO HVO m ruvx to VX^W > < 0 HH g fU HVO T rvxx ro o \Ul\ VJl VX VJl vx vx > > O t) < < 3 ffl P H a hH ro a ro h*vo 3 Svo P ct ct Ct > :n 'to^'ro ho 3a o vx VX s*JX v>xx a h N WJl'v, VJl'"'> t VJl VJl VX VJl vx vx vx H HHH H NHH * Ov '-4-prjr- H HHH ro vx ro h * P" -F-4=-4=- w 39 Hp" vHjivHxvHx H Vx HH rovx HO * fO.. H -P"CDH H on -J-P" O H a cr HH a Conditioned A ir VJ1 VJlVjlVP ts s Os ro-4 CO VX COOsOS VX OOO vJl CP*Jlvjl A S vo ro ooo O' OOO -3 OOO VJl VJl VJlVJl VJl VJl VJl VJl % * % v CD P" COfUVX P" vxvn OS vji vx h ro K> ro ro o VX OOO o oo 8 9* IPS' ^ai amo |sa tt o wCft H HHH VJl vx osOS * -P" Os -V4J1VWJ1 -op- ->3 OOO H VO H CD CD < * * pr P^-hvo CD VJl O O VX OOO f a t\oo H HHH O MOO (B V S HH vx -4VO V !* V t> P aa tJ o vo hoh P" ovo o OS VJl VJl o o -0 OOO vJl o OO oo xao Cft NEV 016609 WATER_PCB-SD0000030557 I 5:1 H P* 0 cf a __ H* Cf O' Oa H*->* V1 a h* i t> t vx vo 1 p 1 a M rv> vx vjT VJi VJI f* VX1 -P VO VJI -P njp aa aa- *t-*vi cp* S3 h a o oa P* i 3* co ft* r><a ro a > vx a [<a VJI vx HM ro hvo VO\x''I\3' VJIVJT^ VXVXVJJ S?S? P cf cf cf a o wosa o a P- a> ta vx Hro H m Hro Hro H H H M Ra M , MHH HO OCT OvO-P* hJ 4=--P"-i=- O H O a p* o o a ef (V) (-* Oa i a a P* p CT a No On % Mi * 00 VJIVJl <J\ tv><v o4=- M CJV owjivji < ro o ov CO ovovo p* P"OXX o M* o o o |VX o oo 3MO gPo w3<3_ Pi Cf UO wO VO ro owe h V HM VX OV0 a AO ao Ov VX H--J o MVO H o o o ON O O O vx o oo vx Oo Oa Qo Bv) 0 cf 2 NEV 016610 WATER PCB-SD0000030558  r- t IUM < PP MM o3 pC to is tSt* H P* i* 3 *3 oo Pp 33 ft et fH C3 t-> 3a <to9 p to ct ct rt 3 ET* 3g* t?fr Po ft O' ft OP cr h> ft 3 Po H- (3, 3 oo as oo 3 O ft O3 -3 O ' Ct M .3 W o H* S'? r* tD CO t*T*1 cUq 30 OI OP' HChO*' 3at cCOt p *G QCqO SOP3' HC53HC0CO*t*3PP t*o OCqo M 3 CD O H- <t> CO o *3 > 3Ufc a p H CO V^JK^J0 1 vOo~(\jj ONVJ1 Vo: cOoN aO' ON^n o->i P09OH* > ,< Xta 3 P oq B <d0 o o if S3 ct- CT\ C o o ciHCO a6o< tX=) p i? o 3H* 3p 3ct 3 SO3! MM M l\J VJl T<OCDOl a cot o o 3a ^Mo Mrol CMH3-VNJ3 *3OO> OH3 \_Hron f=- nOVHO Ul H ro o3OH>o3'.. Hro > < MiO 3 P Oq o FCO*t 3<aD r"; v*n on a op- H .i ` On 09 vx 3 > 3H .- Hro*vMx Mvo !robr" 8<0 rovji ru f-4 - -V : 3*"- o H'-P $CO > Hip 3 N gP* P oq p t3W P3 0<-4 get co ffi 3 3O ao O P p tfl 3 pi P O 3 a> o 3O o 3 vj . O'. MP ** oq g5 \3 HO ocq OM O 3 o' H H- 3 VJ1-J aon 3 lI*P-3 P > !p<! 3 IS a id5S p 3u O -4 M 3 *r o 6 to 10 p` .3 }-3 P 3 *--* P M I > < P 3 P fPe a o p 3 ct 3 P ct H* O 3 O*3 NEV 016611 WATER_PCB-SD0000030559 ( f c+ (1 XI r 1 XI to 33 Q P0 3F o' M O3 ct P CO F 03 CO 33 3 PC O' Ho' 3 H* O ct P CO *c F 0<t CO >W 3 o 3Oo3 t-4 t-* a ]u to H ^ovx F CDK 31 F3 -* 3 * N t3o 3 F + ot O FSO H >4 O s* OC CtoDcH oc OOP- o 3 H* MH to i M 03 X to ft. a ! oo a cl* o 3cr OVX FI O1 oo s o H cr* oc 1* OVT c * ro Oro-Po" > 03 o > 3 FO FO vO\X o 00 rv o 3 !( H o FO 3n 3 H- + 3 to 1 M ~ oo H to VJ1 h 6 tv Fro onvji Ps-H"HM P" 1 p--0 c "OVC i -0-0 H'P' ON wV noJIVcJI oo OP" 'Nst *H* o o HC OH 11 O O ro c 03 VX OtV H --J novm CO HIV I? F IV CNF 3 Ol H CO NQl p 'A O O) oOj-vpj-1 ccsc ON0- o. P 4 oc ororov ii o o HOVOjn V6 O O 31 O OH 3$ gS S* 3O 0< O (a \cr 3B tt at OS* h>3 FO 009 OH O 3 o' FF 3 crfa rt-ro S'1 3-R NEV 016612 ns o J) e >3 o _o 3Wo ATER_PCB-SD0000030560 W eight, Grams k NEV 016613 WATER PCB-SD0000030561 I Figure 2 NEV 016614 WATER PCB-SD0000030562 W e ig h t, Grama i t I NEV 016615 WATER PCB-SD0000030563 i I- NEV 016616 l WATER PCB-SD0000030564 r W e ig h t, Grnnw NV 01661? l WATER PCB-SD0000030565 Y /a lg h t, Gram3 i WATER PCB-SD0000030566 ^-- f t ./VAL. v5WH. L W e ig h t, Grama Time, Weeks r.'zper.Vinettt No.l 8*6 Mlcrograma of Aroelor. 1242/liter Figure 7 9 NV 016619 I WATER PCB-SD0000030567 I t \ 'i i k i NV 016620 a1l i WATER PCB-SD0000030568 I W e ig h t, Grams Rabbits 1374 to 1377 /\ 1375 Kllled 4700 //4600 4500 4403 4500 / / _.........\J j r. V /\ /.\ ... 1 4200 iru r ! v y4100 i \ /. i r^' 4000^ i , .1 ! !' j _i f \ i 3500 i f v \ A. 3800 * * tx / . ... /V3700 : * 3600! / ! 3500 i 1 - 34001 A/ ^--Period of Exposure __ * - Time, YJeeks Experiment No, 1 8.6 Mlorograms of Aroclor 121(2/11 ter Figure 9 NEV 016621 WATER PCB-SD0000030569 I W e ig h t, Grama 2750 2700 2650 Cat AM4.5 Al+1+5 4 Killed 1 / 14 / [ V.' /; .K.//\l ' </!1 2600 2550 2500 2U.53 \ 1 'l* 1 .jj / f . ./ ! ` */ il ' 1 j< 1 \^f ' V 25>" 2303 ; !X 1 ; ! /1. r' ir a t) .. - ._ -. - .......................... 1r . .' 225o 1 i 2200 2150 m .......... rerloa or 1 ............................. Exposure Time, Weeks Experiment No. 2 6.8 Micrograms of Aroclor 12li2/litor Figure 10 1 - fr NEW 016622 WATER PCB-SD0000030570 W eight, Grama i Exnorim.'nt Wo. 2 6,8 Iilcrograms of Aroclor 12lt2/llter Figure 11 NEV 016623 WATER PCB-SD0000030571 c u in jo i NEV 01662^ WATER PCB-SD0000030572 W e ig h t, Grnns 85O Guinea Pigs 83^5 to B330 and Bl+19 800 1 750 Expoaurs . .1^''*^, s, :A ................................ /r . 700 650 i .. A.V ^ ,/r H'+V ,J 600 550 ! 1 \ y' .1 f . rp 'A MA 7^ \ /; 500 tr l\ 1+5 0 11 ! ./ u ^u .v,f' l+oo 1 l/ J 350 J"'1 j / .. L..'| j... r , i-t'' L -I-'4 ". Tine, Weeks ..... L'/j:perittient No. 2 . 6,8 Mlcrograma of Aroolor 121+2/1 Iter Figure 13 . .. i i i ___ NEW 0166/5 WATER PCB-SD0000030573 Y /olght, Grnms J1 Mice B21+1 to B250 end B376, B464 --------------------------------------Period of------------------------ -- h-- i F Exposure 1 /' / *A 51 \ i . v V \ Ki ` 1 \ i . 1 ` ii iVi' ' 1 4i (\ ' * VJ , _____________________ __________ L/tK t =^1,, .A i vr1, 1, i ------------ 1 --v! J|; 1 i' J , i \* T IT ! /IX, t i K/ Nr i \! X/ v i 1 * i 1 c i1 .1 \l v >. t-i : i K,/; :1 ` ' 1 / V| *i i i ... ' - . Time, Weeks 'xos.rijr.ftnt JIo. 2 6.8 Hicrograras of Aroclor 1242/Iiter Figure ll+ NEV 016626 WATER PCB-SD0000030574 W e ig h t, Grams Experiment No. 2 6.8 tflcrograras of Aroclor 1242/llter Figure 15 NEV 016627 WATER_PCB-SD0000030575 i W e ig h t, Grama I E NEV 0L6b28 I WATER PCB-SD0000030576 W eight, Gram3 Rats L526 to L535 and L836 320 i i 310 /t-i 11 300 /I \ A f 290 / V! \ 280 j i; 270 1 T / r. .4 i,,4 \ --*"1i n___/v______.________________ i i ! \j ' Y-x- - i 260 < 5\ 1 \r m J *' j 250 ' i li+ 1A\ t 'K , 1 i240 i 230 /\|7 1" vi 4/ . .'k"k ........ - ii i HH ! 220 ____* 1 vi____ "7 < __1 * * 210 k\ 11 1 190 180 r , i. , J rt xw xj \ j 17 T Exposure . Time, Weeks "speylir.nnt No. 2 6,8 Mlcregrams of Aroclor 1242/llter Figure 17 . ' NEV 016629 WATER PCB-SD0000030577 ] Rabbits 1370 to 1373 and 1964 k ppN 1370; tIt ifTT killed '1 J. f Ferlod of - -j Exposure i W e ig h t, Grams 11373 _JCL_. Killed P* //!N J j 1| /4-" ' ... -....... 3200 3100 3000 2900 2830 y/ A ,J-*H A pr------- \ 1904 `- Time, Weeks Usperircent No. 2 6.8 Micrograma of Aroclor 1242/liter Figure 18 - 27OO 2600 2500 214.00 NV 016630 WATER PCB-SD0000030578 i W e ig h t, Grama Rabbits 1370 to I373 and I9b4 ItfOO 4400 * ' ' i'erioa or -- "` ' 4300 4200 1 x-t~H 4ioo TR\ /Ij--r1 s 4000 \13900 3800 i ________________ / t )./ t: * 3700 3600 1 1| :_______v__'-_____YS/,_H_/_ J 3500 /' \S_______________ v, 3400 /^/f" Y>J / \/ 330c 1 3200 11-- 3100 a 3000 /\ X ^i b-r ___%Lj;_ ,/ a. r_____ . v......... i A / / '*. 2900 ? \7r / p* ^ ~ _ : 2800 2700 2600 2500 !/!f / ,r> /' Jr - iMy _ . _. 2400 Time, Weeks Experiment No. 2 6o Micrograms of Aroelor 1242/llter Figure 19 J NEV 0I6o31 WATER PCB-SD0000030579 W e ig h t, Grama -t*** Tine, Weeks iixperircent No. 1 5.4 Microgroos of Aroclor 125VHter Figure 20 NEW 016632 I WATER PCB-SD0000030580 W e ig h t, Grams i Tims, Weeks -zperiment No. 1 5.4 Mlcrogrnraa of Aroelor 125lj/llter Figure 21 NEV 016633 WATER PCB-SD0000030581 I W e ig h t, Groms NEV Qi663<* l WATER_PCB-SD0000030582 W e ig h t, Grima 1 j Gulner? Pigs B319 to B324 nnd BI4I5 ^nd Bl+lS BOO r 700 _________ :______________________________________ y\ .. . 600 Hj ____________________ pi ,rH-^ 1 uK e i } ' . i LHPFl TT7 V. 1 300 i41 i ' i200 1rr1-- ------------- i__ - -... Period of Exposure ' --------------------------- - Tlme, Weeks jc'jeiriK.er.t Ko. 1 ` 5.4 Mlerograras of Aroolor 12514/liter Figure 23 I- HEV 016635 WATER PCB-SD0000030583 i r W e ig h t, Grntns T Time, Weeks ''xperlmer-t N6. 1 5.4 Mlcrograras of Aroclor 125l4/llter ' Figure 24 NEV 016636 WATER PCB-SD0000030584 W e ig h t, Grnm3 I NEV 016637 WATER PCB-SD0000030585 i 250 225 t r-Uj Rata LS21 to L52S /]----^---- K"t- '+.. tv ^ ' K+-'^ 21 20.1) Killed 275 1 250 \ i225 K y- - ; * ~i t * t5i2 Klllod 203 g 252 fi 225 * 250 S 175 hO S 5?0 !t 1-- ' -1 Wt '!1Xll1/H~h1--TL--SfI31 \/j Killed * 275 , 1^1 ,.u,i Lj-4-Hdir l 1 v'. 1 l IM 1 w`\ --'I tcp Killed 2`,3 225 f . , J-I-+-1' '1.525 200 ' | '--r' -! 175 , ~M Pr,T1n/^ fir Exposure .... Kill* - d ' Time* Weeks _xxppcerlmont No. 1 5.4 Mlcrogrnras of Aroclor 125lj/llter .. .. ....... " ......Figure ci NfcV 01663d WATER PCB-SD0000030586 I W o ig h t, Grama ' Time, Weoka xp^rlx'sn.t No.'l 5.L Mlcrosrrms of Aroclor 1254/litor Figure 27 HV 0L6639 I WATER PCB-SD0000030587 W o lfth t, Qrnm3 NEV 016640 I WATER PCB-SD0000030588 i W e ig h t, Groma Tine, Weeks ilrperimor.t Ho. 1 5.U Mlcrogrnroa of Arcelor 12514/Uter ' Figure 29 NEV 016641 WATER PCB-SD0000030589 k I _ Time, Weeks .: 5VJ4. Mlcrogrnins-of Ar oc"lo l^k/liter----- L-_ . '- - FlgurB; 3^ r; - ", - v 1 NfeV 01.66^*2 I WATER_PCB-SD0000030590 W e ig h t, Grnms ! t I Time, Weeks Control Subjected to Conditioned Air? for '"xroriment No, 2 Aroclor 12l|2 and rsperiweiit Mo. i Aroclor 12^14. Figure 31 NV 0166A3 l WATER PCB-SD0000030591 i W eight, Grams Guinea Pigs B313 to B316 j 700 KBil5l1e3d / 633 500 ___________ "\ 630 5,33 ri'T'1^ - lt33 .. B314 _. jyS,'r_Jj^nied - .633 '733 603 ' ' ' " "" . B3I3"Klllec / * , , 4. : ,.5pD. ipD ? HcHj3ib Died 300 Exposure Time, Weeks Controls Subjected to Conditioned Air; for Experiment No. 2 Aroclor 121\2. and "xperlment No. 1 Aroclor 1254 .- Figure 32 I NEV Qibhkk WATER PCB-SD0000030592 W e ig h t, Grama lr Figure 33 p--- NEV 0166 <5 I WATER_PCB-SD0000030593 'W eight, Gram3 v j I; WATER PCB-SD0000030594 W e ig h t, Grama t Figure 35 New 0166^7 1 WATER PCB-SD0000030595 W e ig h t, Grams Rats L507 to L512 2^0 22? >. , . . . ..1 1 l*A--!--i r r1 1 " ^L507 200 Killed 2p0 225 "V A1 1 . I --4--i--4 J y k `' 1 ! __ f--+" LyOB 200 .Killed 250 225 :_____ ___1_______ a ?--a-k 200 ___________ , 275 !1:!1- 250 , t i-M v/r^>-vLJ-'i 1 225 _/ ^ 1 200 2^0 | ' SI-- - L309 JCtlled 11 --I--1_ ~ L310 i-Killed 225 200 \A 175 150 275 250 22i> 200 175 7^ ~ 1 ... \ LSll Died 71' ` i-M-HL 1-1! -rV"Na- ] 512 Killed i . .i t Exposure " Time, Weeks Controls Subjected to Conditioned Air; for ixncriment No. 2 Aroclor 121(2 and Experiment No. 1 Aroclor I25U ' Figure 36 NEV 016648 1 WATER PCB-SD0000030596 Rats L513 to I5l6 and L748 275 250 ____ 225 ___________ _L. 1--t ^' L513 Killed 200 11^ 250 225 k , i --k.^^1 4-t 1 __ 1 200 ^ l5i4 i i --^------------------------------------ Killed 175 . . 275 J^l 1_________________L u-4n-tei5 250 rSv i ! ^! 1 1 ' \ ' ' ' Killed 225 l_< 1 200 1 1 250 225 CfN iJ-H-I-------- H/l --1-- 1^16 200 |\ \/\ . ivxiJLea 175 275 250 225 jj V ,4-T r 200 ' i i i. i jr Killed Period of jsxposure . . " Time; Weeks Controls Subjected to Conditioned Air; for Experiment No. 2 Aroclor 1242 and Experiment No. 1 Aroclor 1254 Figure 37 NEV 016649 WATER PCB-SD0000030597 W e ig h t, Grams Time, Weeks , Uootrols Gubject6d.to Conditioned Alrj for . "jnsrinent No. 2 Aroclor 121+2 and ' XT'oriftent "No. 1 Aroclor 122j. ` -- . * Figure 38 NEV 016650 WATER PCB-SD0000030598 W e ig h t, Gram3 k NEV 016651 l WATER PCB-SD0000030599 W eight,- Grama Rabbits 1368, 1369, and 1975 ifrp ' 1368 . , /' TCllled ITT ]J 1 i K ----------------------------------- ------------------------------------- --- ------------ --- " -r -....- 4000 3800 3600 3400 3200 4600 4400 1+202 ______________ * L^J L_U_____ :^ / " -f/ .......... ' H7- .f .I-'' ^. '' \ \ . - 3000 2800 2600 , 1+0D0 3800 1H A\ I l/tsj ________ fc 1 !i ' Died 3600 [Aj 15 i\\! V ............ . . y _ . 34oo / V 3200 p* - - 3000 2800 - Y ' ' A______ .- . Period of ------------ * Exposure " T9/9' TzCilXedr 1/ ______________________ > 2608 2400 p- * /1 ... . f . . - 2200 Time, Weeks Controls dubiected to Conditioned Air; f0r rxpcrimenfc No. 2 Aroclor 1242 and Experim.fnJi.;No. 1 Aroclor 1254 ' Figure 4 r NEV 016652 WATER PCB-SD0000030600 W e ig h t, Gram.1 i i t* Controls Subjected.tu.Conditioned Air; for ''xperiinent Ho. 2 Aroclor 121+2 end 1?xv>orinerxt No. 1 Aroclor 12^2+. figure 1+1 NEV 016653 WATER PCB-SD0000030601 o I WATER PCB-SD0000030602 I Sequence of Tables 1 The Mortality! Length of Survival, and Changes in Weight of Animals Exposed to the Vapor of Aroelor 1S4& in Air* 2. The Mortality, Length of Survival, and Changes in Weight of Animals Exposed to the Vapor of Aroelor 125^ in Air* 3* The Mortality, Length of Survival, and. Changes in Weight of Animals Exposed to Conditioned Air* ' 4* Sumary of the Data on the Mortality Among Experimental and Control Animals* . 3* The Average Changes in Weight of the Survivors Among the Original Groups* .. i NEV 016655 WATER PCB-SD0000030603 k ! I 1 1- 4 1 Sequence of Figures 1, Weight of Oat A-499 Recorded at Weekly Intervals During the Period of Exposure to the' Vapor of Aroclor 1242 in Air - 1.9 miorograms per liter (Experiment No. 3) . 2. Weights of Individual Guinea Pigs B-599, B-600. B-6oi and B-626 Reoorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroclor 1242 in Air - 1.9 micro grams per liter (Experiment No. 3) . 3* Weights of Individual Guinea Figs B-602, B-603 and B-6o4 Recorded at Weekly Intervals During the Period of Expo sure to the Vapor of Aroclor 124-2 in Air - 1.9 miorograms per liter (Experiment No. 3). - 4-* Average and Range of Weight Reoorded at Y7eekly Intervals of Guinea Pigs B-599 and B-601 to B-6o4 that Survived throughout the Period of Exposure to the Vapor of Aroclor 1242 in Air - 1.9 micrograms per liter (Experiment No. 3). 5* Weights of Individual Mice B-567. B-566, B-569 and B-600 Reoorded at Weekly Intervals During the Period of Expo- . sure to the Vapor of Aroolor 1242 in Air - 1.9 miorograms par liter (Experiment No. 3) 6. Weights of Individual Mice B-570, B-571, B-572 and B-921 Reoorded at Weekly Intervals During the Period-df Exposure to the Vapor of Aroclor 1242 in Air - 1.9 miorograms per liter (Experiment No. 3) 7. Weights of Individual Mioe B-573, B-574, B-575, B-576 and B-OOl Reoorded at Weekly Intervals During the Period of ' Exposure to the Vapor of Aroolor 1242 in Air - 1*9 mioro grams per liter (Experiment No. 3)* Average and Range of Weight Reoorded at Weekly Intervals of Mice B-567, B-566, B-571, B-572, B-573 and B-574 that Survived throughout the Period of Exposure to the Vapor of Aroclor 1242 in Air - 1.9 micrograms per liter (Ex periment No. 3)* NEV 016656 I WATER PCB-SD0000030604 I 0$ 4 G Hi 9. Weights of Individual Rabbits H-106l. H-lOfe, J-7d4, 1-622, 7-62$, ff-66l, J-663, J-664 and- 7-666 Reoorded at Weekly Intervals During the Period of g> osure to the Vapor of Aroolor 1242 in Air - 1.9 miorograms per liter (Experiment No. 3) - 10, Weights of Individual Rabbits H-IO63. S-1064. J-64. ' 7-324, 7-760, 7-623,- 7-662, 7-665 and 7-667 Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1242 in Air - 1.9 micrograms par liter (Experiment No. 3) 11. Weights of Individual Rats L-992, 1^993 and L-994 Re- oorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1242 in Air - 1.9 micrograms per liter ( Experiment No. 3) 12. Weights of Individual Rats L-995, 1-996 and L-997- Re oorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1242 in Air - 1.9 micrograms per liter (Experiment No. 3) ' 13. Weights of Individual Rats L-99$, L-999, L-1000, M-l and . M-445 Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1242 in Air - 1.9 mioro- . grams per liter (Experiment No. 3) 14. Average and Range of Weight Reoorded at Weekly Intervals of Rats L-992 to L-1000 that Survived throughout the Period of Exposure to the Vapor of Aroolor in Air - 1.9 oiorograms per liter (Experiment No. 3) 15. Weight of Oat A-500 Recorded at Weekly Intervals JXafing the Period of Exposure to the Vapor, of Aroelor 1254 in ` Air. - 1.5 micrograms per liter (Experiment No. 2) ' 16. Weights of Individual Guinea Pigs B-605, B-606, B-607 B-672 and B-799 Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1254 in Air 1.5 micrograms per liter (Experiment No. 2) . 17. Weights of Individual Guinea Pigs B-606, B-609 and B-6lO Reoorded at Weekly Intervals During the Period of Expo sure to the Vapor of Aroolor 1254 in Air - 1.5 micrograms per liter (Experiment No. 2). NEV 016657 WATER PCB-SD0000030605 i iv 4 Jr t id. Average and Range of Weight Recorded at Weakly Intervals of Guinea Riga B-bOb and B-bOd to B~6lQ that Survived throughout the Period of Exposure to the Vapor of Aroclor 1254 la Air - l5 miorograas per liter (Experiment No. 2). 1$. Weights of Individual Mice B-577, B-57d and B-579 Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolcr 1254- in Air - 1*5 miorograms per liter ( Experiment No. 2). - 20. Weights of Individual Nice B-5d0. B-5dl, B-5d2 and B-919 Recorded at Weekly Intervals During the Period of Expo sure to the Vapor of Aroclor 1254 in Air - 1.5 miorograas per liter (Experimant No. 2). 21. Weights of individual Mice B-5*3, B-5d4, B-5d5, B-5dd, B-602, B-603,. B-673 and'B-do3 Recorded at Weekly Inter vals During the Period of Exposure to the Vapor of Aroolor 1254 in Air - 1.3 miorograas per liter (Experiment No. 2). 22. Average and Range of Weight Recorded at Weakly Intervals of Mice B-577 to B-580, B-5d2 and B-5S5 that Survived throughout the Period of Exposure to the Vapor of Aroolor 1254- in Air - 1.3 micrograms per liter (Experiment No. 2). 23*. Weights of Individual Rabbits H-IO65 and E-1066 Rsoorded * at Weekly Intervals During the Period of Exposure to the Vapor of Aroclor 1234- in Air - 1*3 miorograms per liter (Experiment No. 2) 24. Weights of Individual RabbitsH-10fi7t and H-lOdd Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1234- in Air - 1.5 micrograms per liter (Experiment No. 2) * . 25. Average and Range of Weight Recorded at Weekly Intervals of Rabbits H-IO65 to H-lOod that Survived throughout the Period of Exposure to the Vapor of Aroclor 1254 in Air *1,5 miorograms per liter (Experiment No. 2). 2d. Weights of Individual Rats M-2, M-3 wd l!-4 Recorded at Weekly Intervals During the Period of Exposure to the Vapor of Aroolor 1254 in Air - 1.5 miorograms per liter (Experiment No. 2). NEV 0166S8 WATER PCB-SD0000030606 i 27. Weights of Individual Hats M-5 H-6, H-7 and M-d Re- oorded at Weekly Intervals During tha'Period of Exposure to the Vapor of Aroclor 125^ in Air - 1.5 miorograms per liter (Experiment No. 2) - 23. Weights of Individual Hats H-9, 15-10, H-ll and U-l^tO. Re corded at Weekly Intervals During the Period, of Exposure to the Vapor of Aroclor 125^ in Air - 1.5 micrograms per liter (Experiment Ho. 2) . 2? Average and' Range of Weight Recorded at Yfeekly Intervals of Hats 15-2 to 15-11 that Survived throughout the Period . of Exposure to the Vapor of Aroolor 1254 in Air - 1*5 aicrograms per liter (Experiment No. 2). 30. Weight of Control Cat A-510 Recorded at Weekly Intervals During the Period of Confinement in Conditioned AJr (Ex periment No. 2). ' 4 31 Weights of Individual Control Guinea Pigs B-6ll, B-&L2, B-613 and B-79$ Keocrded at Weekly Intervals During the Period of Confinement in Conditioned Air (Experiment No. 2). 32. Weights of Individual Control Guinea Pigs B-6ltf>, B-615 . and 3-6l6 Heoorded at Weekly Intervals During the Period ' of Confinement in Conditioned Air (Experiment No. 2). 33. Average and Hange of Weight Recorded at Weekly Intervals of Control Guinea Pigs B-612 to B~6l6 that Survived throughout the Period of Confinement in Conditioned Air (Experiment No. 2) 34. Weights of Individual Control Mies B-5^7, B-5S6, B-5$9, ' B-9S6 and B**99 Recorded at Weekly Intervals During the Period of Confinement in Conditioned Air (Experiment No. 2). 35. Weights of Individual Control Mice B-590, B-591, B-598, B-doif, B-936 and B-967 Heoorded at Weekly Intervals Dur ing the Period of Confinement in Conditi. oned Air (Experi ment No. 2) - 36. Weights of Individual Control Mice B-593 B-59^, B-595, B-590, B-933i B-935 and B-970 Recorded at Weekly Intervals During the Period of Confinement in Conditioned Air (Ex periment No. 2). NEV 016659 ! WATER PCB-SD0000030607 Ti Jf, Average and Range of Weight Recorded at Weekly Intervals of Control Mice B-5&6, B-569, B-591 and B-59I4- to B-596 that Survived throughout the Period of Confinement of Conditioned Air ( Experiment No. 2),. 33, Weights of Individual Control Rabbits H-IO69 and H-1070 Recorded at Weekly Intervals During the Period of Con finement in Conditioned Air (Experiment No. 2). 39, Weights of Individual Control Rabbits H-1071, a-1072, T-183 and 1-202 Recorded at Weekly Intervals During the Period of Confinement in Conditioned Air (Experi ment No 2), 1)0, Average and Range of Weight Reoorded at Weekly Intervals of Control Rabbits H-10b9, H-1070, J-l$3 and J-202 During the Period of Confinement in Conditioned Air (Experiment No. 2). * hi. Weights of Individual Control Rats M-12, M-13 and H-l4 Recorded at Weekly Intervals During the Period of Con finement in Conditioned Air ( Experiment No. 2) 42. Weights of Individual Control Rats M-15, M-l6 and 14-17 . Reoorded at Weekly Intervals During the Period of Con- ` . finement in Conditioned Air (Experiment No. 2). 43. Weights of Individual Control Rats M-l$, H-19, M-20, M-21 and M-35I Recorded at Weekly Intervals During the Period of Confinement in Conditioned Air (Experiment No. 2). 44. Average and Range of Weight Reoorded at Weekly Intervals of Control Rats H-12 to H-lS, M-20, M-21 and M-531 that Survived throughout the Period of Confinement in Goa ' ditioned Air (Experiment No. 2), ' 4 `& NEV 016660 WATER PCB-SD0000030608  The Toxicity of the Yanar of Aroolor i242 ana of Aroolor US* (Supplement to Report of June 22, 1955) Soone This supplemental report presents the results-obtained when separate groups of animals were exposed, respectively, to Aroolor 1242 and Aroolor 1254- in lower concentrations in the atmosphere end for a longer period than were those employed in the experiments described in the report of June 22,. 1955* Since these experiments involved a different period of time and a dif ferent season of the year, a new group of control animals^ was included* - ' SmnffleT'V Of Results 1* In an experiment in which, for 7 hours per day on 150 days over a period of 214- days, animals were exposed to air bearing Aroolor 1242 in the concentration of 1*9 micro grams per liter {0ld ppm), the incidence of mortality among the various species, with the exception of the rabbits (which died during an epldemio of pulmonary disease), was no greater than that encountered mnng a similarly constituted control group* The growth of the experimental animals was unaffeoted* NEV 016661 WATER PCB-SD0000030609 I / i lv* -2- No gross or microsoopio evidence of general or specifio in jury was found in the tissues of the animals as a response to their prolonged exposure to Afoolor 1242 2* In another experiment, in which animals, were exposed to air hearing Aroclor 1254 in the ooneentration of 1*5 micro* grams per liter' (0.11 ppm), for 7 tours per day on 150 days over a period of 213 days, the incidence of mortality was no greater than that ohserved among corresponding groups of con trols, The growth of the experimental groups did not differ significantly from that of the oontrols. No relevant gross pathological ohanges were observed in the animals, hut micro- . soopio lesions of a mild, nonspecific, toxic type were found In the livers of some of the guinea pigs and mice and in n of the rabbits and rats* Similar ohanges were also found in the renal tubules of the rats* Although miorosoopio abnormali ties of this type are often found in the tissues of "normal" animals, those observed in oertain of the oontrol animals were not as severe as they were found to be in the corresponding speoias of animals that had been exposed to Axoolor 1254* The conclusion seems to be warranted, therefore, that prolonged respiratoxy exposure to Aroclor 1254 is oapable of causing some injury to the tissues of susceptible animals, under conditions in which the atmospheric concentration of the material is of the order of 0U parts per million* NEV 016662 1 WATER PCB-SD0000030610 3 Exposure to the Vapors The method of volatilisation of either of the Aroelors from & heated glass well was similar to that described in the earlier report (June 22, 1955) except that in these experi ments the temperatures of the Aroelors in their respective wells were decreased and the rates of air-flow over the surface of the . liquids were increased. In the oase of Aroolor 1242, which was heated to 55s to 6o0, the air passed over the liquid at the rate of 00 liters per minute as measured by a venturi-meter attached to an inclined manometer. Aroclor 1254 was maintained between 115* and 125*0 while 700 liters of air per minute passed over its surface before entering the chamber, . During a 7-bour period on each of 5 days per week, 3 groups of animals (2 experimental and 1 control) were confined simul taneously in separate 600 liter ohambers as desoribed earlier. Zaoh group consisted originally of 1 oat, 6 guinea pigs, 10 aloe, 4- rabbits and 10 rats* When an animal in any of these * groups died, it was replaced soon thereafter. ' The oontent of Aroolor in the air was determined daily by the turbidime trio method described in the previous report. In these experiments samples were collected by passing air at the rats of 1 liter per minute for 45 minutes through a Willson NEV 016663 WATER PCB-SD0000030611  4- combustion furnace and then through 2 midget bubblers in 'series, each containing 10 ini of 0.1 N sodium hydroxide Experimental Results Aroolor 124g Experiment No. The pertinent data relat ing to the fate of tha individual animals of this exp rimant are . detailed in Table 1 and summarized in Table 4* One oat, 5 guinea pigs, 6 mice and 9 rats survived, following their subnotion for 7 hours on each of 150 days (over 214- days) to air bearing the vapor of Aroolor 124-2 in the concentration of 1*9 micrograms per liter (O.ld ppm) e Seven additional animals, introduced es sub stitutes for animals that had died during the experimental per iod, were alive at the conclusion of tha final period of expo sure; these ere listed to show their speeies end, in parenthe ses, the number of periods of exposure to which eaoh was sub jected is given, as follows * 1 guinea pig (26), 2 mioe (4$, 135) 9 3 rabbits (d, 13 and 34-), and 1 rat (lOf) ,, In this initial group, 10 fatalities occurred as follows: 1 guinea pig after 124- periods of exposure; 4- aloe after 14-, 14-, 102 and 150 periods, respectively; 4- rabbits after 17 lid, ll and 129 periods, respectively; and 1 rat after 4-3 periods. In addition, a substituted mouse died after 133 periods of i NEV 016664 I WATER PCB-SD0000030612 5 ezpo8uret and 11 substituted rabbits died, for the most part, during the first 2 weeks after they had been' lntroduoed. fen of the 15 rabbits died within a period of J weeks during a severe epidemic of pneumonias . Exoept in the case of the rabbits, the incidence of mor tality was no greater among the experimental animals than among the oontrols (Tables 3 and 4) Among the original group of controls 1 cat, 3 guinea pigs,. 4 mioe, Z rabbits and 9 rats survived throughout the entire period of their confinement, for 7 hours on each of 150 days over 213 days in a chamber supplied with conditioned air Three other animals, 1 rabbit and Z mice, were killed after 3^ 109 and 113 periods of con finement, respectively, in a chamber in whioh air containing * no Aroolor was conditioned with respeot to dust, humidity and temperature Other oontzol animals, introduced as sub stitutes for those that had died or been killed, survived during the indicated numbers of periods of confinement, re spectively* 1 guinea pig, 55* 6 sice, 5, 5, 13, 37, 4l and 66; 2 rabbits, 115 and 119* and 1 rat, 115 Seven fatalities occurred among the. original group as follows: 1 guinea pig after 94 periods of confinement; 4 mioe after 64, 112,' 137 and 141 periods, respectively; 1 rabbit after 30 periods, and 1 rat after 29 Tro mice used as substitutes died after 4 and 33 periods of confinement, respectively. NEV 016665 WATER PCB-SD0000030613 I -6- . In summary, in each of tbs original (experimental and oon- trol) groups, 1 guinea pig, 4 mice and 1 rat* died, No specific signs of intoxication were observed in the members of either the ec^erimsntal or control groups* .. The weights, observed at weekly intervals, of the individ ual animals (cats, guinea pigs, mice, rabbits and rata) exposed to Aroclor 124-2 (Experiment No* 3) are shown in Elgares 1 to 3, 5 to 7 and 9 to 14-* The average weight of the original groups and the greatest and least weights attained by individuals within 1 these groups of guinea pigs, mioe and rats that survived are plotted in Figures 4-, 3 and 14, respectively* Comparable 'data for the controls are to be found in Figures 30 to 44* The av erage changes in weight of the various survivors of the original groups (experimental and control) are shown in Table 3* The ap plication of the widely used nt" test indicates that the growth of the animals exposed to Aroclor 1242 did not differ signifi cantly from that of the control group* , All of the animals were examined post-mortem, and the vis- oera of most of them were examined miorosoopioally* Among the exposed group, 1 guinea pig, 5 nice and 15 rabbits that died, and 1 rat that was killed when moribund, were viotims of in cidental lnfeotiotts disease* The cat, guinea pigs, mioe and rats that survived had normal viscera* The 3 rabbits used as NEV 016666 WATER PCB-SD0000030614 I -7- . . replacements survived. Two of them were found to have hepatio lesions of ooccidiosis and the other bad no gross microsoopio abnormalities* - The control animals that died were found to have pneumonia* Of those that survived, 1 rat, 2 guinea pigs and 1 mouse had focal or diffuse oytoplasmic vacuolation of the hepatio cells, tbs etiology of which was not apparent* The remaining animals ' that survived had essentially normal viscera. Aroclor 1254. Experiment No. 2. The pertinent-data relat ing to the changes in weight and fate of the individual animals subjected to prolonged intermittent exposure to the vapor of Aroclor 1254 are shown in Table 2 and summarized In Table 4* One oat, *4- guinea pigs, 6 mioe, 4 rabbits and 9 rats from the . original group survived following their exposure for 7 hours on each of 150 days over a period of 213 days to air containing Aroclor 1254 in the concentration of 1*3 micrograms per liter {0*11 ppm)* One guinea pig and 1 rat from the original group were killed after 30 and 14$ periods of exposure, respectively* " Seven animals introduoed into the group later survived* They are listed by speoies and periods of exposure, in parentheses, as follows: 2 guinea pigs (4? and 120)j 4 mioe (49, 66, 115 and 135); 1 rat (139). Among the original group only 4 mica and 1 guinea pig died after 12 to 101 periods of exposure, and NEV 016667 WATER PCB-SD0000030615 a mouse, whioh was a replacement, died after id periods of ex posure, the incidenoe of fatalities among them being slightly less than that encountered among the controls. No general or specific signs of intoxication were noted among the experimental animals during or after their exposure* The weights, at weekly intervals, of the individual animals (i*e*, oat, guinea pigs, mioe, rabbits and rats) exposed to Aroclor 125*1- are presented In Figures 15 to 17, 19 to 21, 23 and 24-, and Z6 to 2d* The average weight of the original groups, and the greatest and least weights attained by Individuals within these groups of guinea pigs, mice, rabbits and rats that survived are presented graphically In Figures id, 22, 25 and 29, respec tively* The rates of growth of the experimental groups did not differ significantly from those of the control group (Table 5) All of the animals were examined post-mortem, and the vis cera of most of them were examined microscopically* The guinea pig that died exhibited chronio pyelonephritis, pulmonary hyper emia and edema, and degenerative lesions in the brain and liver* ' The visceral lesions were related, no doubt, to the renal infeotion* The deaths of the 4- mice were attributable to aoute bronchitis and pneumonia* The visoera of the oat that .survived were normal. Of 7 guinea pigs that were living whan the exper iment was terminated, 3 had normal visoera and four had slight NEV 01666a WATER PCB-SD0000030616 alterations or hepatio cells characterized by cytoplasm!d vaeuolation. Ten aloe survived and of these 6 had noxmal viscera and 4 had slight degenerative changes in the liver. Four rab bits killed 1 to 15 days after the last period of exposure had diffuse hepatio degeneration. The character of the lesions var ied from cloudy, hyalins or hydropic degeneration and.Included varying degrees of fatty metamorphosis. The other visoera of these animals ware normal. All of the rats were examined and found to have degenerative lesions of the liver of-slight to moderate degree. The lesions of greatest severity were'found in tha rat that was killed and examined on the first day .after the last period of exposure. Two rats had chronic pyelonephri tis, and the remainder had slight degeneration of the renal _ tubules. Discussion The preceding report (dated June 22, 1955) indicated that no harm resulted to animals that inhaled Aroolor 1242, inter mittently, over periods not exceeding 4 months when tha con centration in the' atmosphere approached that of saturation. The present report demonstrates, in terms of rates of growth, incidence of mortality and non-oocurrencs of pathologio changes, NEV 016669 WATER_PCB-SD0000030617 I 'i* i i 10 - that no injurious effects resulted from more prolonged, inter mittent exposure (more then 6 months) to the- concentration of O.ld ppm in the atmosphere (1,9 micrograms per liter). This concentration is nearly twice that (1 microgram per liter) . . recommended by the American Conference of Governmental hygien ists (Arch. lad. Hyg. Oooup. Med., 2:530, 1954) for a .chlorin ated diphenyl of unstated chlorine content. The exposure of animals to Aroclor 1254 failed to induoe harmful effects in the form of retardation of growth, or of mortality, but histopathologic evidence of apparently reversible ' hepatic cellular injury was found in the animals. These find ings cannot oertainly be attributed to the effeots of ArOolor 1254 because of the appreciable incidence of pneumonia among both experimental and control animals. When these nonspecific toxio changes in the visoera of the animals were associated with pneumonia, they were readily explained thereby, but they were also found in animals that had been exposed to the vapor of Aroclor 1254 and were free of pneumonia. That these may have . represented toxio effeots of exposure to Aroclor 1254 finds support in the fact that the livers of the exposed rats (report of Tune 22, 1555) were significantly heavier in relation to their body weight, than were those of control rats. Zt would appear that this material, which is reported to contain 54 NEV 016670 I WATER PCB-SD0000030618 per cent of chlorine, is somewhat more toxic than is Aroelor 12kg. which contains only kg per cent of chlorine * OeB* Drinker proposed (7* Xnd. Hyg* Toxicol*, 21:155. 1939) as a threshold concentration for a sample of a chlorinated diphenyl that con tained 50 to 55 pa? cent of chlorine, the value of 0*5 micro gram per liter, which is about one-third of the concentration of Aroelor 1254- to which the animals were exposed in this ex periment Proa The Bettering Laboratory in the Department of'Pre ventive Medicine and Industrial Health, College of Medioine, University of Oinoinnati, Cincinnati, Ohio* Project Mo* 47* Experimental Work and Report by: Joseph ? Treon, Ph,D. ' Prank P* Cleveland, M.D* Frederic B. Shaffer, M.S* John Oappel, B*S* Robert A* Boiler, M,S* John P* Torbeok, B*S* ' June 26, 1955 Director NEV 016671 WATER PCB-SD0000030619 1 m d I 4* n+ 9 o m fe'gSw op. Vi 4 *h . O Vi P 4 rH W H tC 4 O lM Iq a aaaaa aaa a & & * *d > < > d 49 a <d <d H uo-flo trstrvao H-80 H H d ' ad <Od (>0O Q<0<Q<0 HH'd, r*"t< Hf-4 Hi--I H<--I Hi--I ti H fH *H *r4 *H **4 44 ww*5 g g H H g:m-wh jri hw 'h ^ d *d > >d >d d <o > m i am *0l H TiS* _ - _ 3$ 3 CO O CQ CO M CO 60 `>`<d'd`l2''a'3 3h3ii4oHiH3MS9hSb4cti| cocopQcoracocoQa s$.2 t *g^ sd o o si SS?* S8 *H 43 H S60 a d<rl 0 0 43 91 3W JhlO.o ffii hhOOOO VO 0? 1 6 H o Ps I + 1 O o^ON-rt ! ON-KVCUX** 1 + ++ *60 CVJXO 0_ t>--<-( r-t WxKOVOVO fArt OJ KVf<~Sl KMfN ++++ ++ + M . . r*o Ht- 'J5* ovao^so 1 CVJ t lift 1irst-j-H . 043-10 \T\0 0 x*vq H ITNOJ h>CM KVCW H H CWtC\HH<-4tCN + + '+ + + 1 1 r-t fl 3 hi3 *4 4oH* 5 " S' CW KNO OW mr-sp-eo r--< _ CeUgxwi*ehu-!cTu\scjv* iHw1ew.=w* KcuMcvfj\ iTMCNiCMCN *o1 tX! Ml ti . P4 s" 39 ONO H CVJ t<Vi"03 g ONOOOOO w ICNNOVOVONOVO'BO II IIII I 43 Annnnnn M o ON QH OdHttCH O*HQOttOHtt vx> 2 P<P<P4AIOP<P1 nnnnnniini&n i! oO O O woo asaaaas iilliiiili H o 3933939 0000000 0000000000 maaaaaaaxa 4 4 ' o 00*00000 OOOOOOOOOO 5 f** 1*-- NNNNNNNb^NS H HHHHKHH HHHKHHHKHK ^ssssass*& i_oc_n_da_i_'So_S_oi_n_uo_N_toc_N_aca pod'Noooood- H HHHHHH fl o +rl H c? au O Is o ON M o rt NEW 016672 WATER PCB-SD0000030620 a <L* O' 0 , o | VlB4 Juj Pi0o soaa o^htn < Pt *i-t 0 4* C O C > P.J; SMfiS q 0& <0 0 in H ?a*w0 a ass >0 > Ua 00 Pai oa e oa 00 & 0 >. 3 0 0 43 03 fMn >>4in "i SH Pi 0 Q 0 0 0 SSSNSN0 s5 MI0 60>40 >.0 0 S 000000 0000000H00O43C-HlA^BI 00 HO 00USrO4 0O rlrirl I^H r-Jj^ H CM `iN5H2 00 O > 000000000p>i0000000000p>i0000p>j0 BQQqQOwoOP-Poorafiolfl 0 I* S' 00 H0 00 OO H <H H *4 r"Ht MM 00 s 0 UT\ .. N- vO-si-ON H >- bob -4 0 0 P $ o *o4 no M> <0 0 til_0 <ri o a BOM 5h M o 3 BO O 43 So H^j.. s 0q 43 P4W o la ^sd +i in h-tncvi H4evj CM CM + I+ I I I I I I I I I * I in CM trvd- jd-MI + I + t TON'PO-OinWOsK'lOONCMONSOCNjnCMN- wosH'toH'oaor-KisOHr'-oioinsi^j'Bo rlHH KlH HH ` H H ++ I ++++ I t I I I++ 1 I + I II "BOO WIO + aV I 3* (OIOS TOTO Q CM lf\H roo ONO N-tnfOlAH O g W QP- cm cm cm in incM-BO rotoNO eM^so-eo rjjd-eu ir\inoso> if os Sr-1 OSOHotOJd-eomN-ovBOd-frui-BOvOMMfv CM H fncM CM KNCM CM CM CM K\K\CM K\K\<VnCM CM K\ 3u o0 d 0 H 0SD ON nnn e ill aa sa HCMSO^f SOSO OVO Q4* CM KVBO H CM KVd-ITtNO N O O H 03d- CMnSv CM CM CMVONOVOVOTbTOTO rlH I H- mN-h-BO'WBOW'SO'W'BO'BO'W'BO Ad2*lw4.i.wlulululw 4*4*4*4*4*434343434*434*434*4*4*4*4* %P1 SS14*<5B<B5<S50<*S<0 MA iO0"2aCO*83i'OS' O"SPOSlQS<I0SI'0 CMK\ ON OS s$ II OOO t nnn MHH g> rlH KKXXXXXXXXXXXXXXXX ONTO NNO H Usd- 01J KV'*mt'~KSCM CMTOVO WrjrlrJ (^NIAH H OO NN XH 0in0it. 0H i H 4* 0 Pi Is 0 O 0o: o TO H O ON NV 016673 WATER PCB-SD0000030621 1 day days nd. 15 iol lr iebdu Survived K ille d 15 days S urvived K ille d 15 days SurvL ved K ille d 13 days Survived K ille d 6 days Survived K ille d li days f1 days $ days Lv ievnaglthAoftfe rSur iXast Period o f Exposure Unless Otherwise S tated KK iillllee dd KM K ille d . S u rv iv e d * S u rv iv e d S u rv iv e d Fate Table 1 (Page 3) D u rin g P e rio d of Exposure isQh <maJ u0x+<6r:Ci aifc _ +S.h3 ro^M)Vol *pQo <21 M d <H 60 0)04^ Co WK oM-HC J3 ftH O a oasirvo inVaO- I r-- ^KVrlOd'OO H ++ t 1+ I 1 + vcoutorv\oodw-vcouo\Hiergvccegs\ +++++++ i Duration o f Exposure (hours) H terr-gJeKegg\jHeognhOoSCmevgJ'oBHOveHeg ceOgg^hhi* 0 a o u SI o on ONONS'O'^OsO 7 'O H 43 43 43 43 43 43 43 43 43 PS PS os ot os os os 000000000 few t ^hhhhhhmh HHrlHHHH H H g o vo 0 s ra4ooeco* H c\ H . 0166?< WATER PCB-SD0000030622 iAg ihrt We in Changes I n roolor 1254 and of A f S urvl-ral. th e Vapor I 0 95 6 4o* q4a .rO*.a.I4MoQ0'OhoDein"O & .m 0d>*0d^ mam a d ^d0^d*dd^0d*0 moo 6o?foc*d?2d? dd d d o o >0 Hdo a"BO IHTNHirNr-TBO H OnONON ON 0saa sill's <o0 2 <0 *0 <3 *0 <poqoq4>g)<d H HHiflHrlHHH "*5 '0 0o 0 So a0dd0 WW WWWW SS5Swm00S 8888 843 H 00 g 00000 0000 0 VV. V4 ' >0 >00 > I IsSsSSas SS^5S55| WCQQtaCQCQtOW WCQCOCOQODQQ W 4 *S B 4-tS, aHdoe0d SSShi So<H WA049 JfS I fc M O O ss on cvj .i ri cm .i rr--Nr-DoNso _i oon +++ CM CNOCTnHfOUNdO CM in r-Hji- mno inxNdo H CM K>0 CM r4 CM .+ + + + + + + + 2TO O I __ I I I I + ONOOTOoKNOdO O ?iS8"^-srs 1 + 44 I 4 4 I 4 (Experiment No. 2) -M00403-SX50toJjf--e--,c*c to dptnrj-pntriNOho UN f~*r-l3"T0 UN do jd* mini?.:*.=* inm M UNNO f'-TO On6 ivi ON srsissi&Ra&sj g 0 4do3 eh O O O QO r-th-ON NONONO NONO NONO III II I II nnnnnnnn 0O0dt0C0O0O0O0O0O I Kiln'S "So do 4o'b? mnm wftnnnnn 4 oooooooo 800808080 0H o13 o3o3o3o3e33o3oao 339S33S33 oo o oo oo o o aasaaaasa 4O4 codSd-H' o o ooooooo O"O# O* OO%OOOO si*e H HHHHHHHH hhhhhhhhh .5 a HNjno rtrlrtrtHrl SSSSSSMS al0o 43 o eP 8Ol m 8o0o ; H NEV 016675 . Length o Exposed to to rta llty A n im a ls WATER PCB-SD0000030623 k o no I Vi tip U 090 a . _h+s 03 Vi <9 Ota 9 0S VlPH 9 op u o a &,,<& 9ftpi PH 9 G COOP O 9 G f 9 Pr.C 373SS n * 9 999 9 O 9 *9 m H BBS? S*9 K>9 <9 <9 03<o 9 9 9 9 O, rHH'O_ rHH rHH HiH rijH H H H wH kS W can 9 9 0 9 ftKK 9 >9 *9 >9 X* *w *0 *0 "d HWO IT\ HV9VV 9 *9 >9 >9 9999 H HH rl HPHH *2* 9 9 9 9 >9 99990 rH iH rH rH (H rlrlrlrlH H2**H *H*HH 9 *9 >9 9 9 <9 <9 >9 9 >9 >9 19 *9 9 P P* 9 009 r t _jH *H t 0 > > > P9HPU 2Q? c3acSt wfl w9 S9 9000 * * * p > >-5J^ inwraoj QA0A0 hhh 333, W CO W 03 I ss +> . *9 on vus, oo. a h coed O-rtViB 9 ^ *rt P o o o >9 9 9 P 9 ~ *?* <P4 m r-in r-in ^SS-^S + 1 1 * 1. H in . mr-ovo tCNHtO w Hines +++ 1+ 1 h n p h 9g9 9 M O *H CM PlOP Sf-H* f-- CM-BO H tnoH N-N-fneuvo ISJe or w ph Vi fc O ~ VOVO CM CM in omrnH cm cm r--mo mCM CM l*-H ^inO-4- cMKvnmH BAo CM r-t 9P . o H G^^l rH p ud to CM KSCMTO as pfcroto r>-0"oo woo t HHi? CM CM CM CM H H #lno m j-hwoo 0 9-- t^9*ovo CM CM CM CM CM H5 cMjnmeM so cm mmmes iruO3O\D\Pot9aHas iiiiii mnneamm 009099 3 000000 xsssbss tH S'* HBV Id 000000 HHHHHH I jnmto ir>o 3. BO KJH HVO-d* HH tr>o n-to V0S6VOVO 0000 T't't'S KWWB pppp HHHH 3333 3jS>jo 0009 eMtnsi'irvo 111 1 1 aasaa P P p p 43 900 9 9 Ketafosas 0 0*0*0# 00000 r-r'-r--h-h- HHHH HHhHK pppp pl'p'k HHHH HHHHH gH H P H 9O Vt c. t* 9 9 in 3g C3 NEV 016676 I WATER PCB-SD0000030624 9 & Uo oM u Pi A 43 u a Pi 3 3 A 4 *g *d 1 15 h * 3 NEV 016677 WATER PCB-SD0000030625 WATER PCB-SD0000030626 ' day days days 1 day d days days days days days 9 days 9 days 9 days 9 days llldeeeadddys151d 15 15 15 15 dentally,, 1 hour KK iillllee dd KK iillllee dd 0 . 5 day KK iillllee dd 1 day d d K ille d acci LLv ieavnstaglthPAeforitfoe rdSuprf Exposure Unless Otherwise Stated iilllles KK iillllee dd l l l K2i Ki Ki KK 1 1 Fate Survived Survived Survived Survived Survived Died Survived Survived Died Survived Survived Survived Survived Died Survived Survived Survived Change in Weight Expressed as IPneirtcieantlagWeeiogfht . During P e rio d of Exp osure ` Table 3 (Page 2) d s U O 28 sa 4 or- ^ d- icn i + i+ vo P<i\ + h-vo cvjoj r-ff, + i ++ VHO oWoOC'oMVoJf_mvtsH?ot oHNtCwd\ - t Hovtaciv* oKH^chhu-virrol\veo\jvVico0v^8cJvBj- + + * + + + + + + 2 3 C * dwr icrvsjc^v-icovjHcuccuvjohvcdvj*ccuvcj vcjjcovjj vfMC'oV-JOnQovJ-HrcKrvV\jiOoOoJV'SOHOJoCkHVvnJ 8 ONO H CVJ tL jd- itnoJ-roumh-voo\o 'ooootoo o 0 X <UiU'UiUiU 3*1 O 0080900000 ^9 49 49 49 49 49 8. 3 3333333333 SSSSSS H a aaaaasasaa Duration o f Exposure (hours) 0 O O O O O O O O O O '000000 f"" ^f**f*"f*-?*"r-- H HHHHHHHHHH khhhhh cn p ovo HK*vrKV=}-tr\irv eood-oMn H HH ' rtH H H NEV 016679 WATER PCB-SD0000030627 days days days 5 5 5 1 1 1 K ille d 15 days 1 day ft d a y s $ days 6 days 6 days 6 days O therw ise S ta te d E xposure U n le ss of r d te io f r s lA Pe le n g th o f S ur lldeeaddy le d KKKKK iiiiilllllllllleeeee ddddd K ille d ta lll vs KK 5ii Ki lvai ed ge We ss tla Change I n W e ig h t xpre ri tc iean E IPne ^ ^^ ^ ggggggg ggg +> wcarararararaQtnwra t ao sf ig h o uS S uoo Up a. | .=t osinoNcsiTfeo ! k>nd HWrIWOOO HrH H ++++++ + + J+GVlHOKNK'vOvOi J+t+D W jrf- p O O-SKO K'NO'BOVO H KVM in ++++++ 1+4 + D u rin g P e rio d r0"l*O0 MS<OWOiK->IeHvJ CrUtH K'MJ MNCgojCOCVICUCUCUOtH ______ ( k g ) ___________ E x p o s u r e W eight o f In itia l .Id e n tific a tio n Number D u ra tio n o f E x p o s u re (hours) M IM II Ml? saaaasaaaai 4S494344*-0-P4>+4*0 ($&&((*&&&&& OOOOOOOOOOO f--t--r--f~r--"f--f~--f--" * HHWHHHHHHHH rlrlrlHrlHrt HrlH NEV 016680 WATER PCB-SD0000030628 f. \ ; eTH(DaS. *'nVooH P . 43 O 1 ?0* 0* m i Ha<<psd0 s 11 >e>0----a h3 Iu 2* AgjS! i-- Heg h * >Q5d -oHH 5o s < g<3 Vi m i 0 A o a|| 7Mo3 >H--aoi 3V * Kel V< |15 |a ca Voi BH tl I5 i were controls days; on 150 . 1 i tf0oo#- i 1 w co 3rro-*! O' 1 IITT\ Pk\Crj\ S HiHn JHH\ H e eg H day per nsuubjneccotnetda tmionateexdpoasturmeosfpohrere7#hours 4a 0 I 3Hhe- I I 3 1 5O' cBHrat :9*0 oroicOgS. I H 0- H rt 3 rj 0^ o 55 IH o rJ O3' s eg M fO 5 H in eg H 3\ nctoarlr easn ip omna ldsinwg le rye i E x p e rim e confined 2a Wr00l aHo +a oa<ri K +a* B Oat NEV 016681 WATER PCB-SD0000030629 k Table 5 The-Average Changes la Weight of the Survivors Among the Original Grouts; Species of Animal Average Change in Number Average Initial Weight Expressed of Weight : as Percentage of Animals ______ Os)_____ __ Initial Weight Experiment Nq9 3 - Aroclor 1242 Oat 1 1.730 + 115.0 Guinea Pigs 5 ' 0.514 + 43.1 nice 6 0,023$ + 21.5 Rats 9 0.214 ......... 39.9 Experiment No. 2 - Aroolor 1254 'P >0.05 >0.05 >0.05 Cat 1 Guinea Pigs 4 Hice 6 Rabbits 4 Rats 10 2.656' + 15.2 0.467 * 32.5 0.0243 17.0 2.95s 44.4 0.222 + 26.7 Controls - Experiment No. 2 >0.05 >0.05 >0.05 30.05 Oat 1 Guinea Pigs 5 Mice *6 Rabbits 4 (l) Rats 9' 3.329 0.500 0.0257 2.622 0.216 Cl) Includes 2 early replacements. - 7.6 * 54.7 + 13.2 52.2 + 33.7 . -. - m .- NEV 0166E2 I WATER PCB-SD0000030630 NEV 016683 WATER PCB-SD0000030631 Experiment No. 3 Time, Yeeks 1,9 Micrograms o f A ro o lo r 1 2 4 2 /lite r ` Figure a X O 3B 0 3 IHS b 4o* bobco `^uSie/A NEW 016634 WATER PCB-SD0000030632 4 * & f6t0 NEV 016685 WATER_PCB-SD0000030633 016666 I WATER_PCB-SD0000030634 SJ J4 9 V\ * B NEW 016687 WATER_PCB-SD0000030635 ]5 e o * 3 I- 121*2/11ta r A ro o lo r o3 f E xp a rln e n b No. 1 *9 tflo ro g ra m a Wft rOt Wn nOnWO^Ocvi \fAtnO NNO nV\ fOtUp\ Oi Btasag 'V^Te/i NV 016688 WATER_PCB-SD0000030636 n iot M 1 'B u - E xp e rim e n t N o. 3 . * 1 .9 Micrograms o f A ro c lo r 1 2 l* 2 /lite r Vm\ On \A n W O IA o o wov\ mnpINrlNNH u\o NN u\o Hn V\ M O N V\ H nnc{) '^vdS-fit NEV 016689 WATER PCB-SD0000030637 f & f ! i X 9 O as 6 to cv H fa Ho Oo fa n o4 1 +c> ftoao SS -o*H b S 0.9^ jS.4 NEV 016690 WATER PCB-SD0000030638 NEV 016691 WATER PCB-SD0000030639 F ig u re 10 Tim e, Weelce 1 .9 Uicrograma o f A ro o lo r 1 2 4 2 /lite r Experim ent Ho. 3 oo o OOO O V\ H R8R8R8&8 <n cm n <n n ccum -a- lSAOoVpNoo R5SJ0 ^qSTM Vo\ oo Wo o 5 O V3AOo XoA' Oo 5o\ NEW 016692 WATER_PCB-SD0000030640 I smj^o '^nS-ps/^ X Q 3' ua 22 Hft, 3 CM h O oo h no ii a 43 H) CO geH <hoH ha AO_\ H C9H NEV 016693 WATER PCB-SD0000030641 n X or. -r * DO 43 5 (M C. O o <8 mo 55 3 fca acK 5 snmao '^9*^ NV 016694 WATER PCB-SD0000030642 I ! V 016695 WATER PCB-SD0000030643 Experiment No. 3 1.9'Hlorograma o f *ro o lo r 1 2 l|2 /lite n X 9 * is9 0 h & Oi u o H O 2 no is *2 43 !s3 2 Se *f2es i D. On. K NEV 016696 WATER PCB-SD0000030644 f' !*. \ E 016697 WATER_PCB-SD0000030645 & o c* 1 e o pH * `o P I 60 1 2 5 4 /llto r A ro o lo r o2 f Experiment No. 1 .5 'M lc ro g ra m a NEW 016698 WATER PCB-SD0000030646 StBB^O `3t*S?8/ NEV 016699 WATER PCB-SD0000030647 i ii a a CO o o ri *O u 3 I 60 Sh Uo CHM 5 Ho o *1 CM O & *2 13 ff 1o *.a oa.v\ K . WH pOv Oo oOof- vop-3 o 1_A p scrsao NEV 016700 WATER PCB-SD0000030648 0\ bo 4* V\ CM bo <-t oo b cm u SB 6a 43 0 Co IS "-t rt b SB OAA NEV 016701 WATER PCB-SD0000030649 r BTST3J0 #^9T0A o M b P b 6H0 <M H b O-< o o b b, NO |g bat OAA ft, NEV 016702 WATER PCB-SD0000030650 CM "O Mica B583, B602, B$6i|, B603, and B673 OIAVNO'AOOVN OOlA O O IA' O V\ . n tM rl N ' N n N CM cn CM CM CM CM H NEW 016703 WATER PCB-SD0000030651 / I I i CM aUto u CM h O H O2 < u CM O M tB ca S8 *O s*a e. mau\ Wrt NEV 016704 WATER PCB-SD0000030652 & <ac J k CM CM 5 8s, u 2 H T7S ci u e V i U (M O a sr bl eLn <3,4 NEV 016705 ! WATER PCB-SD0000030653 NEV 016706 WATER PCB-SD0000030654 * CM 0 z1 M l o ua S.V\ NEV 016707 WATER PCB-SD0000030655 k NEV 016708 WATER PCB-SD0000030656 "8 O .3?: *_ls i x 4I a 'O 0 01 I2 H fit K fa o f it 8 3 11 'll fOa *s. <2 <3 01< foa i 0s 0i faa. *'! i ir as.v*\ itj a, g 8 S 8 8 S 8 8 & S m n n nn n n SB9JQ NEV 016709 WATER PCB-SD0000030657 8$ 8S8 O OO O WO_ W - W O O wO O o **% CM CM CM CM n n n n m n pi .SOT40 ^qST*ft NEV 016710 WATER PCB-SD0000030658 .! i v 'a c i i ! I s^8 S8 n n ct 8*88^8 <m <m M\ CM CM saruiO NEW 016711 WATER PCB-SD0000030659 oo \A O _ m nn m sc*.xo NEW 016712 WATER_PCB-SD0000030660 f. \ \ I ;{ \ \ I n eoO o *a0 n "a o O - X <A 3 <**C0 12 38 a QH .CO uw *%xC3vgi NV 016713 WATER PCB-SD0000030661 i \ f I tatufj NEV 01671^ ;f l f \ WATER PCB-SD0000030662 I WATER PCB-SD0000030663 ry e * ry a t, cs o A e0 oc H 5 e9 *o a n a! 15" 12 C.C ,* U0 NEW 016716 WATER PCB-SD0000030664 . *BJO NEW 016717 M 0 .m QU WATER PCB-SD0000030665 BxporInant No. 2 Controls Subjootod to Conditlonod A ir $ u NEV 016718 WATER_PCB-SD0000030666 ExpariMonfe M o..2 C ontrola SubJoe ted to C o n d itio n e d A ir NEV 016719 WATER PCB-SD0000030667 Exparlatent No. 2 C ontrols Subjected to C onditioned A ir I OlAO U%OV\ J CM PM r* re**10 'WT NEV 016720 WATER PCB-SD0000030668 X Or*>k 5 Tt NEV 016721. WATER PCB-SD0000030669 WATER PCB-SD0000030670 Experim ent No* 2 C ontrols SubJoetod to Conditioned A ir I f \ : i t u , ft. u iI ( S u b ja o te d to C ondit` ramJ-Q *4M9tA* NEV 016723 WATER_PCB-SD0000030671 I curaao "9 N *a O O 9 v 1h tz ac NEV 016724 WATER PCB-SD0000030672 "0 eco e u <M <2 a o ft M CO e 13 hf O.C a* NEV 016725 WATER PCB-SD0000030673 fig u re 43 m a 0 i sb < 1 o u 0 wi5 O "0 is tg 0a4s* Mw NfcV 016726 WATER PCB-SD0000030674 *4. a* t 8 3 *3 CM 3 O NEW 016727 WATER_PCB-SD0000030675 U II tuduilrio The Toxicity of the Vapors of . Aroclor 1242"and Aroclor 1254' J. F. TREON, Ph.D., F. P. CLEVELAND. M.D., J. W. CAPPEL. and R. W. ATCHLEY The Kettering Laboratory, Department of .Preventive Medicine and Industrial Haaith Collaga of Medicine, University of Cincinnati Investigation of the physiological re Aroclor 1242 per liter of air Is equivalent sponse of animals to the inhalation of to 96.9 ppm by volume. two Aroclor*, namely 1242 and 1254, was . Aroclor 1254, which corresponds to penta- undertaken because in the earlier literature1 chlorobiphenyi, is a light, straw-colored, dealing with the toxic effects of Aroclors, viscous liquid. The positions of the chlorine no distinction has been drawn between the atoms have not been established. The mo toxic 'affects of chlorinated biphenyls and lecular weight of pentachlorobiphenyl Is those of .chlorinated naphthalenes, despite 326.445, of which 54.3% is chlorine. Ben the differences in' the chemical composition, ignus has reported that Aroclor 1254 con physical characteristics, and Industrial ap tains 55.0 = 0.5% of chlorine. Bulletin No. plications of these classes of compounds. In O-P-115 describes Aroclor 1254 as having a the case of the chlorinated biphenyls, at specific gravity at 25*/25*C of 1.538 to least, further toxicological investigation and 1.548; a distillation range of 366* to 390*C, consideration were required to demonstrate a refractive index (D-line at 20*C) of 1.639 the relationship between the extent of their to 1.G41, and a Saybolt Universal viscosity chlorination and their toxicity. - ' at 100' F of 1,800 to 2,500 seconds. The vapor pressure at 150*C Is about 1.3 mm, Properties and at 200* C is about 9 mm. The material A gocLOK 1242 is a light, straw-colored, mo is soluble in most organic substances, but it bile liquid. According to Benignus, of is insoluble in water and glycerine. At 745 Monsanto Chemical Company, it contains mm of mercury and 25 *C the concentration . 42.0 0.5% of chlorine, an amount which of 1 mg of Aroclor 1254 per liter is equiva corresponds to a chlorinated biphenyl with lent to 76.5 ppm by volume. three chlorine atoms in unassigned posi tions. The physical and chemical properties Experiment*! Method are given in Monsanto Application Bulletin EXPOSURE to THE vapor: In a preliminary No. O-P-115. Its specific gravity at 25*/ " experiment,(No. 1) with the vapor of 25*C Is 1.376 to 1.388; it has a distillation Aroclor 1242, a group of animals was con range of 325* to 360*C; a refractive index fined for seven hours on each of five days (D-Ilne at 20*0 of 1.627 to 1.629; a Saybolt per week in a rectangular plywood chamber Universal viscosity at 100*F of 80 to 93 (volume 600 liters;, of which the inner seconds, and a flash point (Cleveland Open metal lining was coated with a baked chemi Cup> of 176*-180*C. Its vapor pressure is cally resistant plastic, through which was about 4 mm at 150*C and about 30 mm at passed a stream of air laden with Aroclor 200*C. Although Insoluble in water and gly 1242 in a known concentration. The stream cerine, it is soluble in most organic sub of air, conditioned with respect to tempera stances. At 745 mm of mercury and 25'C, ture t75* = 3*F inside the chamber), dust the concentration of 1 mg of the vapor of and humidity, entered the chamber through an "Anemostat," located at the center of its PmaU4 at tha Straataanth Annual Matting of tha Amuwak Ikoustmal HYCitNC AiaocurtON, PMUJaipbia. April tl, list. - top, at the rate of 500 liters per minute, as measured by an inclined manometer attached SHiv to a vt ulthdrr througl wall at rides. Aroc heated 138*C. the Hr l Fig. In a on the tore of WhL 1242 * ing th carries the fir inner over li tained rninut In t expert their on eat week clor v of cor was c fora i DEPOSITION EXHIBIT /f WATER_PCB-SD0000030676 -?4 CHLEY IH> i'Ment i L ataolored. hlorine h mo- >{ .1 is Ben54 con- Vo. Ia 538 t 300* r. j x. The 3 nun. i rial it it At 743 tration ' *: iva- riary i r of iL eone day* leather f iner t (mi ll wa* iroclor l earn 4 era, du.-t trough J:i:; tWheu Industrial Hygiene Quarterly tos to the Aroelors, in another cham ber supplied only with condi tioned air. , A second set of experiments with each of the Aroelors at lower concentrations was con ducted in the same chambers. In the third experiment with Aqs- clor 1242, which was heated to 55* to 60*C, the air passed over the liquid at the rate of 800 liter* per minute. In the second ex periment with Aroclor 1254, the liquid was maintained between 115* and 125*C while 700 liters of air per minute passed over ita surface before entering the chamber. The animals exposed in each of these experiments, as well as a second group of controls, ScktmtlU rfUgrtm f quipnwnt lot voUtllit'ing Aiaelor, diamWr. end qulpmni lor eombuillon and coil. ctlaa ef lamplaa. were confined in separate 600 liter chambers during a seven hour period on each of five days per week during the week pre to a venturi meter (Fig. 1). The air was ceding the initial exposure of the test ani withdrawn by auction from the chamber mals to the vapor of the respective Aroelors. through an exit tube located on the rear In all experiments, the period of exposure wall near the floor, equidistant from the or confinement, (controls) was seven hours sides. ' per day on five days per week for several .Aroclor 1242 was volatilized from a weeks. In all instances, the original group bested glass well maintained at 132* to consisted of one cat, six guinea pigs, ten 138'C. The air passed over the surface of mice, four rabbits, and ten rats. Among the liquid before entering the chamber both the experimental and control groups, (Fig. 1). several animals died from extraneous causes In a second similar experiment performed during the course of these experiments and on the same Aroclor (No. 2), the tempera were replaced very soon thereafter. In the ture of the liquid was kept at 100* to 105*C. second experiment with Aroclor 1242, in While the second experiment with Aroclor the first experiment with Aroclor 1254, and 1242 was in progress, an experiment involv in the comparable group of controls, repre ing the'vapor'of Aroclor 1254 (No. 1) was sentative animals were killed throughout the carried out in a second chamber similar to experiment ' the first except for the absence of a plastic Method fob the determination of the inner lining. The rate at which air flowed aroclors IN air: These materials were de over liquid Aroclor 1254, which was main termined quantitatively by virtue of the fact tained at ISO* to 135*C, was 400 liters per that, on thermal decomposition, they yield minute. hydrochloric acid which with silver nitrate, In order to accustom the animals to the forms a suspension, the density of which experimental procedure, they were kept in could be measured by means of the Beckman their respective chambers for seven hours spectrophotometer at 500 mp. on each of four consecutive days during the On each day, two samples of air from each ^eek prior to the introduction of the Aro- chamber were collected by passing air at the elor vapor into the chamber. For purposes rate of one liter per minute (for 15 min of control (No. 1), a third group of animals utes in the case of the higher concentrations ** confined, throughout a like period be and for 45 minutes in case of the lower con fore and during the period of the exposure centrations) through a fused silica combus- m WATER PCB-SD0000030677 i %" sot ' June, isjij *1 fritted-glass bubblers (Mine Safety Ap pliance Company No. 43867). The 0.1 K sodium hydroxide was pre pared by dissolving 4.0 g of pellets (ACS specifications 0.01% Cl) In about 60 ml of double-distilled water. To this were added 10 g of arsenic trioxlde (chloride-free) dis solved in water and filtered through What man No. 42 filter paper. The combined solu tions were diluted to 1,000 milliliters. The sample was transferred to a 25 ml graduated cylinder to which one drop of a solution of phenolphthalein was added (1 g of phenolphthalein dissolved in 100 mi of CP methanol). The solution was neutralized ._ Rs- * Homldifitr, furnsca, and aampKnf (swart. with 3N nitric acid (190 ml CP concen- * trated nitric acid diluted to 1,000 ml with double-distilled water) and diluted to 23 ml with double-distilled water. One mi of 3N nitric acid was added to obtain a pH of 1. After adding 1 ml of a solution of silver nitrate (3 g of AgNO., diluted to 1,000 ml with double-distilled water), the sus pension was mixed by inversion. After standing 30 minutes, the transmission of wave-length 500 m/i was measured in a 50 cm cell by means of a Beckman spectropho tometer which was set against a correspond ing cell containing a reagent blank. The amount of Aroclor was estimated by means of a standardized curve prepared from known quantities of sodium chloride. A curve presenting the transmission values Fig. J. Sampling lawan. (MiJgat lubUart) at 600 mp of suspensions prepared from sodium chloride ranging from 5.0 to 120 micrograms per 25 ml of final suspension is tlon furnace, and then through two midget shown in Fig. 4. On the basis of 42.0% of bubblers, in series, each containing 10 ml of chlorine in Aroclor 1242 and 65.0% in Aro 0.1 N sodium hydroxide. A quartz tube (13 clor 1254, one microgram of sodium chloride mm outside diameter, 7 mm inside diameter is equivalent to 1.442 micrograms of Aro and 16.25 inches in length) contained sev clor 1242 or to 1.1028 micrograms of Aro eral strips of folded platinum foil. The mid clor 1254. -, dle nine inches of the tube were wrapped with a heating unit. The heating unit of Experimental Results the furnace (Fig. 2) consisted of 20 feet A/TORTAUTY--AROCLOR 1242: No signs of of B. and S., gauge 22, nichrome wire (1 * intoxication were observed in any of ohm per foot), and was covered with as the members of a group of 31 animals (Ex bestos cement The sample of air was hu periment No. 1), all of which survived midified by a cotton wick saturated with throughout a period of 24 days, on 17 of water placed 0.5 inch upstream from the which they were subjected to the inhalation quartz tube. With the furnace maintained of air bearing 8.6 micrograms of Aroclor at 660*C, the Aroclor was decomposed In 1242 per liter (0.83 ppm) for seven hours the presence of H:0 to form HC1; this was (Table I). - absorbed in 0.1 N sodium hydroxide in One cat, four guinea pigs, six mice, two midget bubblers (Fig. 3) equipped with rabbits, and eight rats survived without . Industrial t -lirns of Int-, tholr exposr j. for seven tiny* over a nir bearing 1*142 in the mlcrograms Certain otguinea pigs tiit. and two so fewer pe Table !)-* trnneous c: posed aniir. those whic!' correspond:; i No. 1, Ta : mice, thre*. rabbits, an following s hours on l of 122 da which the ilust, hum: no vapor o vur* (thre bit, and t\ periods of S MalvHftt \rv<lr tUi . Nrter ISIS Wtat IU1 I'pntM____ . ( tk % . <ft* III* i lla# #t tk# MaUttil \erWr 1*4. Itt * Mf*t Hut ( tk ( Ike Hi* mi lint WATER_PCB-SD0000030678 I G tO tttlfA ItC f' ^ ltW A R Y tou ; ifety A|i. de was pro(ACS 0 80 ml of Lara added ide-free) din. :--gh Whst>i lined soluII...era. 1 to a 25 ml n* drop of h a added i 1 I 100 ml of ia neutralized i CP conceit.,1 0 ml with u. d to 23 ml na ml of 3X n a pH of 1. :l of alive r it i to l.OOii sr), the atii<tralon. After ' 'viaalon of d in a 5<i. n apectrophoa c* apondb i ;ln<__ed by v.4 prepared Hum chloride, n' don valuer r ared front m S.O to 120 luapenaion U i * 42.0 (c of \l i4* in Arod ;m chloride rams of AroT'Tna of Aro- Vo signs of e' in any of I imtU (Exlich aurvived ava, on 17 of tj i Inhalation r{ of Aroclor r seven hour.'* s' : mice, two ed without Industrial Hygitnt Quarterly tor signs of Intoxication throughout their exposure (Experiment No. 2) for seven hours on each of 82 ==j_____ 1__________ too os Mm60TL days over a period of 120 days to air bearing the vapor of Aroclor 1242 in the concentration of 6.83 micrograma per liter (0.66 ppm). Certain other survivors (two 1 guinea pigs, three mice, one rab l!bit, and two rata) were subjected to fewer periods of exposure (cf. Table I). The deaths from ex 1 1Hu --* Iwsail a an o s*wr oa, mM C s i i|4in| a Vet s MOg OM, SM yHOM traneous causes among the ex posed animals were fewer than those which occurred among the corresponding group of controls (No. 1, Table I). One cat, three mice, three guinea pigs, three l >1 l *1 rabbits, and eight rata survived following confinement for seven hours on 84 days over a period of 122 days, in a chamber in H.4. Standard cum Ur the detarmlnettee of ArocUr 1141 and Arecler I2S4. . which the air, Conditioned with respect' to In another experiment (No. 3, Table I-A) dust, humidity and temperature, contained in which animals were exposed to air bear no vapor of either Aroclor. Ten more survi ing Aroclor 1242 in the concentration of 1.9 vors (three guinea pigs, four mice, one rab micrograms per liter (0.18 ppm) for seven bit, and two rata) were subjected to fewer hours per day on 150 days over a period of periods of confinement (cf. Table I). 214 days, the incidence of mortality among Table L Summasy or Data on Mortality Amono animals Exposed to the Vapor or Ahoclos 1242 os Aroclos 1254 in Am l (Higher concentrations) - . ` Material coaoootrouom T/l >pa Rapecara (Rear.) Cato Number of Aaimala that Suirlvod Caiaoa Pin Iftoo Rabbits Sato East. Ne. Aroclor lift Aroclor 1141 Aroclor 1114 Coatrol Ml Ml Ml 441 iM 1.41 if x f Uif Uit 94 x f 1 1 1 1 I IS 4 IS 1 I'.1 S',*.* S' 10*, S If.* 4', IS* I 1*.* 4* IS*.* 1 'One of tho aaioaali was apoaod on only M to 11 dayo *086 of tho animals was aspoood on only 41 to II days Kino of tho animals was oipoood oa only 11 to II days 4firs af tho asiec wort oxpoood on only f4 days *Throo of tho mleo wero oipoood on only 14 to 19 days *Oac of tho rabbits vu aapoood on only 19 days -Tails I-A. Summasy or Data on Mortauty Amono Animals Exposed to the Vapor or AiocLO* 1242 os Akoclo* 12S4 in Ais (Lower concentration!) t - Keterie! Artxtar lilt Arwlor 1U4 Coatrol Ceee.elr.Ue. in v t.M ft.ll Ml Ml (hears) 111 x f 111 x f US x 1 Cato t t 1 Numborof Animals that Sanrlvod CuJnon Pits I* . I> MIm EihUta , l,v 10*.'.'.* 4 IVsVsVs* I*.*.1 Bete ! 11* 101 Siyb N*. S s t of tho animats u tspooed on only It to I? fan of tho animals u oipoood on only 41 to If days *Ooo of tho animals was axpod on only 110 to 111 4ars 'Om of tho animals was tapoood aa only I to II days *Onc of tho animals woo tapoood on only lOt to 111 days *Ons mouM was tapoood on only II days WATER PCB-SD0000030679 iOS /ne, l9St Industrial Hyg tht various species, with the exception of the rabbits (which died during an epidemic of pneumonia), was no greater than that en countered among a similarly constituted control group (Experiment No. 2, Table 1-A). . Tails II. Thk average Chances in Weioht or Exrcxj- MENTAL AND CONTROL ANIMALS (Higher concentrations) -4 .One cat, five guinea pigs, six mice and nine rats survived throughout the entire Species f ' period of their subjection to the vapor of Animal Number Aalub Avyrese Errnwl m 3 t.HUI Wefskl <k*l UllUI Wrf'kt Aroclor 1242. Seven additional survivors (one guinea pig, two mice, three rabbits, and one rat) were exposed intermittently to the vapor of Aroclor 1242 over a shorter period of time (cf. Table 1-A). Among the group of controls (Experiment No. 2, Table I-A), one cat, five guinea pigs, four mice, two rabbits, and nine rats survived through out the entire period of confinement, for seven hours on each of 150 days over 213 days in a chamber supplied with condi tioned air. Eleven other control animals (one guinea pig, six mice, three rabbits, and one rat) survived during a shorter total period of intermittent confinement (Table I-A). No signs of intoxication were ob Amtsf 1141 . 0.40 >/l Experiment Ma. 1 Cat Guinea Flf Mmm Rabbit Rat 1 9 If 4 19 UU e.ii 0.024 2.901 O.lt* 4- at - a* +u + r.r 4 at Arcelor IS4S C4I 7/1 Experiment No. 2 Cat Guinea Flf Itouae Rabbit lUt 1 1 10 2.221 0.404 0.021 f-121 * 0412 +tsa 444.1 4ia 44M +11.4 Aroclor 1294 4.40 7/1 - Experiment Ma. t Cat Guinea Flf Meuse Rabbit Rat 1 1 If 4 10 UU 0.400 .on tm 0422 i4tu 440-1 * +*as *- +I0.T + a* served among experimental or control ani- Conditioned Air Control Experiment He, 1 mala. Mortality--aroclor 1254: One cat, three guinea pigs, four mice, two rabbits, and nine rats survived throughout their expo Cat Gulaea Flf Mouse Rabbit Rat 1 0 2 4 10 UU 0.4 IS 0.022 2.244 0411 + 0.0 4ia< 411.4 4444 411.4 sure, for seven hours on each of 83 days over a period of 121 days, to air containing Aroclor 1254 in the concentration of 5.40 micrograms per liter (0.41 ppm). Certain other animals (three guinea pigs, six mice, Table II-A. The Average Changes in Weight or the SurvivoRS Among the Original Grouts (Lower Concentrations) two rabbits, and one rat) were killed for Arertte Chief* examination after 33 to 74 periods of expo sure (Table 1). The incidence of mortality Spetln from extraneous causes among the exposed AalmaJ In Weiibt Number Averete Expruued ae at loltUl Weltkt Feree.Utt el Aalmete Iks) Initial Welfht F * and control animals (Experiment No. 1), with the exception of the rabbits, was com parable (Table 1). One cat, four guinea pigs, six- mice, four rabbits, and 10 rats survived (Experiment 2, Table 1-A) following their exposure for seven hours on each of 160 days over a period of 21? days to air containing Aroclor 1264 in the concentration of 1.5 micrograms per liter,.(0.11 ppm). Eight other animals Aroclor 1242 1.9 7/1 Experiment Me. 1 Cat Gulaea Flf .Moum Rat 1 i 0 0 L2I0 0414 o.ont 0414 4UO.O 4 4SJ 4 na 4 IM Aroebr 1294 14 7/1 Experiaeat Ma. 2 Cat Guinea Flf Move Rabbit Rat 1 4* 0 4 10 2418 9.4IT 0.0242 loll 0.222 4 ias 4 oas 4 IT.0 4 44.0 4 *11 _ ^0.01 >a -- >0.00 $0.00 $100 li * (three guinea pigs, four mice, and one rat) Coedittoned Air Control* Experiaeat Ko. 2 survived through 30 to 139 periods of ex posure. The incidence of fatalities among the exposed group was slightly less than that encountered among the controls (Ex Cat Guinea Flf Mouee Rabbit Rat 1 0 4 4' 9 2429 0.100 9.9202 tiu 0412 -u 4 oar 4 las 4 oat 4 oar -- -- -- -- periment 2, Table I-A). 'UcMes tve early replacement*. No general :ion "era noti animals during Growth: Th the cbsnges In various groups mid II-A. Although co: weighed durin; Aroclor 1242 grams per litei the gulnet pig. |v In weight dt pesplte a small in this experlm Ing weight at t nt exposure. The animals dor 1242 in micrograms pe nr to Aroclor . 5.40 mlcrograa (i, with the e that were expc 1254, grew eqi Experiment, * The growth r nnimais in the the lower coni Aroclor 1242 ( Summary or th the Body W: ! Arot Aro* . Con- 1 i im Amin UIS Areeter III! Ar,,lor IU4 Antler IU4 A feeler ItSO Afeeler ItU Arecler ISIS lfeeler ItU 'feeler IU4 Afetter 1114 Areeter IU4 lentrel '*treO . eiwrel 'tMr.1 . t'eeirel I'eMrel WATER PCB-SD0000030680 I I lnduitrial HygUna Quaritrly SOB . p xrai. mtt Chan. Ahl <"1i; iintf ibl Wtltki IKD > MU +IM No general or specific signs of intoxlcstton were noted among the experimental animals during or after their exposure. GBOWTH: The pertinent data relating to the changes in weight of the animals of the various groups are presented in Table II and II-A. Although comparable controls were not weighed during the Arst experiment with Aroclor 1242 (17 x 7.0 hrs.--8.60 micro* grams per liter) all of the animals, except the guinea pigs, appeared to gain normal* ly in weight during the period of exposure. Despite a small net loss by the guinea pigs in this experiment, they were actually gain ing weight at the termination of the period of exposure. The animals exposed to the vapor of Aro clor 1242 in the concentration of 6.83 micrograrae per liter (Experiment No. 2) or to Aroclor 1254 in the concentration of 5.40 micrograms per liter (Experiment No. 1), with the exception of the guinea pigs that were exposed to the vapor of Aroclor 1254, grew equally as well ss the controls (Experiment No. 1, Table II). The growth of the surviving experimental animals in the groups initially exposed to the lower concentrations of the vapor of Aroclor 1242 (Experiment No. 3,1.9 micro grams per liter) and Aroclor 1264 (Experi ment No. 2, 1.5 micrograms per liter) was unaffected. No significant differences in the average change in weight of the expertmental and the control groups wera found by the use of the "t" test (Table II-A). Weight op liver and kidneys: The weights of the livers and kidneya and the relationships of their weights to. the body weights (expressed as grams per 160 (yams of body weight) of the animals that sur vived following exposure to the vapor of Aroclor 1242 in the concentration of 6.83 micrograms per liter, are shown in Table III, which also gives the corresponding data on the controls. Comparison by the "F" test of the variances of the ratios of the livers or kidneys to the body weights of rats, guinea pigs and rabbits exposed to the va por of Aroclor 1242 (6.83 micrograma/ liter), with those of the controls, revealed no significant differences (P>0.05) ex cept in the case of the livers of the rats. Application of the "t" teat to the mean values (Table III) revealed no significant differences between the test and control groups of guinea pigs, rats, and rabbits. Comparable values for rats, guinea pigs and rabbits exposed to Aroclor 1254 in the concentration of 5.40 micrograma per liter t ac SunihwUP* tUlf #! u *1,-44 lUiial ffdlbt P 3 "** 1. > St. 1 h . Table III. or or orSummary the Data on the Relationship the Weight the Liver ob the Kidneys to orthe Body Weight Animals Exposed to the Vapor or Aroclor 1242 or Aroclor 1264 Aroclor 1242 - 6.83 y/1 - 82 x 7 hrs. Aroclor 1264 - 6.40 y/l - 83 x 7 hrs. Controls - 0 y/1 - 84 x 7 hrs. Cn petti Ones Avemfe Specie* 0(|ii ef Weicbt Aiiael a (t) Average Body Welfbt <*> Retie ef Weicbt ef Ofvu m 144 to BUy Weicbt t r' Aroclev 1949 Antbr ties . Antbr MIS Awiw mi Antbr IU4 Antbr !U Antbr Uit Antbr mi Antbr tilt Antbr IU4 Antbr Mil Antbr 1UI Ctttnl Ctttnl Ceetrei Ctttnl Ctttnl Uver Ur ' Liver Liver Uvr Liver KJ4eye XUier* KMttrt ICUeere fl4ert KMeers Liver * Liver liver Ki4e*r* Kidney* Kidney* Ktt Guloe* Plff lUkUt Ktt Golem Pif Rebbit Ret Guloee Pit RebUt Rtt Guloee Pis RebUt Ret Gutee* Pic Rebblt Ret Guinn Pll R.bkJt 4 S 11 4 4 9 14 4 14 4 4 14 1 4 114 91.1 till It* 91.4 110.4 Ml M III 9.1 4.4 III M ao-i tOtrt L4 4.1 ll.K 944 444 4.499 914 111 MIT 944 444 4.491 914 ' III 4,111 949 914 Mil 949 444 i.m 4.44 4.11 4.44 1.94 441 Ml 4.119 4.441 0.414 4.114 4.144 4.444 4.11 441 9.11 4.141 0.111 4.144 UUl 4.4941 14414 9.9499 0.0004 1.SIU 4.1149 4.1911 4.41TI 4.4144 44411 4.9141 ----n -- -- S.IS-MS >MS s.is-sas ^4.41 >UI >S4S MOAH >MS s44l >u -- -- A s v y fl; m i- v m o 7 D I'- WATER_PCB-SD0000030681 no /line, ISst iifMrisl are also riven In Table III. Application of centration of 6.83 micrograms per liter; (2j the "t" teat to the differences In the mean to the vapor of Aroclor 1254 in the concen values which characterized the experimental tration of 5.40 micrograms per liter; and and control animals (Table III) shows that (3) to conditioned air alone. In Table IV the weights of the livers of the exposed rats the percentile relationships of the dotting were significantly greater than those of power have been calculated arbitrarily by . the controls, the ratio of the liver to the dividing 100 times the average clotting time ' body weight of the former being 5.34 g per of.the blood, of the control animals by the 100 g of body weight, that of the latter being clotting time of the blood of the experl- ' 4.16 g per 100 grams. In all other Instances mental animals on the same day. No dimin tested (livers and kidneys of both guinea ution in the clotting power of the blood, as a pigs and rabbits and kidneys of rata) the measure of the Impairment of the function ? differences were statistically insignificant. of the liver, was induced by the exposure of ' The organs of the animals exposed to the the animals to the Tapor of Aroclor 1242 - lesser concentrations of the vapor of the in air in the concentration of 6.83 micro- . Aroclors were not examined In this manner grams per liter, or to the vapor of Aroclor ; because of the borderline character of the results associated with the higher concen trations. 1254 in the concentration of 5.40 micro- grams per liter. i This functional test was not applied to V Liver FUNCTION: Data were obtained as animals subjected to the lower concentre- to the apparent prothrombin activity of the tions, because of the negative results ob- blood (measured by the method of Kato1) talned when the animals were subjected to of certain animals that had been exposed the higher concentrations. i (1) to the vapor of Aroclor 1242, In the con- Hematological results: Determine ________________________________________ \ Table IV. *1 The Effect or Exposure to the Vator or Aroclor 1242 on Aroclor 1254 in Aw for Setzx Hours Per Day on Five Days Per Week Over a Period or Several Months Upon the Ap parent Prothrombin Content or the Blood or Cats and Raibits - JUkUon of "Prothrombin Tim#" of Control ond Expoaod Antmob gxprwid Arbitrarily In Trm# of FrcnUo Chonvo In Ckttiny Tim# #f Blood ' 190 i Cbttlnv Tim# #f Blood of Control Anlmoti Cbttlnv Tim# of Bbod of Expoood Anlmok Tim* of SftmpUnr In lUktlen to FtrW of Expoonr* __________ Dor* Boforo Tint F.rM briit HnmWr of Toriod AfUr Whbh Bbod Woo Drawn Arocbr 1141 l.tt y/I Experiment Ho. 1 " Cit A-441 JUbblU (Avirut) 100.0 00.0 I1M MM 1.4 IU4 . a, 00.0 fl.O 100.1 M M lOM 10I.S too.o `* 1 s 1* " SO 4S IS ot 1 Arobr 1114 0.40 7/1 * Exporimont No. 1 Cat A444 JUbbtta (Afinit) ' 1M.S 100.0 u USA MM Hi M.I i.t N.O 1.0 N.O 0. lOM Ht.i 01.0 1.4 f.l 1N.0 s 1 i0 10 u so 11 Ia 00 00 i Dotormlnolbn mod* U don ofWr find poriod of oxpmuro. 'DoUrminoUon mod# 14 doyo oftor llrd ported of txpotiir^ fur. Ayerjm ;ip Hem- \m S24R ' ..*rbr ISS4 MtroU - *Vo*wo iWtUW Ilona of tt li-ucocytes . the periphi rabbit* sul Aroclor; mirrogram V. which a control ani to dlfferen. -xperiment Ixtrderline vncea th t` the coneen btood of tl were small Item of tei. miimals w<. lent was Y individual; regarded ;; Somewhat; mined wh:; subjected : lion of 5.4 logical p ..lightly elf guinea pig' Pathol; perimentar 15 days ai In the 8 8.60 tnic: mala were* gross xa, reveal an< sues of c rat, four g and six n; microscop' ' found In i In the! 1242 (&8f. : t t WATER PCB-SD0000030682 c ta tta iA tuj^ *, t *-. t eoncenInert and v Table IV H` dotting lij irily by on.n* tlm tats by thf tt expert\ > diininl.jod, at it ne (unction :r~osurs of d or 1242 Se'.,J microof Aroclor 5/.G micro* i jpiled t- concentmresults oh*ii lected t I. Determlnn- a Seven roN THE Ar- iul NitUr ' frM Alur IM V Drava 18 it 41 it 88 8 18 18 88 ti 48 8 88 Industrial Hygiene Quarterly sti Taslc V. The Avnuet Numscrs or Erythrocytes and Leucocytes and the Average Concentration or HcMOGLoatN in the Peripheral Biooo or Experimental and control Animals Ceafn4 Arwter I1U Amhv 1884 Cntnh CMmtniidi WII 8.88 8.4$ Numtar 8 1 1 f Aitnul Gultra FI* lU&Wt CuIm ris lUfcWl Gultra FIc SUbfeU Crytkracylrat <Th*gtn4) V*f rara* 1.781 4.888 8.188 . 8.418* 8.788 8.888 Ltueocytrar Mr raa* ie.ui> 8.788 18.887 18.811 18.184 11488 HratfeWti S/I88 rat 14.4* iu IU> IU IU IU I iViIm Iftlllmlb' (rraUr ihaa tkit ylildit by wttrahb Horn of the numbcra of erythrocytes and first experiment with Aroclor 1254 (6.40 leucocyte! and of the hemoglobin content in micrograms pec liter), which were carried the peripheral blood of the guinea pigi and out simultaneously, the deaths, that occurred rabbita subjected to inhalation of the vapor among the test and control animals were of Arochr 1242 in the concentration of 6.S3 the apparent result of an appreciable in micrograma per liter, are given in Table cidence of pneumonia. The lesions of cer V, which also includes comparable results on tain of. the animals were those of frank control animals. Application of the "t" test pneumonia: in others such lesions were not to differences in the mean values for the fully developed. Degenerative changes in the experimental and control animals yielded viscera were usually found, in varying de borderline evidence of significant differ grees of severity, in association with the ences in the number of leucocytes and in pneumonia, but In certain animals, test and the concentration-of the hemoglobin in the control alike, the degenerative changes were blood of the guinea pigs. These differences more evident than the pneumonia. The were small and of opposite sign (the num similarity of the lesions in test and control bers of leucocytes in the blood of the test animals, and the lack of characteristic evi animals were low, while the hemoglobin con dence of chemical pneumonitis, led to the tent was high), as well as being subject to reasonable, but not altogether certain, con individual variations, and they cannot be clusion that all of these fatalities resulted regarded as of physiological significance. from intercurrent disease among the ani Somewhat comparable results were ob mals, and not from the effects of their ex tained when guinea pigs and rabbits were posure to the Aroclors. For practical pur subjected to Aroclor 1254 in the concentra poses, this conclusion was subjected to the tion of 5.40 mierograms per liter. No physi- critique of further experiments involving .. ological significance is attached to the more prolonged exposure of animals to some Is slightly elevated hemoglobin content of the what lower concentrations. Except in the |K guinea pigs. esse of the rats exposed to Aroclor 1254, the " Pathological findings: In all of the ex survivors subjected to the higher concen periments, animals were killed from one to tration of either Aroclor had normal vis 15 days after the final period of exposure. cera. In the first experiment with. Aroclor 1242 AH of the animals exposed to the vapor (8.60 micrograma per liter), ali of the anl- of Aroclor 1242 in the concentration of 1.9 . mala were examined postmortem, and since. mierograms per liter were examined post # grass examination of the viscera did not mortem, and the viscera of most of them rereal any significant alterations, the tis were examined microscopically. The few sues of only representative animals (one deaths among the exposed group were at cat, four guinea pigs, six mice, four rabbits tributed to incidental Infectious pulmonary and six rats) were sectioned and examined disease. All exposed survivors, except two. microscopically. No abnormalities were rabbits that had hepatic lesions of coccidlo- found in the viscera of these animals. sis, had normal viscera. The control anlmala In the second experiment with Aroclor (Experiment No. 2) that died were found 1242 (G.83 micrograma per liter), and in the to have had pneumonia. Of those that sur- WATER PCB-SD0000030683 .i its /**, isst lnduatri>^ Tired, one rat, two guinea pigs, and one mouae had focal or diffuse cytoplasmic vscu- oUtlon of the hepatic cells. The remaining control animals had normal viscera. . All of the animals exposed to the vapor of Aroclor 1254 in the concentration of 1.5 micrograms per liter were examined post* mortem, and the viscera of most of them ment, liver function or hematological . changes. | More prolonged exposure of animals to a lower concentration of Aroclor 1242, (l.$ micrograms per Uteri over the period of seven months was likewise without harm in terms of growth, mortality and the absence . of pathological changes. ' were examined microscopically. A guinea In view of this evidence it is suggested . pig that died exhibited chronic pyelone that the tentative allowable concentration of . phritis, pulmonary hyperemia and edema, the vapor of Aroclor 1242 should be at least ^. and degenerative lesions in the brain and two mlcrograms per'liter (2 mg per cu liver. The visceral lesions were related, no meter), which is twice that recommended . doubt, to the renal infection. The deaths of by the American Conference of Govern four mice were attributable to acute bron mental Industrial Hygienists* for a chlorin chitis and pneumonia. The viscera of the cat ated diphenyl of unstated chlorine content that survived were normal. Of the seven The exposure of animals to the vapor of : guinea pigs that were living when the ex Aroclor 1254 in the concentration of either , periment was terminated, three had normal 6.40 or 1.5 mlcrograms per liter failed to viscera and four had slight alterations of ' induce harmful effects in the form of re- , hepatic cells characterized by cytoplasmic tardation of growth (except in the case of vacuolation. Ten mice survived, and of these six had normal viscera and tour had slight degenerative changes in the liver. Four rab bits killed one to 15 days after the last guinea pigs exposed to the higher concen tration), or of mortality, but histopatho logic' evidence of apparently reversible hepatic cellular injury was found in the . piriod of exposure had diffuse hepatic de- animals. These findings cannot certainly be g.meratlon. The character of the lesions attributed to the effects of Aroclor 1264 be vi-ried from cloudy to hyaline or hydropic d< generation and included varying degrees of fatty metamorphosis. The other viscera of these animals were normal. All of the rats ware examined and found to have slightly to moderately severe degenerative lesions of the liver. The. lesions of greatest severity were found in the rat that was killed and examined on the first day after the last period of exposure. Two rats had chronic pyelonephritis, and the remainder had slight degeneration of the renal tubules. Diteutsion: In terms of mortality, growth (except that of guinea pigs) and non-occur rence of pathological changes, the vapor of Aroclor 1242 in the concentration of 8.S mlcrograms per liter (approaching satura tion) appeared to be non-lnjurious to ex perimental animals subjected thereto for seven hours on each of 17 days over a peri od o^ 24 days. Experimental animals subjected to but a slightly lower concentration of Aroclor 1242 (6.83 micrograms per liter) for seven hours per day on each of 82 days over the period of 120 days suffered no injury on the basis of any of the following criteria: mor tality, growth, pathology, 'organ enlarge- cause of the appreciable incidence of pneu- ' monia among both experimental and con trol animals. When these nonspecific toxic changes in the viscera of the animals were associated with pneumonia, they were readi ly explained thereby, but they were also found in animals that had been exposed to the vapor of Aroclor 1254 and were free of ' pneumonia. That these may have repre- - sented toxic effects of exposure to Aroclor 1254 finds support in the fact that the livers of the exposed rats (6.40 mlcrograms per liter) were significantly heavier in relation to their body weight, than were those of control rata. It .would.appear that.this rat- , terial, which is reported to contain 55 (a of .. - chlorine, is somewhat. more toxic than is J Aroclor 1242, which contains only 42# of j chlorine.'Therefore it is suggested that the t threshold concentration of 1 rag per cu meter t of air recommended tentatively for safe in- . dustrial practice by the American Confer-( ence of Governmental Industrial Hygien- ists* is reasonable.' -.} * It should be noted that it was necessary to ] heat these Aroclors in order to Increase the s rate of volatilization sufficiently to at* *1 tain the concentrations maintained in these ` 'i A .xperlm* rial usa. ; orature may wel- Summery paotos rmals t so 8.63 r no injur rable ct: rulted it. certain ! The V A: beet Soe ; Am Fee. to ' eng by: teel* this: Tht* spo; th*: r. :;St:. pre : gin/ xe? Of of cer; out: is tht: HA; pitln<-' *i: cei; th.: ini? jraj of j WATER PCB-SD0000030684 *E! m; Ma Indiutrial Mygint Quarterly Iff tc'-fical ti i* to a . (1.9 II^d of h m in s, ince igeested at of , lean per eu . unended G rem* orinntent. vapor of if Ither . fa; id to n f re- . i ease of. v| tceni thos...-slble . In the ' a'-' be f: ai r.u- tnd con s'" toxic a! were rti :eadiere also p?*ed to. a ; ea of ' a I epre- Aroctor he liver* a per v ation those of this ms- ' % of i! in i* 4xtt of that the c> meter i! fe intjnferHyffien- J try*0 e-iS the at , tese experiments. To the extant that their Industrial usage is carried but at ordinary ternperatures, the hazard of their inhalation may well be slight or entirely absent sponsored by the Monsanto Chemical Cornpiny, whose Snandal support Is gratefully acknowledged, as is also their assistance in supplying the materials for investigation. Summary . . References . Prolonged intermittent exposure of ani- 1 mala to the vapor of Aroclor 1242 (1.00 to 8.63 micrograms per liter) demonstrated no Injury. Prolonged exposure to compa rable concentrations of Arodor 1284 re 1. Cawsuu C. K.t Purikf OWniUan* Ikb 8ys(nU TosUltr ! CrUla f Ckkriuld HHroctrboni (tk 8ufcUocu for fcmUglkU^CMOMH trilUu ia th* Air of Workroom*. J. tmd, iiiii*. im. 4 To*, t. Kato. Li Micro-protkromWft Tact with C^flDarr Whob Blood: ModlAcatioa of Qulek'a QiuatlUtfot Mtb* sulted in reversible degenerative changes in osL Amtr. J. CUm. HiHT, 1141. certain viscera. . I. America* Coafcnacc of CmrcroacoUl IMutfUI HhUbUu. TferaafeoM Limit* for 1111. AUA Ank, /ii I The work described in this article was Huttk, 11:111. till. I American Sanitary Engineering Intersociety Board Arrza almost ten yssrs of hard work on the psrt of a handful of Interssted and de termined persons, the American Sanitary Engineering Inter-society Board haa become a reality. The aseib is an incorporated organisation sponsored by the American Soelety of Civil Engineers, the American Society for Engineering Education, the American Public Health Aiioeiation, the American Water Works Association and the Federation of Sewage and Industrial Wastes Associations. The purpose of the Board ia to improve the prictlce, elevate the standards, and advance the causa of 'sanitary engineering. The term "sanitary engineering'' Is employed in the broad sente at defined by the National Research Council, and embraces the entire concept of engineering bio technology application for control of environmental stresses. As most reader* know, this definition includes industrial hygiene, air pollution control end radiological health. The Board of Trustees of the aseib is nude up of three representatives from each of the sponsoring bodies and three elected at large by the Board. We are happy to annbnpce that two of the Trustees st Urge are industrial hygiene engineers, namely lt. col. ALVIN r. mxyex, it., Headquarters Strategic Air Command, end alle.v d. brakdt, Bethlehem Steel Company. One of the most Important functions of the aseib will be examining qualified llcenied professional engineers and granting certificates of special knowledge in sanitary amgineering or one of its specialties to those who show acceptable competency in the examination. In this respect the Board will function in a manner not unlike that of the American Bsirde of medicine. For an Initial period, outstanding members of the profession having not lest than 16 years acceptable experience will be granted certificates without examination. Pertons who have been certified by the board will automatically become members of the American Academy of Sanitary Engineer! which is the regieter of holdera of such certificates. . i The board was incorporated In October, 1965. At the present time the Chairman of tha Board U raoresso* earnest sovce, University of Michigan; Vice-Chairman ii w. A. HAJLEZNBEIO; Treasurer is R. s. Rankin; Secretary ii FRANCIS s. elder. It if contem plated that applications for certification will become available in June. AU qualified Industrial hygiene engineers, air pollution control engineers and radiation safety en gineers having not less thsn eight years experience in this work are urged to apply for certification. The benefits that will result from this movement and the advantages that will accrue to all who become certified are many. What ia more, they will increase with the passing years. Application forma may be obtained from the SecreUry, Francis s. eider, S3 West 39th Street, New York 16, New York, or from members of the Board of Trustees. ' I WATER PCB-SD0000030685 PROJEC'i NO. jJ-U3 n/J REPORT fILE Younger Laboratories BbohemUU ... PkaimaodoqUii.... AnalyiU k IBS CLIFF CAVE ROAD St. Louis as, Missouri $c- PHONBl TILDBN 6-2 5 40 SUBJECT Toxicological Investigation Of: Inerteen PPO Monsanto Sample Number 22 Monsanto Projeet Number I-6J-12 STUDY CONDUCTED TOR Monsanto Chemical Company, St. Louis, Missouri M&roh 4th, 196J EXPERIMENTAL PROCEDURE - A) Oral LDj0 (Rats) The undiluted compound was fed by stomach tube to Sprague-Dasley strain albino rats* After the approximate Minimum Lethal Dose was determined, groups of mala and female rats ware fed in increasing doses at increments of 0*1 fractional log intervals at four levels designed to blanket the toxicity range thereby supplying data for calculation of the LD90 which was done according to a modi fication of the method of E. J* de Beer* Observations were made for toxic symptoms and the viscera of the animals that sucoumbed were examined maerosoopically. The data are shown in Table 1* . B) Skin Absorption KLD (Rabbits) The undiluted compound was applied in increasing doses at increments of 0*2 fractional log intervals to the closely clipped, intact skin of New Zealand white female rabbits* The treated areas were covered with plastio strips and the animals placed in wooden stocks for periods up to twenty-four hours, after whiah time they were assigned to individual cages* Observations were made for toxic symptoms and the viscera of the animals that succumbed were examined maerosoopically* ____ The data are shown in Table XI NEV 01T31-8 2 DEPOSITION EXHIBIT 1ST Thi* confidential report U cot to be uaad in any form of advarttdnt without written parmWion WATER PCB-SD0000030686 To: Monsanto Chemioal Coai..<ny St* Louis, Missouri Younger Laboratories Certifieate of Analysis - Page 2 (5/4/63) - 1-65-12 X7 EXPERIMENTAL PROCEDURE - (Continued) C) Skin Irritation (Rabbits) The undiluted compound was applied to the clipped, intaet akin of albino rabbits and removed after tsenty-four hours. The application was oovered with plastic strips to retard evaporation* Observations were made over a period of several days for irritation* The data, soored according to the nethod of Draise, Woodard and Calvery (Journal of Phar. and Exp. Therapeutics, Voluae 62, Deoeaber, 1944) are shown in Table III. D) Eye Irritation (Rabbits) 0*1 Milliliter of undiluted sasple was placed in the conjunctival sac of the right eye of each of three albino rabbits and observations nade over a period of several days for inflammation. The eyes were rinsed with warn isotonic saline solution after twenty-four hours* The data, soored according to the nethod of Drain, at al, are shown in Table IV. E) Vapor Inhalation (Rats) Pour 150-gran sale rats were placed in a glass desiooator, 250 an in diaaeter, and exposed for six hours to a concentrated ataospbere of vapors produced by passing a stress of air through 100 milliliters of the cowpound oontained in a 250-milliliter tall for* gas washing bottle with fritted disc* Vapors fro* the bottle passed into a one liter bottle to reaove droplets and then into the ohaaber* Air flow through the sanple was 6 liters per ninute as neasured by a calibrated rotameter* This was sufficient to violently agitate the liquid* No supplementary air was introduced inasmuch as the above supply was aaple for the animals oxygen requircnents. The animals were observed for behavior and, since there were no deaths, all were held for ten days observation. The data are Shown in Table V. SUMMARY In9rteen_PP0 A) Oral LDj0 (Rats) The Oral LD,0 for male and female rats was placed at 2570 milligrams per kilogram with lower and upper limits of 2095 to 2690 milligrams per kilogram* The compound was classed as slightly toxic by oral ingestion in male and female rats. B) Skin Absorption KLD (Rabbits) The Minimum Lethal Dose by Skin Absorption in female rabbits was found to be greater than 5160 milligrams per kilogram and less than 5010 milligrams per kilogram. The compound was classed as slightly toxic by skin absorption in female rabbits. NEV 017319 WATER PCB-SD0000030687 To: Monsanto Chemical Company St. Louis, Missouri Younger Laboratories Certificate of Analysis - Page 3 (3/4/63) - Y-63-12 SUMMABY - (Continued) C) Skin Irritation (Babbits) The compound ess classed as a aoderate skin irritant* The average naxinun score as 4.6 out of a possible 8 in twenty*four hours. D) Eye Irritation (Babbits) The eapound was classed as a aoderate eye irritant* The average aaxinua score vae 33*6 out of a possible 110 in one hour. E) Vapor Inhalation (Bats) All aniaals survived the six hour exposure as veil as the following ten day observation period. It was concluded that the vapors were slightly toxic under conditions of the test* & I0UH3EB LAB08ATOBIES The material in this report i3 to be used in development of the product nnd may be given to responsible aalea contecta, but it is not to be used by thorn in advertising copy. The source of this material is not to bo divulged until it appears in formal publications. No exceptions to the established rale may be made without the approval of the Medical iJspaiV ment in St. Louis. Customers' inquiries regarding mattava of toxicity are to be referred as before to the Medical Dtpaarb, ment in St. Louis for reply. Monsanto Cbemkal NEV 017320 WATER PCB-SD0000030688 To: Monsanto Chemical Coa.,-.ny St. Louie, Missouri Younger Laboratories Certificate of Analysis - Page 4 (5/4/63) - Y-63-12 TABLE 1 THE ORAL LD so OF 'Inerteen PPO' FOR RATS Sample Fed Undiluted Animal No. - Sex Weight Gm . 1- Female 2- Female 3- Male 4- Male 5" Female 6- Female 7- Male 8- Male 9- Female 10- Female 11- Male 12- Male 13" Female 14- Female 15- Male 16- Male 17- Female 18- Female 19- Male 20- Male 220 210 235 240 215 225 245 240 210 220 235 245 205 215 250 235 210 220 235 245 Dose Mgi/.Kgi 1580 1580 1580 1580 1580 2000 2000 2000 2000 2000 2510 2510 2510 2510 2510 5160 5l60 3160 3160 3160 Fate Survived Survived Survived Survived Survived Survived Survived Died Died Survived Died Survived Died Died Survived Died Died Died Survived Died DISCUSSION - The Oral LDJ0 for male and female ratB ms plaoed at 2370 milligrams per kilogram with lower and upper limits of 2095 to 2690 milligrams per kilogram. The compound was classed as slightly toxic by oral ingestion in male and female rat6. Survival time was approximately twenty-four hours to six days with most deaths occurring in three to five days. Toxic symptoms included severe diarrhea, loss of appetite, increasing weakness, and oollapse. At autopsy there was marked liver and renal hyperemia as well as pulmonary congestion. NEV 017321 SI WATER PCB-SD0000030689 I To: Monsanto Chemioal Company St. Loui6, Missouri Younger Laboratories Certificate of Analysis - Page 5 (5/4/63) - Y-6J-12 TABLE II THE MINIMUM LETHAL COSE OF 'Inerteen PPO' BY SKIN ABSORPTION IN RABBITS Sample Applied Undiluted Animal No. - Sex 1 - Female 2 - Female 3 - Female 4 - Female 5 - Female 6 - Female Weight Jju_ 2.3 2.1 2.4 2.3 2.5 2.7 Dose Hei/ABi 794 1260 2000 Jl60 5010 7940 Weight Change 5 Da ya Later ______Ul______ + 0.2 0.0 0.0 - 0.1 - 0.4 - 0.6 ________ Fate__ ______ Survived Survived Survived Survived Died -- 5 Days Died -- 6 Days DISCUSSION - The Minimum Lethal Dose by Shin Absorption in female rabbits mas found to be greater than Jl6o milligrams per kilogram and less than 5010 milligrams per kilogram. The compound was classed as slightly toxic by skin absorption in female rabbits. Survival time was fire to six days. The animals that succumbed lost appetite and wasted away. Movement reduced as the animals weakened but no definite paralytic symptoms were observed. At autopsy the liver had a jaundiced-like appearance and there was pulmonary hyperemia macroscopically. NEV 017322 WATER PCB-SD0000030690 !\? l /I I / i i , . Tot Monsanto Chemical Company St. Louis, Missouri Younger Laboratories Certificate of Analysis - Page 6 (3/4/6?) - Y-63-12 i ^ TABLE III SKIN IRRITATION IN RABBITS AFTER APPLICATION OF 'Inerteen PPO' Sample Applied Undiluted Anlpal_Nuiiber Nuaerical Evaluation At The End Of l_Hour 24_Hours 46_Houro ]2_Houra 120_Hours 168 Hours 1 24 4 3 2 1 2 35 4 2 2 1 3 35 5 4 2 1 Average 2.6 4* 6 4 3 3.0 2.0 1.0 DISCUSSION - The compound was classed as a moderate skin irritant. The average maximua score was 4*6 out of a possible 8 in twenty-four hours. After one hour there was well-defined erythema with barely perceptible swelling for an average score of 2.6 out of a possible 8. Inflammation increased overnight at whioh time the average score was 4*6 based upon moderate redness and slight edema. Daily improvement was noted after the application was removed. Irritation reduced to barely perceptible erythema on all animals within seven days. NEV 017323 WATER PCB-SD0000030691 i To: Monsanto Cheatcal Company St. Louie, Missouri lounger Laboratories Certificate of Analysis - Page ^ (3/4/63) - 1-63-12 ` TABLE IV EYE IRRITATION IN RABBITS AFTER APPLICATION OF 'Inerteen PPO< Sample (0.1 Milliliter) Applied Undiluted Aidaal_Nuaber l_Hour Numerical Evaluation At The &id Of 2J_gours ^8_Boura J2_Houra 120_Hour l68_Hours 1 37 30 21 15 8 4 2 29 21 15 9 4 0 3 41 30 19 12 6 0 Average 35.6 27.0 18. 3 12.0 6.0 1.3 DISCUSSION - The compound was classed as a moderate eye irritant. The average maximum score was 35* 6 out of a possible 110 in one hour. Moderate discomfort eas shown following application. Within one hour there was aild edesa, copious discharge, mild to moderate redness of the conjunctivas, and slight iris congestion and corneal dullness for an average soore of 33* 6 out of a possible 110. Discharge and cloudiness decreased overnight. Iris clarity was normal within five days and within seven days two of three animals were free of inflammation. NEV 017324 WATER PCB-SD0000030692 i To: Monsanto Chemical Company St. Louie, Missouri lounger Laboratories Certificate of Analysis - Page 8 (3/4/63) - T-6J-12 TABLE V INHALATION OF 'Inertsen PPO' VAPORS BT BATS Average Temperature Inside Chamber ....................................................... Average Relative Humidity Inside Chamber............ .. 76* F. 47 % Animal,No.- Sex 1 - Male 2 - Male 3 - Male 4 - Male Survived Survived Survived Survived _____Observationa_During_&cgosure Mild eye and nasal Irritation after twenty to thirty minutes ... Moderate discomfort ... Lethargy ... Occasional moderate dyspnea ... No serious weakness ... Slight nasal discharge. DISCUSSION - All animals survived the six hour exposure as well as the following ten day observation period. ' It was concluded that the vapors were slightly toxic under conditions of the test. Mild inflammation of the nasal nusoca was evident when the animals were removed from the chamber. Breathing was slightly irregular but no rasping sound was detected and the animals were only mildly weakened. Animal behavior was praetioally normal, in forty-eight hours. No respiratory complications developed during the ten day observation period. NEV 017325 WATER PCB-SD0000030693 . RQJECT no; t>, AJ./i REPORT FILE Younger Laboratories BtoohimlsU... Plt&MaadfifliU ... Attahf&U 128 CLIFF CAVE ROAD 1 St. Louis ss>, Missouri 7? Q&di^iccde AtuzbfUiPBONII TILBEN 63 540 SUBJECT - Toxicological Investigation Of: Pyranol 1470 Monsanto Sample Nunber 21 Monsanto Project Nnnbsr 1-65-11 March 4th, 1965 STUDY CONDUCTED FOB Monsanto Cheaioal Company, St. Louis, Missouri EXPERIMENTAL PROCEDURE - A) Oral LD,0 (Rats) - The undiluted compound was fed by stomach tube to Sprague-Dawley strain albino rats. After the approximate Minimum Lethal Dose was determined, groups of male and female rats were fed in increasing doses at increments of 0.1 fractional log intervals at four levels designed to blanket the toxicity range thereby supply ing data for calculation of the LD90 which was done according to a modification of the method of E. J. de Beer. Observations were mads for toxic symptoms and the viscera of the animals that sucoumbed were examined maeroscopioally. The data are shown in Table I. 9} Skin Absorption KLD (Babbits) The undiluted oospound was applied in increasing doses at increments of 0.2 fractional log intervals to the olosely clipped, intact skin of New Zealand white female rabbits. The treated areas were covered with plastic strips and the animals plaoed in wooden stocks for periods up to twenty-four hours, after which time they were assigned to individual cages. Observations were made for toxic symptoms and the viscera of the animals that succumbed were examined macroseopically. The data are shown in Table II 5 DEPOSITION EXHIBIT K NEV 105131 Hits eoaRdentie] Nporl If so) to be ueed in any form of advertising witHo* t written permiavion WATER PCB-SD0000030694 To: Monsanto Chaaical Co&pany St* Louis, Missouri lounger Laboratories Certificate of Analysis - Page 2 (3/4/6J) - r-63-U EXPERIMENTAL PBOCHJBBE - (Continued) C) Skin Irritation (Babbits) The undiluted eoapound was applied to the clipped, intact akin of albino rabbits and rsnowed after twenty-four hours. The application was cowered with plastic strips to retard ewaporation. Observations were nade ower a period of seweral days for irritation. The data, soored according to the aethod of Dralse, Woodard and Calwery (Journal of Pham, and Exp. Therapeutics, Voluae 82, Deeenber, 1944) are shown in Table III. D) Eye Irritation (Babbits) 0.1 Milliliter of undiluted sanple was placed in the conjunctival sac of the right eye of eaoh of three albino rabbits and observations nade over a period of several days for inflaanation. The eyes were rinsed with warn isotonic saline solution after twenty-four hours. The data, soored according to the aiethod of Draise, at al, are shown in Table IV. E) Vapor Inhalation (Bats) Pour 150-gran male rats were placed in a glass desiocator, 250 as in diaweter, and exposed for six hours to a concentrated ataosphere of vapors produoed by passing a stress of air through 100 si111litera of the eoapound contained in a 250-ailliliter tall fora gas washing bottle with fritted disc. Vapors froa the bottle passed into a one liter bottle to reaowe droplets and then into the chaaber. Air flow through the saaple was 6 liters per ainute as aeasured by a calibrated rotaaeter. This was sufficient to violently agitate the liquid. No supplesentary air was introduced inasauoh as the above supply was aaple for the aniaals oxygen requirements. The aniaals were observed for behavior and, sinoe there were no deaths, all were held for ten days observation. The data are shown in Table V. SUMMARY - SJKS521-14I9 A) Oral !,,, (Bats) The Oral LD',0 for aale and feaale rats was placed at 2230 milligrams per kilogran with lower and upper Units of 1990 to 2555 nilligrans per kilogran. The eoapound was classed as slightly toxic by oral ingestion in n&le and feaale rats. B) Skin Absorption MLD (Babbits) The Minimus Lethal Dose by Skin Absorption in feaale rabbits was found to be greater than 2000 ailligraas per kilogram and less than 3l6o ailligraas per kilograa. Animal 3, dosed at 2000 ailligraas per kilogram, very nearly succuabed. The compound was classed as sUghtly toxic by skin absorption in feaale rabbits. NEV 105132 WATER PCB-SD0000030695 To: Monsanto Cheaioal Company St. Louis, Missouri -- - lounger Laboratories Certificate of Analysis - Page 3 (5/4/63) - >63-11 SUMMABT - (Continued) C) Skin Irritation (Babbits) The ooapound was olassed as a aodarate skin irritant. The average aaxiaua seore was 4*6 out of a possible 8 in twenty-four hours. D) Bye Irritation (Babbits) The ooapound was olassed as a aodsrate eye irritant. The average naxiaua seore was 326 out of a possible 110 in one hour. E) Vapor Inhalation (Bats) All aniaals survived the six hour exposure as well as the following ten day observation period. It was concluded that the vapors were slightly toxic under conditions of the test. I0UN3ER LABORATORIES The material is this report is to be used in development of the product and may be given to responsible sales contacts, but it is not to bo used by them in advertising copy. The source of this material in not to be divulged until it appears Is formal publications. No exceptions to the established rule may be made without the approval of the Medical Depart, scent In St. Louie. Cueiomers' inquiries regarding matters of toxicity are to be referred as before to the Medical Depart ment in St. Louie for reply. -- Monsanto Chemical Company NEV 105133 WATER PCB-SD0000030696 To: Monsanto Chemical Cow^abj St* Louis, Missouri lounger Laboratories Certifieate of Analysis - Page 4 (3/4/63) - -65-11 */ TABLE I THE ORAL LD 30 OF 'Pyranol 1470' TOR RATS Sample Fed Undiluted Animal No. - Sex Weight On. 1- Female 2- Female 3- Male 4- Male 5- Female 6- Female 7- Male 8- Male 9- Female 10- Female 11- Male 12- Male 13- Female 14- Female 15- Male 16- Male 17- Female 16- Female 19- Male 20- Male 225 210 255 250 215 220 240 245 230 215 235 240 220 210 235 225 220 215 235 245 Dose Mg./Kg. 1580 1580 1580 1580 1580 2000 2000 2000 2000 2000 2510 2510 2510 2510 2510 3160 3160 3160 3160 3160 Fhte Survived Survived Survived Survived Survived Survived' Survived Survived Died Survived Died Survived Died Died Died Died Died Died Died Died DISCUSSION - The Oral LD}0 for male and female rats was plaoed at 2250 milligrams per kilogram with lower and upper limits of 1990 to 2555 milligrams per kilogram. The oonpound was classed as slightly toxic by oral ingestion in male and female rats. Survival time was approximately twenty-four hours to three days with most deaths occurring in two to three days. Toxic symptoms included increasing weakness, severe diarrhea, tremors, and collapse. At autopsy there was inflammation of the gastric mucosa, liver discoloration, and renal hyperemia macroscopically. NEV 105134 WATER PCB-SD0000030697 To; Monanto Chemical Coa^<ny St. Louis, Missouri Younger Laboratories Certificate of Analysis - Page 5 (3/4/63) - Y-63-11 TABLE II THE MINIMUM LETHAL DOSE OF 'Pyranol 1470* BY SKIN ABSORPTION IN RABBITS Sample Applied Undiluted H 1 a. Animal^ No. _;_Sex 1 - Female 2 - Female 3 - Female 4 - Female 5 - Female 6- Weight JfAi- 2.5 2.2 2.3 2.6 2.5 2.5 Dose Sfc/.Sfc 794 126o 2000 3160 5010 7940 Weight Change 5 Days Later ______ fe______ - 0.1 - 0.1 - 0.2 - 0.3 - 0.2 - 0.4 ________ Fate________ Survived Survived Survived Died - 10 Days Died - 7 Days Died - 7 Days DISCUSSION - The Minimum Lethal Dose by Skin Absorption in female rabbits as found to be greater than 2000 milligrams per kilogram and less than 3160 milligrams per kilogram. Animal #3, dosed at 2000 milligrams per kilogram, eery nearly suooumbed. The compound was classed as slightly toxic by skin absorption in female rabbits. Survival time vas seven to ten days. Toxic symptoms included loss of appetite, lethargy, and increasing weakness. Weight losses ran to forty per cent of starting weight. At autopsy there was liver discoloration and renal hyperemia maoroecopically. NEV 105135 WATER PCB-SD0000030698 Jo: Monsanto Chemical Company St. Louis, Missouri Younger Laboratories Certificate of Analysis - Page 6 (3/4/63) - I-63-ll <53 TABLE III SKIN IRRITATION IN BABBITS AFTER APPLICATION OF 'Pyranol 1470' Sample Applied Undiluted Animal Number I_Hour Numerical Evaluation At The End Of 24 Hours 48 Hours 2LI22I* i20_Houra 168 Hours 1 35 4 3 2 1 2 24 3 2 1 0 3 35 4 2 2 1 Average 2.6 4.6 3.6 2.3 1.6 0.6 DISCUSSION - The oompound was classed as a moderate skin irritant. The average maximum score was 4*6 out of a possible 8 in twenty-four hours. Well-defined redness with very slight edema developed within an hour for an average score of 2*6 out of a possible 8. Overnight there was well-defined to moderate erythema and alight edema increasing the average score to 4*8. No blistering was observed. Inflammation gradually reoeded following removal of the application. After seven days one animal was given a score of mero but very slight redness was still present on the remaining animals. NEV 105136 WATER PCB-SD0000030699 To: Monsanto Chemical C\__pany St. Louis, Missouri lounger Laboratories Certifieate of Analysis - Pag* ^ (3/4/63) " 1-63-11 'V T A B L E IV - EIE IMITATION IN BABBITS AFTER APPLICATION OF 'Pyranol 14701 Sample (0.1 Millilitsr) Applisd Undiluted Animal Number Nuasrieal Evaluation At The End Of 1 Hour 24 Hours 48 Hours 72 Hours 120 Hours 168 Hours 1 2 3 Average 35 28 35 32.6 26 21 30 25.6 17 13 20 16.6 12 8 12 10.6 6 4 6 5.3 0 0 0 0.0 DISCUSSION - The coapound was classed as a moderate eye irritant. The average maximum score was 32.6 out of a possible 110 in one hour. There was intermittent pawing at the eyes for several minutes following application. After one hour the average score was 32.6 out of a possible 110. Slight edema, slight to moderate redness of the conjunctivas, copious discharge, and slight iris congestion developed. The corneal area cleared somewhat in twenty-four hours and within five days iris and cornea were normal. Conjunctivitis and edema cleared completely within seven days. NEV 105137 WATER PCB-SD0000030700 To: Monsanto Chemical Coa_4 _ny St. Louis, Missouri lounger Laboratories Csrtifieate of Analysis - Page 8 (3/4/63) - Y-63-11 TABLE V INHALATION OP 'Pyranol 1470' VAPORS BY RATS Average Temperature Inside Chaaber ...... Average Relative Humidity Inside Chaaber 76* F. 42 * Animal No. - Sex 1 - Male 2 - Male 3 - Male 4 - Male __Fhte__ Survived Survived Survived Survived ________ 2k2*ritiona_During_Ex08ur9________ Occasional pairing at the eyes in ten to Slight nasal discharge ... Soae dteooafort .. No real weakness ... Occasional irregular breathing. DISCUSSION - All animals survived the six hour exposure as sell as the following ten day observation period. It was concluded that the vapors sere slightly toxic under conditions of the test. The vapors produced mild nasal and eye inflammation as well as soae discomfort. No weakness of consequence was noted at the end of exposure and no bronchial rales developed. Activity and appetite returned to normal within twenty-four hours. Nasal inflammation disappeared within seventy-two hours. No respiratory complications developed during the ten day observation period. NEV 109138 WATER PCB-SD0000030701 I' PROJECT REPORT FILE *' . ( L >( '. $ttdu&Uial BIO - TEST lafuwcdiyUeA, Urn. 1110 FRONTAGE ROAD NORTHBROOK, ILLINOIS Ttphon CKmNwxxI 2-3030 REPORT TO MONSANTO CHEMICAL COMPANY SUBACUTE DERMAL TOXICITY OF AROCLOR 1254 CP t NEV 009120 WATER PCB-SD0000030702 ( I REPORT TO MONSANTO CHEMICAL COMPANY SUBACUTE DERMAL TOXICITY OF AROCLOR 1254 I. Introduction and Outline of Procedure A sample identified as Aroclor 1254 was received from Monsanto Chemical Company for evaluation of 20-day subacute dermal toxicity in albino rabbits. . Skin applications of the test material, in the form of a corn oil solution, were made daily to four test groups consisting of two rab bits each (one male and one female) for a period of 20 days. These test groups corresponded to daily dermal.doses of 10, 50, 100 and 200 mg/kg/day respectively. In addition to the four test groups, a treated control group of two rabbits was also employed. The animals in this group received daily dermal applications of corn oil comparable to the volume administered to the highest test dose group. The rabbits in all groups were housed individually over the period of investigation and observations were made with respect to body weight effects, incidence of mortality, reactions displayed and status of the ' ' NEV 009121 WATER PCB-SD0000030703 ( (2 hematopoietic and urinary systems. Special emphasis was placed on observations for skin irritation produced by the test material. Following the 20-day application period, the rabbits were kept under observation until an additional 14 days had elapsed. At the end of this period, final mortality tabulations were made and all surviving rabbits from each test and control group were sacrificed for pathologic studies. Gross autopsies were performed and a complete set of repre sentative tissues and organs was taken from each rabbit and preserved in formalin. Microscopic examinations of selected tissues and organs - were made to ascertain the presence or absence of histopathologic.change as a result of dermal applications of the test material at the dose-levels employed. ' The dosage schedule and other pertinent data are presented in Ta ble I. The tosterial in this report is to be used in development of the product and may be given to responsible sales contacts, but it is not to be used by them ia advertising copy. The source of this material is not to be divulged until it appears in formal publications. No exceptions to the established rale may be tnsdo without tbs approval of the Medical Depart ment in St. Louis; Customers' inquiries regarding matters of toxicity are to be referred as before to the Medical Depart; ment in St Louis for reply. ... -- Monsanto Chemical Company NEV 009122 WATER PCB-SD0000030704 (( 3 Group T-C T-l T-n T-ra T-rv TABLE I 20-Day Subacute Dermal Toxicity - Albino Rabbits Dosage Schedule Number of Animals 2 Dose* (mg/kg/day) ** "Number of Scheduled ' Consecutive Applications 20 2 10 20 2 50 20 2 100 2 200 20 . 20 Administered as a 4. 0 per cent solution in corn oil. * ** The treated control rabbits received doses of corn oil comparable in volume to that applied to the rabbits of the high dose group. NEV OO^3 WATER PCB-SD0000030705 ( {4 II. Invest! Rational Procedure A. Experimental Animals The animals employed in the 20-day subacute dermal toxicity study were adult, New Zealand strain albino rabbits. The body weights o the test animals were in the two-to-three kilogram range. A total of 10 adult rabbits were selected from a larger population after examina tion of each animal for general physical well-being. Each animal selected for the experiment was housed individually in a wire-bottomed stainless steel rabbit cage for the duration of the test. All rabbits were fed a . standard laboratory rabbit ration* plus water ad libitum. B. Method of Application , In preparation for the test, the back of each rabbit was shaved using electric clippers. The exposure site thus prepared on each ani mal constituted approximately ten per cent of the total body surface area. The animals were then returned to their stock cages, and 24 hours were allowed to elapse before the first percutaneous- applications were made. This period permitted the skin to recover from the slight disturbance of the stratum corneum caused by the clipping procedure and also permitted the healing of any microscopic abrasions possibly produced during the shaving procedure.- At the end of the 74-hour recovery period, the calculated doses of the test material in the form of a 4.0 per cent (w/v) corn oil solution * Rockland Rabbit Ration, Rockland Farms, New City, New York. NEV 009124 WATER PCB-SD0000030706 ( (5 were gently distributed (without rubbing) over the entire application sites on the test rabbits. ` The animals in the treated control group were treated in the same manner as those in the test group with the exception that corn oil alone was applied to the skin. Following the dermal applications, the animals were returned to their individual cages to await testing on the following day. The above schedule was followed every day, including weekends, for a total of 20 consecutive days. ' C. Body Weight Effects . On the morning of the first test day, the body weight of each rabbit in every group was determined and recorded.. Thereafter, indi vidual weighings were made weekly and the data recorded as an index to body weight effects. Dose calculations were made weekly to coincide with changes in body weight. D. Mortality and Reactions -Checks for mortality and abnormal behavioral reactions were made daily during the 20-day test period as well as during the 14-day observation period. Special emphasis was placed on observations for local skin reactions in order to determine the degree of skin irritation produced by the test material. E. Hematologic Studies and Urine Analyses Blood studies including determinations of hemoglobin concent ration, erythrocyte count, and both total and differential leukocyte WATER PCB-SD0000030707 (( 6 counts, were made at the beginning of the study and at the end of the . 20-day test period. Urine analyses for reducing substances and micro scopic elements were also done at these time intervals. The blood studies and urine analyses were done on the individual rabbits in each group. F. Gross and Microscopic Pathologic Studies Arrangements were made to subject any animal which might die during the test to a gross autopsy. Also, in those instances where post-mortem changes were not advanced, sections of representative tissues and organs were scheduled to be taken for histopathologic study. At the conclusion of the investigational period, all surviving rabbits in each group were sacrificed and gross and microscopic studies were performed. The following tissues and organs were in cluded in the microscopic studies: heart, aorta, lung, trachea, liver, gall bladder,' pancreas, esophagus, stomach, small intestine, caecum, colon, spleen, cervical and mesenteric lymph nodes, kidney, urinary bladder,'testis, ovary, prostate, uterus, pituitary, adrenal gland, submaxillary salivary gland, thyroid, parathyroid, skeletal muscle, bone marrow, peripheral nerve, spinal cord, brain, and skin. NEV 009126 WATER PCB-SD0000030708 I HI. Results A. Body Weight Data Weekly individual body weights for each test group and the con trol group are assembled in Table II. Group T-C T-I T-H T-m T-IV TABLE H Body Weight Data Animal Number and Sex Individual Body Weights (Kilograms) Test Day Number: . 0 7 14 20 Observation Day Number 14 1-M 2-F 2.31 2.50 2.35 2.63 2.49 2.68 2.57 2.80 2.74 2.96 1-M 2-F 2.41 2.72 2.53 2.85 2.60 2.89 2.69 2.95 .2.82 3.16 1-M 2-F 2.10 2.16 1.70 1.96 (1.36) 1.96 2.04 2.18 1-M .2-F 2.55 2.72 2.35 2.80 2.38 2.26 2.18 1.84 2.04 (1.80) 1-M 2-F 2.61 2.53 (2.01) (1.87) - - - Note: Figures in parentheses represent autopsy weights. I 009V*7 r t WATER PCB-SD0000030709 ( (8 B. Mortality The mortality data are summarized in Table m. The data in dicate the number dead and the total number of doses received prior to death. The female of Group m died on the second day of the 14-day observation period. ' The mortality data were statistically analyzed employing the technique of Litchfield and Wilcoxon*, and the mean subacute percu taneous lethal dose (LD^q) was calculated together with the LDq>0j and the LDg^g^* For the purpose of these calculations, the teat per iod was assumed to be 34 days, i.e. a 20-day application period plus a two-week observation period. All animals that died during the inter val, at each dose level, were used in establishing the per cent mor tality for the dose level. The data are presented in Table IV and are graphically illustrated in Figure 1. * J. T. Litchfield, Jr. and F. Wllcoxon, "A Simplified Method of Evaluating Dose-Effect Experiments", J. Pharm. 2c Exp. Ther., Vol. 96, No. 2; pp. 99-113 (1949). NEW 009128 WATER PCB-SD0000030710 ( (9 Group T-C T-I T-n T-m T-IV TABLE m Summary of Mortality Data Number of Consecutive Number Number Doses Received Tested Dead Sex Prior to Death 2 0- - 2 0- - 2 1F 13 2 1M 20 2 1M 1F 5 6 NEV 009129 WATER PCB-SD0000030711 < ( 10 TABLE IV ' Statistical Analysis of Mortality Data Dose mg/kg/day Number Dead Number Tested Observed Per Cent Dead Expected Per Cent Dead 10 0 2 0 0.2 50 1 2 50 26.5 100 1 2 50 66.0 . 200 2 2 100 . 92.5 Mean Subacute Dermal Lethal Dose (LDjq) * 75 mg/kg/day* 95 Per Cent Confidence Limits of LD50 " 30 to 190 mg/kg/day Subacute Dermal LDo.01 "6.2 mg/kg/day Subacute Dermal LD99.99 e 915 mg/kg/day ; I * The LD50 specifically refers to the dose which, administered . each day over a 20-day period, would be expected to kill 50 per cent of the animals so dosed within the 20-day dosing period plus an additional 14 days during which the test material is not applied. NEV 009130 WATER PCB-SD0000030712 &*> U I* 6 Q t \ i 1 i \ i I1 ) NV 009131 WATER PCB-SD0000030713 11 C. Reactions ' 1. Behavioral Reactions No untoward behavioral reactions were noted among the animals receiving doses of 10 mg/kg/day. Generalized inactivity, weakness, lassitude, and loss of appetite were exhibited by rabbits receiving doses of 50, 100, and 200 mg/kg/day respectively. These symptoms progressed in intensity and ev entually ended in profound prostration in moribund rabbits in these dose groups. The surviving female in the 50 mg/kg/day dose group appeared normal by the end of the 14-day observation period. However, the sur viving male in the 100 mg/kg/day dose group continued to show slight gen eralized inactivity and slight loss of appetite at the end of the observation period. 2. 'Local Skin Reactions . The skin at the application site of animals in the 10, 50, and 100 mg/kg/day dose groups became scaly after the eleventh application of the test material. Slight wrinkling and fissuring were noted after eighteen applications. During the 14-day observation period, desquamation occurred and by the end of this period the skin of the surviving animals appeared nor mal. Animals receiving doses of 200 mg/kg/day died before any signifi cant untoward local skin reactions could be noted. N6V 009132 WATER PCB-SD0000030714 (( 12 D. Hematologic Studies and Urine Analyses Values for hematologic data and the results of urine analyses, obtained on surviving test animals at the completion of the dosing period did not differ significantly either from those recorded initially or from the corresponding data secured from the treated control animals. E. Pathology 1. Gross Pathologic Findings Complete necropsies were performed upon all animals that died during the teat and upon all animals sacrificed at the end of the ob servation period. No significant gross pathologic findings were noted among animals in the 10 mg/kg/day dose group. Animals receiving doses of 50, 100 and 200 mg/kg/day respectively showed liver discoloration (lighter in color) but otherwise no hepatic alterations were seen upon gross examination. All other tissues and organs examined from these ani mals were comparable to those of treated control animals. 2. Microscopic Pathologic Findings . Microscopic examinations were.conducted upon the tissues and-organs taken from all sacrificed animals. Animals that died during the test exhibited advanced post-mortem changes. Consequently their tissues and organs were not examined microscopically. No significant histopathologic alterations were noted in any of the tissues and organs of animals receiving doses of 10 mg/kg/day. NbV 009133 WATER PCB-SD0000030715 (( 13 Moderate hepatic necrosis was noted among surviving rabbits in the 50 and 100 mg/kg/day dose groups. This hepatic change was consid ered to be a direct effect of percutaneous absorption of the test mater ial. All other tissues and organs examined from these animals were comparable to those of the treated control animals. ' NfcV 009134 WATER PCB-SD0000030716 (. ( 14 IV. Summary A. Body Weights Examination of the body weight data revealed moderate to sev ere adverse effects among rabbits of Test Groups XI, in, and IV (50, 100 and 200 mg/kg/day respectively). The effects were especially pro nounced in the moribund animals of these groups. The body weight data of animals receiving 10 mg/kg/day compared well with that of the treated con trol group. . B. Mortality and Reactions The 20-day subacute dermal mean lethal dose (LD^q) of Aroclor 1254 for the albino rabbit was found to be 75 mg/kg/day. Generalized inactivity, weakness, lassitude, and loss of appetite were exhibited by rabbits receiving doses of 50, 100 and 200 mg/kg/day respectively. No untoward reactions.were noted among animals in the 10 mg/kg/day dose group. Local skin reactions included scaling, wrinkling, and fissuring among animals receiving doses of 10, 50, and 100 mg/kg/day respectively. These reactions occurred during the second half of the 20-day test period. During the 14-day observation period desquamation occurred and by the end of this period the skin of the surviving animals appeared normal. Animals receiving doses of 200 mg/kg/day died before any significant lo cal skin reactions could be noted. NfcV 009135 WATER PCB-SD0000030717 (< 15 C. Hematologic Studies and Urine Analyses Hematologic data and results of the urine analyses for test group animals compared well with those of the treated control group animals. - k. NEV 009136 WATER PCB-SD0000030718 i (( 16 iI D. Pathology 1. Grogs Pathologic Findings ` . No significant gross pathologic findings were noted among the 10 mg/kg/day dose group. Animals receiving doses of 50, 100, and 200 mg/kg/day respectively revealed liver discoloration (lighter in color). All other tissues and organs examined from these animals were comparable to those of treated control animals. 2. Microscopic Pathologic Findings No significant histopathologic tissue alterations were not ed among animals receiving doses of 10 mg/kg/day. The surviving ani mals receiving doses of 50 amd 100 mg/kg/day respectively revealed moderate hepatic necrosis. This finding was considered to be a direct . effect of percutaneous absorption of the test material. All other tissues and organs examined from these animals were comparable to those seen in treated control animals. Respectfully submitted, , INDUSTRIAL BIO-TEST LABORATORIES, INC. Report prepared by: Richard J. Palizzolo, B.S. Departmental Director Acute Toxicity Department Report approved by: Associate Director March 29, 1963 NEV 009137 I WATER PCB-SD0000030719 October 11, I937 prolonged exposure to Aroolor wapors I wolfed at high ttaper*tursa or by repeated oral ingestion will load to systenlo toxio sffsots, > Repeated bodily oontaat with tho liquid ri Aroolor9 may lead to an aena~fom skin eruption. Suitable draft rant HatIon to control tho ' vapors evolved at elevated temperatures, *v as well at protection by evitable garments " fro extensive bodily eontaot with tbs liq= old. Aroolors, should prevent any untoward ^offset.* In talking with Dr. Kelly before these three paragraphs were written, agreed that they night as well be phrased ee that they eeuld be used not only in the Aroeler booklet, but quoted In oorrespeadenee as that nay be naosssary* L.A. watt nev U7332 WATER PCB-SD0000030720