Document ympgzRnn8YxGDdZG0Lm91Zr73

THE TOXICITY OF POLYCHLORINATED POLYCYCLIC COMPOUNDS AND RELATED CHEMICALS Author Rennie D. Kimbrough* Center Tor Diaease Control Atlanta. G*. Rtltntt; M- M. 8inn j. n. Gr*t| Medical Rwarch Council M nc Tockotogr Unit Madkat Xnuttli Council Litmlariu CanhakrM, Sumy Gotland INTRODUCTION ' Only the small group of the polychlorinated | polycyclic compounds given in Figures I and 2 I will t>e reviewed in (Ilia article. Several years ago when this laboratory was the Atlanta Toxicology Orjncli of the Pood and Drng Adnunisiraiion. enneem was voiced over the possible long-term cficcts of jndy chlorinated bi|>hcnylt. and wc began to study the toxicity of Aroclor 1254 and Aroclor I2f.0. About this time a follow-up study was performed on a group of workers in New Jersey who were engaged in the production of 2,4^-T.' These workers had very severe chlorscne and had previously been shown to have porphyria.1 During that same lime, a petition was filed with the Food and Drug Administration for a residue level of 1 hcxachlorophenc on certain food crops. HexachJoiaphene, as well as 2,4,5-T, it made from tuchJorophenoi. although the cher deal reactions ire quite different. When we checked the available literature on the toxicity of hexachlorophene we felt that adequate toxicity data by present standards were not available, particularly not if the compound was meant to be Ingested. Further more. Larson1 had pointed out that hexachloro phene was more toxic than commonly acknowl edged. Hexachlorophene and similar chemicals behave differently from such polycyclic polychlorinated compounds as Ihe chlorinated biphenyls (PCBs). Hexachlorophene in particular is aculely much more toxic than the PCBs and is fairly rapidly eliminated from the body, while the PCBs have a very low acute toxicity, but are very poorly metabolized, particularly those with more than four chlorine aloma on the phenyl rinp. The long-term efTeels are therefore more critical for compounds such as PCBs. The chlorinated dibenzodioxtns and furans, some of which are extremely luxic on an acme basis, may also show persistence and their long-term effects may be equally as important. Then observations show that it would be erroneous to make predictions on long-term effects from results obtained in acute foxicily studies. formtriy with the bimronmeatii Protection Agwwy. Chombiee Toxieotegy Labotatory, 4770 beford Highway, rhimhtee, tie JO>41 Juwrs 1974 445 mm HONS 207653 FIGURE I. The malerhli shown In (life Optra an mtxtaret of compound! in which mvetel but anally not (I *f the potllloni f*X) ere chlorine and the ml are hydrogen. The propertied of Am materiel! depend on the degree of chlorination of the mixture. At ate inaohtble in water but eotubie in vtiyiwdtfteei in organic aolvnnli and In fata. The trade name Tor the U.S. product (Momento) of the chlorinated biphenyl* and ttiphenyh la Arodor The trade name it followed by the number. For the biphenyk, the drat 2 digit! of (hit number are 12, end for the terphanyla, 54 In each inatanoe the leu 2 digit!(hu the peroentate of chlorine at ucia Wheat bitta.Wheat t, f -tUthy Ira >111 (] .4 trlcMereeheeal) I4Wthyleae-b< ite-chlereeheml) aintiwi. t ,r .Tnt*-eu(e .a-eumenetunei) r< h j J U it t.t.t' trtchler*. (`hyery4t*eey1 ether (TrtclMtr treat** enaa) rrmao* t.l'.Thte*hfi(a.eh1ereehenel| 1,1'*e,*`*Tetreehlerei!Hey!enll1de (TCC> ),l.t'.IrlchlertartoaHief (TCP) FIGURE 2. The compound! in Ihla figure are white cryautUne eolldf with dearly daftnad mailing pointi above ItCfC They are practically buolubie in water and aolubia in rttyiqg amount! hi organic aoirenta and la fata. Thorn contain>n| an OH group, all the above exceptdg 5,4.4'<tricMorocarbealMde, are aolebie in aqueoutalulL 446 OtC Chtictf Rertnm At Toxicology ;u VI.1 Uil llj th If 1(1 If B E Ce HONS 207654 The present review it an attempt (0 correlate various ionic effect* pi udneed by I lie chemicals lined in Figures 1 and 2 in order in better understand and appreciate their behavior. Tire subject* covered in this article have lately enjoyed great interest and a rapidly growing number of scienlific reports arc available. Since a number of Ihe chemicals discussed produce similar or related effects, their toxicology is discussed together raliter than listing each compound sepa rately, which would have led to a great deal of tepetition. A detailed discussion of the various chemical reactions, analytical methods for the deterirhia* lion of the compounds, is not given. The review has been written with the toxic effects of the various compounds as the focal point and the liietaiute cited serves to illustrate these points. No attempt has been made to present a complete review of Ihe literature in this area. DEVELOPMENT. USAGE, AND OCCURRENCE A description of the synthesis of chlorinated biphenyls (PCBs) can be found in Liebig`s Artnalen as early as ISA I.4 Successful production of diphenyl in commercial quantities was not report ed until 1920 * At that time the chlorination of technical biphenyl was also described.4 Biphenyl ilseif is fungistatic and used as a preservative for citrus fruits ' Chlorinated biphenyls ate very stable, and have been used fur protective coalings, is plastic resins or gums for varnishes, and for waterproofing aild flameproofing of wood. The excellent dielectric properties of these compounds were noted as early as 1930. Until the sdvrnt of ihe PCBs. mineral oil was used as a cooling and dielectric liquid impregnant for high voltage electric apparatus such as transformers and capacitors. Since mineral oil is flammable and flimmsble gases could alto be produced, restric tions for Ihe use and installations of these trans formers had to be set, particularly for indoor installation. When mineral oils were replaced by he Aroclors, Ihe (ire and explosion hazards were eliminated. This was recognized by the National Board of Fire Underwriters and the National Electric Code was modified.* Use of Arodort also greatly reduced the physical size of Ihe cipaction.* A recent summary of the varied uses of PCBs has been provided by Broadhum.1" With peak production of 42,327 short tons of PCBs in 1970, Monsanto is the sole producer of Aroclors (U S tradename for PCBs) in the US. Yearly PCB production figures for the period 1960-1971 were reported by Monsanto.11 The production increas ed steadily until 1970 and dropped to 20,236 short tons in 1971 due to a voluntary restriction of the use of PCBs by Ihe company. Trade names in other coon met include Kannedor and San l ha term in Japan, Oophen, Fkndor, Plienoclot, and SovuJ in Europe. The polychlorinated capacitor liquids, com monly called eskareis, are mixtures of chlorinated biphenyls and chlorinated benzenes. A variety of other industrial applications'* have been found for PCBs. They have served n fluids for hydraulic, gas turbine, and vacuum pomps, as heal transfer fluids, as plasticizers, adhesives, textile coalinp, surface coalinp In paints and varnishes, as seal ants, and as part of formulations to prepare pressure sensitive records and color copying papers. The extent of Ihe current use of PCBs in printing processes Is unknown. Other suggested uses over the years have Induded catalyst carrier for polymerization of olefins, conversion of water permeable soil lo a nonpermeable slate, and combined inseclicide and bactericide formulations. Mixtures of chlorinated biphenyls and chlorinated naphthalenes have been employed to insulate electric wires and cables, particularly when they were used in the mining induslry and on warshipa.' * Residues of polychlorinated biphenyls have been reported in the environment by a number of authors hi different parts of the world. These reports were usually from areas where industrial pollution was responsible for the environmental contamination. Holden,11 after finding PCBs in marine fish snd mollusks from Scottish waters, noticed that they were generally highest in samples from the Firth of Clyde and he discoveied that the PCB contamination probably originated from industrial sewage The sewage sludge was trans ported from two Glasgow purification works by boats snd dumped In deep water six miles south of Garroch Head. The disposal of sewage sludge on land could lead to the introduction of PCB residues into the terrestrial ecosystem by transfer through food chains. So far PCB residues have not been detected in fresh water fish tn Scotland. Polychlorinated biphenyls hive also been found January 1974 447 > r . y 9 HONS 207655 in wildlife samples collected along I lie Dutch emu and (lie Rhine River14 and in fall and ml in different parts or (he Baltic Sea along the Swediah coast11 Riscbiough ri at.14 compared ilie presence of PCBs with that of DDT and its metabolites in various birds. These authors found very low concern rations of PCBs in wildlife species in Baja, California, Mexico, and the Gulf of California, wlurii arc very remote areas with a high ratio of DDT to PCB of approximately 9 or 10. In sea birds fiotn (he Pacific, the ratio was between 5 and 10 and at that lime, PCBs were not found in eggs of the Adelre penguin from Cape Croller, Antarctica. Relatively high PCB concentrelioru were found In San Francisco Bay, Puget Sound, and San Diego Bay and the ratio of DDT lo PCBs in wildlife samples from this area was between I and 2. This is again emphasized by the findings of Dustman et aJ.17 Twelve Alaskan bald eagle egp had median PCB residues of 1.65 ppm while 11 eggs collected in Maine, Michipn, Minnesota, and Florida showed a median concentration of 9.7 ppm. Ziiko1 * found low concentrations of PCBs in differenl types of fish taken from aeveral lakes and the St. John River System, New Bruntwick, Canada, and the Nova Scotia banks. Fish from the Milwaukee River and Lake Michigan in the midwen part of the U.S., where the river enters into the lake, contained varying amounts of PCBa1* while fish taken from various small lakes in Wisconsin without industrial discharge failed to show PCBs. Large predators from Lake Michipn contained at much as 10 to 25 ppm PCBs on a fresh weight basis expressed as Arodor 1254 and salmon eggs contained as much as 12 to 17 ppm PCBs on a fresh weight basis. Further studies by Veith1* of the concentration of PCBs in differenl rivers emptying into Green Bay Indicated a trend towards a decrease in PCB concentrations since the roiuruaty pertial ban on PCB tales by Monsanto was imple mented in September of 1970. If PCB discharges arc very high a fishkill may result such as the one in Escambia Bay, Florida in I969.11 In that particular episode the PCBs had entered a plant's effluent through accidental leakap of heat exchanp fluid. The PCB found in this case resembled Aroclor 1254 moat closely. The peak amount found in water of the Eacambia Rivet at the outfall amounted to 275 ppb and the peak concentration found in oysters from Eacambia Bay was 3.0 ppm, Arocloe residues in sediment samples taken near the outfall reached 486 ppm. When the leakage from the plan! w,is corrected, Arodor levels gradually decreased tn this area. The wildlife portion of the U.S. National Monitoring Program analyzed slarhngs, mallard and black ducks, baJd eagles, and Ttsh of several species for PCBs. According to Shekel,11 monitor ing for terphenyls is also done but not for any of the other chlorinated compounds in this repot i. Research surveys that Include fisheating birds, insectivorous birds, ospreys, woodcocks, and cer tain ducks alio monitor only for PCBs and for pesticides. Oysters (Otaouna vlrgintca) are a good indicator of PCB and other chlorinated hydrocarbon concentrations in water and a nation al monitoring program in these moUusks in 15 comtaj areas has been developed.11 So far. moDuiks have only very occasionally contained low levels of PCBa. In summary, the PCB concentrations in water are low or nonexistent in remote lakes and fresh water streams and can be as high aa 50 jigfl (5 ppb) or higher In polluted rivers such as the Hudson River.14 The PCB concentration is usually higher In sediment from these polluted waters because PCBs art only very slightly soluble in water11 Since PCBs are stored in living matter, they jic concentrated in the food chain and the usual "biomagnifleation" is encountered as we know n from other poorly metabolized and excreted sttl> stances. The concentration of PCBs in fish, par ticularly those in the upper end of the food chain from polluted rivets and lakes, is usually estimated In the ppm range and flsheating birds will have somewhat higher levels of PCBs than the fish from the same geographic location.1* Young aninuis may contain lest PCBa than their older counter parts of the unw tptcitt In pifttcutaf lociiion- PCBs are also infrequently encountered in dairy products, meat, flsh, and poultry in ready-to-eat human food.171,1 Occasionally, human food and domeilic animal feed have been more extensively contaminated with PCBs. In these instances, the contamination could usually be traced to local misuse of transformer fluid, leakage of PCBs from heat exchange fluid, or contamination of recycled paper with PCBa where the peper had been used for food packaging such as cereaJ boxes.1* t he US. Food and Drug Administration has recently set temporary tolerances for PCBa in certain food products (see Table l). 44$ OTC Critical Xfnntt n Toxicology * MO NS 207656 TAHl.1- I Tnporiry Tnlium< u FmIi p*r Million (ppm) btuMiihtd bp U.S. Food and Dra| AimaiMnliai (]) Milk (lai biim.) (3) Dm) p'odiM It tin bawl) (3) Poultry (in bam) ill l-ppt (5) rotnplou.' and fmi.tln-d animal frcdi II.I Anmiiil 1'ivd ruwpctni-nit (7) I uli and slirllfuh (edtbli' porlmn) (S) thfani and iuiian food (9) Pal'd lnud-packapinp nultfial 25 2.5 5.0 05 0.2 2.0 5.0 0.2 10.0 himc: Tim lolcrjiiut Cm papet rood-packaging malarial iliall nol apply to aacli material aeparaied from ihe laria'cd food by a functional barrier impermeable to PTH micfjlion, Tim I'ctnd and Drug Admintilranon mil provide upon itqaoi the analytical methodi ii will uae for enforcing the temporary toietancci. mixtures of oriho-, rnela , and paraicrphenyli. Clilorinued lerphenyls ire produced by Monsanto and termed Arodor, followed by numbers. The first 2 digits of Ihese numbers are 54 and the last 2 indicate the percent of chlorine such as Arodor $460. Arodor aeries 25 and 44 consist of mixtures of poiychlo tins led biphenyls tnd polychlorinated let phenyls.** According to Zitko et si.,'7 poly chlorinated lerphenyls have teen found in environ mental samples. Polychlorinated terphenyls do not elute from GLC columns under conditions used for PCBs,** which my explain why they have only been detected occasionally, but the way rimy tre used probably also precludes their extensive spread in the environment. Chlorinated dibemodioxins nd chlorinated dibeniofunnt my occur m contaminants of cydic chlorinated hydrocarbons. In 1959 Tomiti et al,* reported on the AH of these aspects of PCBs in Ihe environment synthesis of polyhilodibeneo-p-dioxifu According I hive been the subject of number of reports, to these authors, chlorophenoh, when heated to reviews,*0 and conferences.*`"* * about 200*C or mote under alkaline conditions Terphenyls and chlorinated lerphenyls are used and pressure, will form chlorodibenzodioxins, as ai reador coolants. The lerphenyls consist of shown below * .o ns n, 1 t `n Ns o' * 2 NaO Undo similar tondnious a clilorudibcncofuran may also be formed. n i c n .5 n tuo ^ r- HiO NaO i. y Ctilonnaied dibcncodioxms and chlorinated diben- show a great deal of difference in the type and Ll wfurans are formed, or can be formed, as contam mount of contamination and different lots of the d inants in the manufacture of a variety of products tame product may vary in composition, making iy which use chlorophenoh and chlorobenzenes as toxicological evaluations of these products very ic iiamng materials, such as 2,4,5-T, penladiloro- difficult. Rigorous production control and cleanup a] pticnol. miTofen, hexa- and pentachlorobenzene, procedures need to be implemented so that the m rd in- and tetrachlorophenol. Polychlorinated composition of different lots manufactured by Ihe :d biphenyls may also be contaminated with chlor same company becomes mote uniform. id inated dibenzofursns.40'4 1 and chlorinated diben- Villanuevi el al.4* and Jensen and Renberg44 sofursnt may resoli from polychlorinated reported the pretence of chlorinated dibenzo-p- `y biphenyls under UV irradiation.4* The toxicity of dioxin in peniachlorophenol. Hydroxy- mJ these products varies depending on the position nonachlorodiphenyl ether was rise found and id number of chlorines attached to the phenyl referred to at "pmdioxin" under the ammption nnp Products from different manufacturen may that it represented a precursor of dioxin. JttMTT 1'?4 441 MONS 207657 According lo iidinnm Cl jI..*' Hic predom inant dioxin in a commercial pentscliiorophcnol was oclachlurodibenzodioxin while trace amount of hexichlorodibenzodioxin and no lelrachiorodibcnzodioxin were round. Since ihc technical pentacliloriiplicnol wai in tonic retpeclt more toxic in animal itudiei, than the pure product, the production proceu of pentachiorophenol was clungrd to minimize the dioxin concentration and a new product has now been re|islered under the Federal Insecticide Fungicide end Rodenlicide Act with a lower dioxin content. Technical 2,4,5-T has also been found to be contaminated with dioxin. In this intlince, unfortunately, the very toxic 2,3,7,8 lefrachlorodibenzodioxin,44*4* was present. The contami nation of the 2,4,5-T with 2,3,7,0-tetrichlorodibenzodioxin ranged from < I ppm to as much as 32 ppm over the years In newly manufactured 2,4,5-T in the U.S. the content of tetrachlorodibcu/.o-p-dioxin lias been substantially reduced and is usually kept below 0.| ppm. Whether other dioxins or furans have ever been detected in 2,4,5-T in the U.S. has not been reported in the literature. Cldotinatcd dioxins luve also been found in so-called "toxic fat," winch produces chick edema disease when fed to chickens. After several yean of study by industry and the Food and Drug Administration, Flick et at.4* were able to separate a purified crystalline product which produced chick edema disease. Two yean later, Cantrell et al.s* announced thal 1,2,3,7,8,9chlorodibcnao-p-dioxin was one of the toxic compounds that produced chick edema. The origin of the chlorinated dibenxodioxin in "toxic fat" was not clear. It was suspected thal il aroae from chiorophenols which were used lo preserve hides for the manufacture of leather. TalJows and greases were obtained from these hides" and a more recent outbreak of chick edema disease was traced io the contamination with chiorophenols of aotpstock used as feed fats. Higginbotham el ai.*3 showed that chiorophenols and their tails, when healed, undergo condensation reactions and form chlorinated derivatives of dibenzo-p-dioxm; furthermore, chlorinated dioxins and predioxins were originally present in some of the chlorinated phenols. Generally no efforts have been made to determine whether various chlorinateddioxins and /imns are present in the environment. However, Baughman and Meselson* * recently reported the presence of dioxins tn fish taken from four locations in Vietnam in Military Region III, which Includes Saigon and aeveral provinces to the north.14 The concentrations ranged from IA to 814 ppt. Zitko*1 was not able to demonstrate chlwfatated dibenzodioxlns and dlbenzofurans m aquatic animats from various locations in Canada. Chlorinated naphthalenes may so far have remained unidentified since they interfere with other chlorinated compounds in any muliiresidue analysis and are delected only when mass spec tronwtry Is also performed.** The various gtrmhhiB gven in Figure 2 have been used as antiseptics, deodorants, and particu larly, as far as hexechlorophene ta concerned, also as preservatives for various cosmetics, soaps, and lotions, A listing of djlTereni products that contained hexachiorophcne prior to the new regu lations was given by the American fturmiceulical Association *1 Registration of bifhlonol was cancelled in the U.S.** because it produced photosensitivity reactions in humans using soaps that contained this product. None of the poly chlorinated polycyclic compounds used at germi cides have been thowr, to accumulate in the environment. Hexachlorophene has additional uses f a fungicide on ornamental plants, on conveyor chains as a mildcwstat. In laundry rinses as s mil dews* si, In shoes and air filters and assorted industrial uses as a fungiilal.and on leather, paper, and textiles.** It is also registered as a pesticide for use on cucumbers, peppers, and tomatoes 40 and for cotton. Dichlorophene Is used as a mildew preventive on cotton and is recommended as a fungicide end bactericide for the protection of textiles and materials from molds.41 CHEMISTRY: METHODS FOR CHEMICAL DETERMINATION AND PHOTOLYSIS Much of the chemistry of polychlorinated biphenyls has recently been reviewed.3 304 Commercial chlorinated biphenyls ,re mixtures of biphenyl groups with diffen .it numbers of chlorine atoms (Figure 1). When hydrogen atoms are replaced by chlorine on the biphenyl ring, a large number of substitution combinations can arise since the number of chlorine atoms on the molecules can vary and the chlorines can alto be 490 CXC Critical heriews In Toxkoiaty MGNS 207*58 ,inched in ilif molecule in different positions, retailing m a variety of isomers. Even If the chlorination may be the tame, different commer cial hatches nuy very in the reliitve concentration of isomers. The PCBs produced in the U.S. by Monsanto fall under the trade name Aroclor. The nnous Aroclor mixtures have a (real number of constituents, and when the biphenyl is chkorinucd, 210 different chemicals are theoretically possible. Aroclor 1254, fur insiance, yielded 69 constituents while the fractionation of Aroclor 1242 and 1260 yielded 4} and-7B components, respectively.*4 The chlorinated biphenyls (PCBs) are chemi cally very inert, resistant to corrosive chemicals, insoluble in water, and have a low vapor pressure. Aroclor 1221, for instance, has a boiling point of 278<>C. The boiling point increases with increasing chlorination. The chlorinated biphenyls are deter mined in biolopcal and environmental samples by s combination of electron capture gas liquid chromatography and mass spectrometry, Nuclear mignciic resonance spectroscopy Is used in addition to the above two procedures in some laboratories PCBs were first identified in environ mental samples by Jensen in 1966.** They were detected as tnieifenng peaks on GLC analysis of environmental samples tlut were analyzed for chlorinated pesticide residues4* The PCBs interfered with routine pesticide snalyscs for chlorinated hydrocarbons. Various authors have therefore published separation methods44 and Reynolds9 * discussed this exten sively in a review. According lo Safe and Hutzinger,4* the use of mass spectrometry for structural studies of PCBs Is lim ad because the chlorine atoms may be ran domized between the phenyl groups when the PCBs are fragmented by electrons. The only exception lo this is 2,2'-dichiorobiphenyl. The quantitation of polychlorinated biphenyl in environmental samples has met with difficulty since the PCBs represent mixtures. Several workers hive compered PC8 components in field samples to Aroclor 1254 because the sample chroma tograms were moat similar to these mixtures.44'4* Rote and Murphy4* quantitated individual peaks of different Arodors by means of s response curve. The semiiogarithmic relationship of detector response (total peak area/16 ng) to average chlorine content was obtained for each Aroclor by the method of least squares From (his relationship the theoretical response of the detector to each chlorinated biphenyl was calcu lated. When this approach was used, the levels of PCBs were lower than when the PCB levels were compered to the Aroclor 1254 standard. Because of these difficulties in quinUtetion, the PCB levels reported from different laboratories may vary greatly. The various problems encountered in the analytical methodology of polychlorinated biphenyls and their quantitation are also described in a U.S. FDA report which can be obtained upon request from the U.S. Food and Drug Adminis tration in Rockville, Md. The title of the report is Analytical Methodology for Polychlorinated Biphenyls (Feb. 1973). Polychlorinated terphenyk (Figure I) are not eluted from ges chromatography columns under conditions used for polychlorinated biphenyls.** By altering their pi chromatograph column, Ziiko et al.*7 were able to detect polychlorinated terphenyk. Chlorinated naphthelenet (Figure I) elute at the same time and cause interference with the determination of chlorinated blphenyk and chlori nated hydrocarbon pesticides in general, unless mass spectrometry k performed to establish the identity of the various chemicak involved.44 The methods presently available for the deter mination of trichhrocarbenUide and other germi cides (Figure 2) are not very well developed. Graber at al.4* outlined thin layer chroma tography method for the determination of these chemicals in soap and very briefly mentioned other methods that have been employed in the past Several methods fos the determination of chlor inated dibenxodioxins and chlorinated dibentofitrans have been described The goal of these methods k two-fold. First of all, quite number of technical polychlorinated organic compounds may contain one or several of these compounds74'71 as contaminants and methods hive been developed to determine them in various technical products. Other methods dealt primarily with toxic fats where many attempts were made to isolate and identify the compounds reiponrible for chick edema disease." Some of these methrjs were described in the paper* that will t cited in connection with the description of chick edema disease in this article. When preparing the various chlorinated dlbenzodioxini and ttibenzofurani for toxicity studies in older lo obtain pure product*, method* alto need to be available for their icparaiion and identifi cation.'4 One of the problem* with letrachlorodibcnzo- dioxin a* well at let richlorodibenzofuran i* their aevere toxicity in the microgram range which will probably not retult in an accumulation within the ppm in biological tittuet, for inatance, for residue analyst*. Baughman and Mcselion1' have recently descrihcd a method for the determination of these material*, within the ppt range. /Jexachlomphene it made from trkhiorophenol. A process for its production was patented by Gump.'1 Additional methods for the produc tion and purification for hexachiorophene have been published and are died in Chemical Abstracts. Recently eeveral method*'*'7* have been developed for the determination of small amount* of hcxachlorophene in blood and biological tissue*. Some methods were based on extraction of liexachlnrophene with ether or ethanol or a mixture of ether and ethanol. An acetyl or methyl derivative was formed and the hexachlorophcne was determined with electron capture gas chroma tography.'* In an earlier method described by Bachnunn and Shctlar,1* simple benzene extraction was used and no derivative of hexachlorophcne was nude, which led to very variable results Tite various advantages and disadvantages of these methods were discussed by Ulsamer.'* In earlier work my co-workers used ether for extraction and prepared a dimethyl ether deriv ative of hcxachlorophene.11 Recovery with this method was only about 75% and we have recently altered our method of extraction to obtain belter recovery.,s A method developed by Kabacoffel al.'1 for the delermination of hcxachlorophene it, according to the eulhors, also suitable for deter mination of dichlorophene and Gutenmann and Uska> have described a method for the determin ation of hcxachlorophene in several agricultural products. The production of hcxachlorophene from trichJorophenol proceeds at low temperature and under acidic conditions which makes the produc tion of a chlorinated dibenzodioxin as an Impuri'/ highly unlikely. Since chemicals that are found in the environ ment are exposed lo sunli^il. the effect of W light on the compound* under discussion ha* been given considerable attention. Huatert and Korte1* synthcli/ed a few PCB isomer*. (miikI*. 2,4.(i,2'.4'l6'-hexachlorobiphenyl. 2.4.5.2'.4\(>'. hexachlorobiphenyl. 2.4,2',4',ieirat:hlofoiupliciiyl 2,5.2'.5'-teifachforobiphenyl, and iiradiaicd ilie various isomers with UV light. A mcicury vapor lamp with quartz filler* was used as a light source The UV spectrum of the synihetized chlorinated biphenyls only shows significant absorption below 300 nm. When these compounds were Iiradiaicd in a solvent such as hexane, acelune, methanol, o, a mixture of methanol and water, dechlorination and polymerization of the compounds was observed. Whan (he PCB iiomers were irradiated in perftuorinated dimelhylcyclohexanc. an inert medium, iaomarizalion. and chlorination dccliiur. button wet* observed There experiments indicate that, under environmental conditions, photolysis products may result with a lower chlorine content but small (mounts of higher chionneted biphenyls may alto occur in low concentrations. Safe ami Hulzinger*1 reported that the irradiation of 2,4,6,2',4',6'-hexaehiorobiphenyl at 310 nm resulted in compounds of lower chlorine ciintcin. such as di-. tri-, tetra-. and pentachlnrohiiiliciivl* In these investigations hexane and methanol used as solvents. It is possible that new com|vmnK arc formed by loss of chlorine. Rcafnngooon ,>J condensation may lead to chhmilnpliL-m !. m environmental samples that are not piescni n commercial PCB mixtures- The 24-hr irraili.<i:,n, <<t 3,3' ,4 ,4'-let richlorobiphenyl. 2 ,2* .o letrachlorobiphenyl, 2,2,.5.5'-tetrachlorobipluiru,t dissolved in hexane resulted in about 707, loss oi the starting material, while with 2.2',4,4'.; ;'hexachlorobiphenyl. only 3.8% remained and lev* than 1% of 2l2'|3,3',4,4',5l5'-ocltch)orobiplK'nyl remained. The irradiation of Aroclor 1254 in hexane resulted in a change of the composition of the Aroclor mixture. The ps chromatogram obtained from the irradiated Aroclor showed shorter retention limes than the standard Arpclor 1254. The irradiation of Aroclor 1254 in j dioxanc water mixture also containing sodium bicarbonate did not lead lo the formation of chlorodibenzofurans or hydroxychlorobiphcnyls The compounds that were observed corres|K>ndcd to PCB molecules lo which water was added and a more polar "carboxylic" acid fraclion. Irradiation of Aroclor as a thin film with a frequent addition of water led to newly formed hydroxylaicd compounds and a more polar "carbosylic'' fraction. The retujta obtained from irradiating *92 CRC Critical Rnim in ToxtmiotX MOMS 207660 ly. ./ ivl, I hi' roi ee. ted iiw I in *t ion vas t in trt torale /i\S eni tyis knd of nm UK >rfc. TIC lids nut in in i of s'. nyl t of ,5'leu nyj in i of am ved Jut ii um of ylt. (Jed ida ion ion led lie" ;inf PTUs therefore depended greatly nn lire solvents used and ihe pll of the solution in addition to the hglti source uaed and may resull in dechlorination, fiirnuiiiiit or polymer*. hydroxylalion. and carboxylic products.** Exposure of various chlorinated dibenzodioxins lo summer sunlight or fluorescent UV light with an intensity of roughly 100 uw/cm1 in the presence of organic solvents such as methanol led to rapid photolysis of 2,3.7,8-letrachloro- and 2,7-dichlorodtbenzodioxin. while 1,2,3.4,6,7,8,9-nctschlorodibenzo-p-dioxm produced a series of chlorinated dioxtnt with uniformly decreasing chlorine content. Some evi dence was also found that reactions other than reductive dechlorination occurred.** When 2,3,7,8-tetraclilorodibenzodioxin was spread on a glass plate in a methanol solution and the methtnol wss allowed to dry, quantitative recovery of the dioxin was possible even after 14 days of UV irradiation. Similar results were obtained when 2.3.7Jt-fcifjchiorodibcuzodioxin was dissolved in meilianol. pul on a glass plate, covered with toil, and irradiated.*'' Time results demonstrated that under favorable environmental conditions, when oigauic hydrogen donors are preient. phoiolysir will t.ikc place. Bare surfaces of soil, concrete, or waier arc not effective inducers of photolysis. Chlunnaicd diben/odfoxins could al limes prove io be quite persistent. Partial dachlorinalion may rlsn occur with chlorinated diphenyls or the higher chlorinated dibenzodioxins and may result in less chlorinated compounds. The possible long lasting presence of the diuxint is also emphasized by Kearney el al.,** who recovered 56 and 63% of originally applied leirichiorodibcnzodioxin after one year in Hagerstown and Lakeland, Md. aoils. Tel rvchtnrmaticyianiiide and similar compounds split off chlorine atoms when expoaed to sunlight. This is briefly mentioned in connec tion with photosensitivity reactions of tha akin. Shaffer el al.** irradiated hextchlofvphene In sbsolute ethanol with UV light and obtained a number of de chlorinated bisphenols result ing from loss of chlorine in Ihe ortho and para positions relative lo the hydroxy group, GENERAL TOXICITY AND EFFECT ON REPRODUCTION Diphenyl itself, without any chlorine atoms, is t fungistatic agent which ta used as preservative for citrus frutta, and it usually impregnated Into the wrapping paper. The protected citrus fruit contains a certain amount of biphenyl residue. In different countries the amount allowed as residue on citrus fruit varies between 70 to 110 ppm of biphenyl. Booth et !.* found a reversible nephrotoxic affect of biphenyl on the rat kidney when rats were fed 0.5% or 1% (10,000 ppm biphenyl in Ihe diet). None of these affects was observed on diets containing 0.1% (1000 ppm biphenyl). Deichmann et al.*1 allowed that pro longed exposure to air containing diphenyl bi high concern rations of 5,40, and 300 mg/m* produced liver and kidney injury as well as branchopulmoniry lesions In mice and rats. In general, diphenyl has always been considered to be relatively non toxic. In 1969, in Finland, a man who had been intensively expoaed lo diphenyl for 11 yean became ill end died.*1 Diphenyl impreg nated wrapping paper had been manufactured in Finland for about 15 years In the plant where the man was employed. The equipment used in the impregnating procera was cleaned S to 10 times a year with tri- or telrachloroelltyiene. The cause of death of the worker in this particular report was not satisfactorily explained and neither were the complaints of 31 other workers ingaged in the tame factory. Since chlorinated ethylene com pounds were tied for cleaning, it diouid be established whether organic solvents containing halogens could lead to the production of toxic substances when they are brought in contact with diphenyls. If this proves to be the esse, then care rfiould be taken that this situation it avoided. The KBt are alio not very toxic when given as a tingle or a few repealed dotes to birds and mammals. However, the juvenile pmk shrimp is very sensitive lo PCBt in water, while trout and eel fish can tolerate higher doees; insects are alto not very sensilive to PCBt. Pin fhh are not quite at sensitive to tingle dose, but will die when exposed for 14 lo 45 days to 5 ppb of A rodor 1254.** PCBs are of a low order of toxicity to rats and rabbits when given as a single dote. The acute oral t'-xlcily decreases in nls with increasing chkw -tie content, while this is not is obvious when the dermal toxicity is letted in rabbits.** TWs may be due to poorer absorption of the higher chlorinated compound*. In studies with adult Slwrmtn strain rata, we found that the oral LDj, for A rodor 1254 and 1260 waa somewhere in the January I9M 445 HONS 207661 1 range of 4 to 10 g. These two compounds proved to be more toxic in weanling ran The tingle oral U>5, for weanlings wis 1295 mg/kg, and 1315 mg/kg for Aroclor I2S4 and 1260, respectively, and the lowest lethal dotes were 1000 mg/kg for Aroclor 1254, and 1200 mg/kg for Aroclor 1260. The intravenous LD for Aroctor 1254 in adult female rats was 358 ntg/kg.** A number of short-term studies hive been reported where the PCBs were given to aninult over a period or one or two weeks When (he PCBi were given in Ihit fashion they were somewhat more toxic than when they were given in a tingle dose. These studies ire reviewed in greet detail by Nelson et al.*' and will not be reported here since (hey do not give great deal of additional information. Vos and Kocnian** studied the toxicity of three PCI! preparations in chickens, Phenoclor DB6, a French product, Oophcn AM), a German product, and Aroctor 1260, a product from Monsanto in the U.S. The birds were fed 400 ppm of PC Us in iltcir diets for 60 days; the Phenoclor and Oophcn were much more toxic titan Aroclor. Tltc birds fsd the two Kumpean samples died hei ween 12 and 60 days after onael or exposure. The findings si autopsy consisted of hydro pericardium, ascites, subcutaneous adema, and liver necrosis. Only 3 of 20 birds fed Aroclor 1260 died during this lime with hydropericardium. None of the Aroctor fed birds showed liver necrosis. With the help of miss spectrometrk and microcouInmetric analytes, the authors*' determined altar the (wo European simples were contaminatad with tetra- and penlachlorodibeiuofurans. As an additional contaminant, chlorinated naphthalenes were found. Dermal toxicity studies with the three com pounds, Ctophen, Phenoclor, and Aroclor 1260, in rabbits** after two days led to reddening of the akin where the material* were applied. The skin reaction war more pronounced after one week, desquamation oT the externa) epidermal layers or the skin was observe j. (be hair grew at a reduced rate, (he skin be ante thickened, and prominent transverse wrinkles developed. The test animals showed gradual lata of weight, and one or the rabbits given Phenoclor and three of those given Clophen died. Ail but one rabbli showed ftuormcence or the liver under UV light, and microscopic changes were also observed in the livers and Hie treated akin. Vos and Nolenboom-Ram*T compared the dermal toxicity of the isomer 2,4,5,2',4',5'. hexachlorobipheny! with that of a PCD mixtuic (Aroclor 1260} in rabbits. Their dermal application resulted in early microscopic akin lesions in the Aroclor group. The lesions in the 2,4,5.2'.4',5'hexschlorohiphenyl group appeared latei and were lest severe. Both experimental groups showed liver changes.The liver damage was more pronounced in the animals that had been exposed to hexachlnmbiphenyl than In ihoat that had been exposed m the Aroclor. In inhalation studies with Aroclor 1242 and 1254, rata, mice, rabbits, guinea pigs, and one cat were exposed to PCB vapors five days i week for several weeks. The concentration of Arocioi 1242 vapor was 1.90 to 8.63 iig/l and no ill effects were observed. The exposure or animals ro the vapor of Aroclor 1254 in the concentration of 5.4 or 1.5 pg/l produced enlarged livers in rats. An ituci current respiratory infection in some coni ml .is well as experimental tils made it difficult ninterpret the results.** Since It is now known ili.ii the chronic toxicity of PCBs is more iniporijiii parameter in establishing their effects ihan shunterm exposures, further studies with lower tw. f. over longer periods of time would give wic meaningful information. Fortunately, must of tire compounds under' discussion have low vapor pressure so that exposure by inhilaiion probably does not present greet problem. A number of reproduction studies in various species have been conducted with PCBs. usually, embryo toxicity was observed hut there were no malformations. Heath el al.** were unable in demonstrate an effect un npruductiou with dici ary levels of 25 and 50 ppm of Aroclor 12S4 in mallards and bobwhites. An increase in eggshell cracking or reduction in shell thickness was also not observed while DDE, which was used as a positive control, induced significant shell thinning and cracking and reduction in hitching successes in mallards but not in bobwhite. In field studies with brown pelican shells, ii was found (hat thinning could be besi correlated m DDE residues rather than dieldnn or PCBs. This finding indicates, according to Blus et al ,'0# the PCBs ire not responsible for eggshell thinning m moat birds following environmental exposure. Paakall**1 fad ringdoves 10 ppm Aroclor I2M 494 CWC Critical Arrti-wr tn Toxkvkmy HONS 207662 lUc I hr ' ',5'. Illtf IlKI the ere tver d n) nftv i to and , for re >r of i-s metil, as t to 11',;;! l.iol tori- I I I i i I | i | ! | i mm' i the :tpoi vjhly I j | rimis .tally, ic no Ic 10 diet54 in gshcll , also as a nmng cesses it was cd io This the ling in 1254 fin h moil ills He injected others with I fiO mg/kg iniuj'critoiscally for 4 days, and evaluated the aslu'd egplicll weights of these birds. Ik found no difference in eggs)tell thickness between the ex|icrimcnlal and Use control groups. In a pheasant reproduction iludy with Aroclor I2S4.101*101 a significantly greater difference in rise number of eggs that were pipped but not hatched was found in the group of hens that received 50 mg of PCBs weekly. Egg production and halchabtlny was lower In this group, (he survival of chickens that did hatch al six weeks of pr was reduced, and Ihe ones lliai did survive were significantly lighter than the controls. Aroclor 1242 at 10 ppm or 100 ppm and Aroclor 1254 at 100 ppm in the diets of chickens did reduce egg production and hatchabllity and caused thin eggslietls. while Aroclor 1242 at I ppm, Aroclor 1254 al 10 ppm, and Aroclor 1260 at 100 ppm did not affect reproduction in chickens.10* In a rat reproduction study, decreas ed survival of pups al a dietary level of 100 ppm of either Aroclor 1242 or 1254 was observed. The results obtained from the reproduction studies in birds particularly seem to Indicate that the lower chlorinated biphenyls affect reproduction more th.m the higher chlorinated biphenyls, and il is very important to determine wire I her Ihe lower cltlooiiaied biphenyls that (save been suggested as replacements for presently employed chlorinated biphenyls have an effect on reproduction. In another iji reproduction study** the lowest dose of Aroclor 1254 that affected reproduction was 20 ppnt (1.5 mg/kg/day) which manifested itself in a decreased numbsr of liners and less pups pet litter. Neither I ppm nor 5 ppm of Aroclor 1254 had an effect on reproduction in the Sherman strain rat; however, an exposure of the dams lo ihe Aroclors at 5 ppm or higher increased the liver-iobody weigh I ratios in weanling rats of both sexes. At I ppm Ihe increase in ifver weight was observed only in Fla and FN> weanling male rats. The dietary levels of 100. 20, and 5 ppm of Aroclor 1260 did not affect reproduction; how ever, the liver-to-hody weighs ratio of 21-day-old pups was increased at all exposure levels. Mink are highly susceptible to the toxic effects of PCBs. A daily diet/ y intake of 30 ppm resulted in death in about 6 months. Exposure to 5 ppm Aroclot 1254 in (he daily diet severely affected reproduction.1 09 Unfortunately, it la often very difficult to compare the various dietary levels (hat affect different species, since the food consumption may vary decisively from one animal species (o another and the daily food intake in g/kg body wt. is often not Included in the scientific report. Some mice strains, for instance, will b exposed to as much of a given substance that Is present in the diet at the concentration of 300 ppm as rats on a diet containing 600 ppm or more. With some species it is of course difficult or almost Impossible to measure the food consumption, but a greater effort m establishing these very basic facts would be helpful for (he Interpretation of toxic effect' :n different species. Nol very much is known at present about the toxicity of terphenyh or chbrinated terphenyh These materials have not been studied to any great extent. The few studies that are available only describe very specific effects such as microsomal enr.ymc induction In the liver and give no overall evaluation of their toxicity. The most toxic of Iht various chlorinated dibcniodioxint and dibetnvfUrum is 2,3,7.S-letrachJurodibcnzodluxin which, in the German litera ture. It sometimes referred to as 23^,7-tetrachlorodibenzodtoxin, Single oral LD* values are female rats1. 44,7 pg/kg; male rata: 22,5 pg/kg; male guinea pigs: 0.72 pg/kg: rabbits: < 30pg/kg: and dogs; between 30 and 300 Pg/kg.10* It is possible that young animals are more susceptible lo the toxic effects of this compound than older animals and LD(0 values should therefore also be determined in weanling animals. One of the characteristics of this compound was that il caused delayed death, aometimes as delayed as 40 days after a tingle dose. During the period of intoxica tion, the animals lost a great deal of weight.10* On the other hand, 2.0 g/kg of dichlorodibenzo-p-dloxm and 4.0 g/kg body weight octachlorodlbenzo-p-dioxln were nol lethal to rats. Hexa-, hepta-, and trichlorodibenzo-p-dioxm were more toxic.10* Schulz'01 In earlier experiments found that oral doses of 20 to 50 pg/kg/body weight of 2.3,7.8-ietrachlorodlbenzodioxin resulted in fatal liver necrosis In rabbits. More recently Milnes'01 reported thal 10Pg/kg was lethal to rabbits. The 2,3,7,8-tetrachlorodibenzodioxin has a variety of specific toxic effects. Whether these effects vary in different species is presently not well defined. With the interest in this and the other related compounds, more information on Jammy 1974 4S5 HONS 207663 their tnxk'ily probably will soon become available. The pniblcim inherent in studying these com pounds are the unavailability of some of (hem. the fact tliat they may not be completely pure, particularly (lie hex* and hepts- preparation*, and of course with respect lo the letrschlorodlbenzcrdioxin, its extreme toxicity, which makes it difficult tohandle. In contrast to PCBs. teratogenic aa well as felotoxic effects have been reported for 2.37,8telrachlorodiben/odioxin. Sparschu et al.1** gave 0. 0.0.1. 0.125, 0.5. 2.0, and 1.0 pg/kg/day of 2.1.7.ft-tetrachlorodiben/.o-p-dioxin to pregnant rati on days 6 to 15 of pregnancy. With the 0.03 fig/kg dosage level, in effect on the fetus was not observed, but doses ftotn 0125 Pg/kg on up resulted in fetal mortalily, early and late resorp tions. and fetal inlcslinal hemorrhage. Khera and Ruddick110 pve doses ranging from 16 to 0.125 gig/kg to Wistar rats on days 6 to 15 of pregnancy. Tltesc ant linn also observed pronounced fetotoxicity al all dosage levels except 0.125 pg/kg, Tlte affected fetuses showed cerebral and intestinal hemorrhage as well as subcutaneous edema at autopsy. The offspring of mothers given 0.5 and 1 pg/kg of Ihe tetrachlorodlbenzo-p-dioxin pined less weight during their suckling period and their survival to weaning was reduced. This could have been caused by the intrauterine effect of the dioxib or by excretion of Ihe dioxin in the milk of tlte dams given tlte material during prepiancy. Courtney and Moore111 observed cleft palates in 3 strains of mice given 3 P|/kg TCDD subcutane ously from days 6 to 15 of pregnancy and an increased incidence of kidney anomalies. Hydronephrotic kidneys were also produced in mouse pups that were nursed by dams treated with TCDD during prepancy or at time of parturition.'11 The 2,3,7 J-teirachlorodibenzo-p-dioxin is extremely toxic in the chick embryo assay." In (he rat, hexachlorodibenzodioxin caused fetotoxtc effects at I and I0pg/k| and teratogenic effects at doses of 100 (ig/kg.'** Harris et al."3 pve female rats daily dotes of 0.1, I, or 10 Mg/kg TCDD for 31 days. Rats given 10 Mg/kg lost weight and most of them were moribund within 3 weeks. Weight pm was abo leas at the I Mg/kg dietary level. The rats raceiving 0.1 jig/kg were not affected. In experiments with guinea pip, 9 of 10 animals died arter a single dose of 3 pg/kg TCDD. while guinea pip receiving a tingle dote of lMg/kf pined less weight. All guinea pip receiving 1 \ (ig/kg/week died wiriun 24 to 32 days ami a weekly dose of 0.2 fig/kg depressed their weight Of the various animal species tested, the guinea pig teemed to be most susceptible to the toxic effects of TCDD, Reduced thymus and spleen weights were obterved in rats that received a single dose >f 25 fi|/kg TCDD, The rats that died it the various dotage levels showed massive hemorrhages into the heart, liver, brain, adrenal glands, and gasirointesn. nal tract. Necrosis and ulceration of the glandular part of the stomach were observed. Occasionally orpnized thrombi were noted and the liver reveal ed necrosis of parenchymal cells. Sublethal doses of TCDD produced microscopic changes in livct. kidneys, and thyroid. In the kidneys the coikding tubules were primarily affected. Doses of 0.1 Mg/kg/dsy for 31 days, a single dose of 5 pg/kg. and multiple weekly dotes of 1.0 and 0.2 pg/kg did nol produce changes in rats discernible wnh the light microscope. Toxicity studies were conducted in our lahiiulory with dictthroplwiie.'14 Tlte oral LDS0 tor adult female rats when Ihe material was pve it in peanut oil by stomach tube was 1,660 mg/kg and in males it was 1.500 mg/kg. The only signs of toxicity observed in these rats were diarrhea and some depression. When 200 mg/kg was applied daily in propylene glycol to the skin of a elipi'tJ area of the back of male adull rats as a 12 5 solution, local irritation of Ihe skin with wmic ulceration resulted but there was no sysiori, toxicity. The acute intravenous LDj0 in aduii male rats was 16.8 mg/kg when it was given m j saline lecithin suspension. In a two generation rai reproduction study, a dietary level as high as 1.000 ppm (50 mg/kg/day) had no effect. Gross ami microscopic examination of (he various organ-, including the brains of the parents as well a3 ilnofTspring did not reveal any morphological change-, that could be related to dichlorophene expo'-mc These fmdinp indicated that dichlorophene not very toxic lo rats. The peal difference between the oral and the intravenous LD*(> suggested either poor absorption or rapid break down of the material in the gastrointestinal tract Martins*' lists the acute LDt for guinea pig-. ,is 1,250 mg/kg and for dop, 2,000 mg/kg. Rats led for 90 days on a diet containing 2,000 ppm showed no evidence of toxicity. The acute toxicity of Hexachlorophcnt is much pester and varies in different animal species and is (54 CltC Critit*l Sfrtfwi In Tonkohgy m HONS 207664 also dependent on the route of administration and die solvent m which it 11 given. Gump11' lisu the acute oral LD,n in mice as 168 mg/kg and cites LDio values reported by oilier invesljgilore rangmg rtom 80 mp/hp to 215 mg/kg. Dogs and sheep weie more susceptible to the toxic effects of hexachiorophene. The interest in the effect of hcxacldnropltene on sheep in particular stemmed Trom the fact that it is effective against liver flukct''* and advocated for its anthelmintic properties. This even led to the treatment of duldren with orally administered hexachiorophene in China."1 Intravenously administered hexachloropheiie m rats resulted in an LD*o value of 9.1 mg/kp amt in tahhiis of 8.5 mg/kg. More recently we found dial the oral LDS# (single dose) for aduli male rats was 66 mg/kg and foi adult female rats 56 mg/kg: for weanlings it was 120 mg/kp wl*cn the material was given in pcanui oil by stomach tube to Sherman strain rats. The acuic mnavenous LDi0 was 7.5 mg/kg in dull male rats.11* Nakaue cl a).11* reported sinulai oral LDje values In Wislar rats. The single oral LDja for sheep and cattle lies between 30 and 60 nip/kp.'30 Single dermal LD*o values have not been reported in the literature. We found in rats11* that single applications of 600 mg/kg Itexachloroplicnc in 95% eiliyl alcohol solution caused death in 1 of 10 rats. Dermal application of 24 and 48 mg/kg/day for 30 days in propylene glycol or in a detergent as a 3% solution caused ulceration of the skin and affected the white maitet of the central nervous system in at least half of the rats. A dose of 12 mg/kg/day as a 1.5% solution caused only mild erythema and desquamation of the skin of rats. No effect on the central nervous system was observed. Whether a single doae given in either propylene glycol or a detergent would be more (oxic than one given in ethyl alcohol was not established. The signs of acute hexachiorophene poisoning following a single oral dose varied with (he age of the rats.'1' Adult salt suffered from aevere depression and diarrhea, but ahowed few signs of leg weakness. Weanling*, on the other hand, developed posterior leg weakness as well as de pression. The leg weakness became more apparent in the adults when they were given repeated doses of hexachiorophene. Death was usually preceded by convulsions in rats dosed intravenously. In rals killed with hexachiorophene, rigor mortis developed very rapidly and was very pronounced. Body temperature elevation following hexachlorophene poisoning has also been recorded in rats.' ` * Both changes, the pronounced rigor morlisand the temperature elevation, may be related to the fact that hexachiorophene uncouples oxidative phos phorylation. Additional symptoms of hexachlorophene poisoning particularly in other species are discussed under neurotoxicity. Nakaue et al."' fed 400 ppm hexachlorophene (28.9 mg/kg/day) to Wislai nts. They developed teg weakness within 5 days, diarrhea, emaciation, and they died in 7 to 10 dayi. No deaths occurred In the rats that were fed 200 ppm (about |3 mg/kg/day) for 16 weeks. In a two generation rat reproduction study which we conducted in our Moratory, we found s reduced survive! In the offspring of the F, generation fed 100 ppm (11.8 to 5.5 mg/kg/day) in their diet for 54 end 166 days. This was sliltslically significant in the second breeding of the F| generation. In the second generation the litters were slightly smaller, but the survival to weaning was not significantly reduced.11* A dietary level of 20 ppm (2.3 to 1.1 mg/kg) did not influence reproduction. Thorpe111 reported that the oral administration of hexachiorophene to male rats as well is single doses of 25 and 50 mg/kg hexachiorophene to sheep produced de generation of spermatogenic cells; multinucieaied cells were observed within the tubules of rite testes. The doses given by Thorpe were higher thin the ones we used In the reproduction study, and microscopic examination of the testes of the rats used in our reproduction study did not reveal any abnormalities. Toxic dotes of hexachiorophene given to the dam during pregnancy may result in malformations of the offspring. Thus far, two studies have been reported In the literature; both studies were conducted in tits,1 ','111 It should be determined whether malformations can also be produced in other species and with lower dotes before any conclusion can be drawn from these findings. A number of human poisoning cases have been reported in the literature and some of these were fatal. Most of the poisoning cases in humans have either been caused by accidental ingestion, because a hexaehlorophene detergent solution was mistaken for milk of magnesia, or by the applica tion of hexaehlorophene to large areas of bunted or otherwise damaged skin.M>>l,4 Larson1 was itauarr 1*74 437 MONS 207665  Ik first to reaipui/c thi hunted patient! JrvolopoJ ciinttllsKMit and other toxic symptoms add icpeatnl exposure to hexachiorophene which was applied to hunted areas of Hie skin. The incidence nf convulsions in burned patients decreased when the use of hexachiorophene was discontinued 1 *1 Eleven cases have been reported'**'11 in wliidi a single dose of 3% hexaclilorophetie deter* gent solution had been taken orally. Gastro intestinal symptoms including nausea and vomiting were observed in ten of these cates: the eleventh case, a child who drank about 250 mg/kg hexaclilorophcne. became comatose and died. One additional death occurred in this group, however, this was niost likely due to other causes. A l7*day*old infant was accidentally given daily ora) doses of 37 mg/kg hexachiorophene for 7 days. On (he third day the child started to have looec stools. On the fourth day of hexachiorophene administra tion the child developed spasms of the extremities with twitching of the face, sucking movements of the lips, and lateral shaking movements of the head. It was also noted in the case report that the child refused to take the medicine, there was drooling and frequent vomiting, and it it not quite clear whether the child received 37 ntg/kg/day. The child survived. A week after the hexachioro phene suspension was discontinued, persistent spastic flexion of the fingers, hands, and arms was still observed. After a month tha hands wara slightly flexed at the wrists but were not spastic. Horizontal nystagmus was observed but cleared later. The child was discharged when It was two months and three weeks old. At this point K had shown great improvement end when checked at six months the child was healthy looking.11* PUapil mentioned In his discussion a number of other unpublished cases, some of which were reported to the National Clearing House for Poison Control Centers. Among these was tha case of an infant who by mistake got a spoonful of PhisoHex In each feeding bottle for approximately two weeks, when it suddenly died. Chung el al.'11 treated a total of 105 patlenti 5 to 15 yean old with hexachiorophene for chionorchitsis sinensis infestation. Sixty of these patients received 1 dose of 20 mg/kg, 32 received 2 doses of 20 mg/kg/day, 8 patients received 3 daily doses of 20 mg/kg, and 5 patients received 20 mg/kg body weight on the first day and 10 mg/kg on the second day. The toxicity reactions observed in these patients were mainly confined to the central nervous systam and to the gastn>intestinal tract. The group that received 3 dotes ut 20 mg/kg/day had the most severe reaction. One patient m this group became comatose nn the fourth day with loss of light reflex and positive pyramid al signs. Fundus examination of the eyes revealed mild papQIary edema. A low grade fever was also obsenod. Complete recovery occurred within three days. One half to two thirds oT the patients had diarrhea within I to 2 hr after medication and in some cases a marked depressive affect on the central nervous system was also observed. An additional case of poisoning after oral ingestion waa reported by Korlof and Winaien11* fat Scandinavia and two fairly recent caaet with fatal outcome occurred in the U S.1 Herter1 *1 was probably the first to report a human poisoning case caused by the dermal absorption of hexachiorophene. Because of a language barrier, the mother of a newborn infant applied 3% hexachiorophene as a lotion after the infant's bath without rinsing the baby. After four days, excoriation* appeared on the child's face and buttocks, the infant developed convubions. Ins fece twitched and hit extremities jerked. A roving nystagmus was present. This child recovered wiiliin 16 days after exposure to hexachiorophene had been discontinued. In addition to the two ceses already mentioned In which poisoning occurred because of oral Ingestion, Mulitck*1* also reported four cases Children who died following the treatment for burnt in two Instances and the treatment for severe ichthiotu with a 3% hexachiorophene bath. Larson1 reported six burned patients who developed central nervous system symptoms following the dermal exposure to hexachioro phene. All six patients recovered. Korlof and Wlntten'1* reported two cases of poisoning following the derma) application of hexachlornphtne. Both patients were fairly exiensivciy burned, and developed central and peripheral nerve damage as wall as tome kidney impairment. Lockhart111 mentioned six deaths that were reported to the FDA ceused by the topical application of hexachiorophene on burned patients. In ill of these cases the skin was damaged, probably enhancing absorption of hexaddorophene, and although skin does represent barrier, it it not impermeable - a fact we tend to forgat at times. Damaged or abnormal akin may be eSS CJtC O-Mcwl ArWewr Si Toxkohtf MONS 207666 I to Uiv f of Due (lit live ryes ever rred the Her HVC also iflCT and ten I i tri [Inal >f a Tint r I lie foul and his IVing vitlihad 3 lied oral cs for Tor )aih. who loms loroand ming lorolively hcial neni. were ipical imed WH hexaent a nd lo ay be more pcnneahlc. and ihc iktn of prematuics. iKnlmini. and children in general may have absorption characteristics other than those of adult human beings, another factor that it often neglected. Recently1,1 ,l>4 41 deaths due lo liexaclilorophcne poisoning have been reported m France. Tlie deaths occurred in infantt who were treated with a powder to which 6% hexachlorophene had been added by mistake. This powder was mainly applied in rite dtapet area where the skin became inflamed, this is well as the diaper which served as an occlusive dressing, probably en hanced absorption. These cites will not he made public until litigation problems have been settled. Little published information is available on llte toxicity of live germicides in Figure 3 aside ftom hexadilorophene and dichkmtphene. The acute oral LD,,, for triclosan (TCC) in mice and rats it bout 4.000 mg/kg.1,5 The acuie subcutaneous toxicity for rats was 14.700 ntg/kg and tlie acute dermal toxicity was about 9.3 g in rabbits. Concentrations of I lo 5% solutions caused eye irritation in rabbits. Daily oral doses of TCC up lo 1.000 mg/kg/day for 4 weeks did not produce an effect in rats. When 3.S and 5% suspensions in gum arable were applied dermaHy lo rats for 4 weeks, the animals did not gain as much weight. Un published material cited in the minutes of (lie OTC (Over the Counter Drug!) Panel listed a "no effect" level as I 70 mg/kg/day in rats and for a 90-day feeding trudy in rabbits, 138 mg/kg/day. In baboons a daily oral dote of 100 mg/kg/day did not produce a toxic reaction. The minutes also mentioned some work where lower doses showed an effect on rats and dogs. Since some of theee results are conflicting, further studies are indicated and should be performed with products used u ingredients in soaps; a chemical analysts of these products it necessary to determine whether im purities might be responsible for varying results. Chlornanitine it a result of healing during manufaclute could bt present it such an impurity. Of the anilines, according to Hamblin,* *4 only p- and trxhloroaniline may cause methemoglobinemia if absorbed through the skm- The o- and m-ch)oroaniline may cause liver and kidney damage. Tridosan itself is also absorbed through the skin and absorption is greater when applied to dameged akin. SPECIAL TOXIC EFFECTS Skin Reaction* Chloracne This occupational skin disease can be produced by a number of chemical compounds117 Poly chlorinated biphenyls as well as Ihe chlorinated naphthalene*, a few isomen of the chlorinated dlbenzodioxins. and chlorinated dibensofutan* etc all able to produce chloracne In a certain pro portion of the human population. Oily skin and large pores teem to predispose lo the disease while the opposite h true for smooth, lender akin and it Is possible that young children mj^tt be less susceptible because of the nature of their Ain. Chloracne has alto occurred In workers engaged in ihe production of 2,4,5-T1. During tha indus trial production of Irichlorophenol a Hidden rise in temperature within the reactor hat occasionally resulted In accidents. In one Intiinee this exo thermic reaction caused an explosion.'* When the rise in temperature occurred. 2,3,7,8-tetrachlorodibenzodioxin wet formed and severe out breaks of chloracne were observed among the workers.1 *t> The term chloracne was first used by Herxheimer in 1899,' ** who thought It wu caused by free chlorine generated in certain factories. Wauer in I9I81** and Teieky in 1927'* suggested ihe term "Pemakrankheit." Another term sometimes used is halogcnwaxacne. As the word chloracne implies, part of the lesion of this occupational skin disease resembles adolescent acne; however, it it generally more severe and the distribution of the lesions is not consistent with adolescent acne, although it may be superimpoeed on adolescent acne. Chloracne is described In the literature as consisting of the formation of comedones with or without cysts and pustules. The follicuiar orifices are filled with sebaceous and keratinous material. Melanosis and a secondary inflammatory reaction may alio exist. In addition to this description, skin changes occur that have been discussed leas frequently in the more recent literature. These skin changes are usually referred to as cable rash or cable itch. Cable rash hat been observed in workers who have insulated cables with various mixture* of chlorin ated naphthalenes and chlorinated biphenyls and in workers handling these cables.14 ' It has been observed also in feelories that make eondenser*.141,14* According to Braun,'4* a few of immry 1*74 43* HONS 207447 Hit cam with manifest chloracne flirted with changes in the skin of the face resembling photo sensitivity and the bearers of these lesions suffered severe puritus in the areas of the skin lesions. This lesion termed cable rash occurred predominantly on the cheeks but alto on the skin of the arms. The outcome of cable rash varied; it could either disappear or it could develop into full-blown chloracne. In addition lu the pustules and comedones of typical chloracne. brownish kerallnization of the skin was observed in some patients. Braun' 41 pointed out that the workers, particularly those with poor hygiene, transmitted the disease to their spouses and children through direct eontaet. Many of the occupational cases of chloracne that have been reported in the literature have been reviewed previously.111,144 Jones and Alden141 reported one of the first outbreaks of chloracne in a company lhai was engaged in l ha manufacture of chlorinated biphenyls. They described one typical case and mentioned 24 men working on the manufacture of chlorinated biphenyls; 23 had had an acneform eruption on the face and body. Particularly during the 1930s and 1940s large outbreaks of chloracne occurred in a number of factories. Much discussion has been devoted in the literature to the problem of whether the chloracne was produced by external contact alone or whether it was caused by systemic absorption, particularly the Inhalation of vapors. One question asked was whether internal administration could cause chloracne, and if so what part it played in any particular case As we know now from outbreaks of poisoning due to the consumption of nee oil that was contaminated with polychlor inated biphenyls in Japan {see under "Yueho"), chloracne can be produced by the systemic absorp tion of chlorinated biphenyls. The exparimenls by Shelley and Kligman'44 and Plewig141 showed that the skin lesions could alio be produced by applying the chloracnegenic compound to the skin. After occupational chloncne manifested itself, the patients also complained of systemic effects such at loss of appetite, nausea, edema of the face and hands, abdominal pain, vomiting, and burning and soreness of the eyes. Particularly with the chlorinated naphthalenes, hepetotoxic effects have also been observed. Application of chemicals to the akin can lead to their absorption and cause systemic toxicity. Whether a systemic toxic el leu results will depend on the skin surface atca covered, the amount of the toxic substance applied, and the ability of the skin In absorb ii. || dermal absorption is poor, a tingle apphe-unm will usually not be very toxic. However, if the c*upound is not easily metabolized and excreted, repeated dermal application may result in sulircienl accumulation in the body tu cause systemic toxicity. Crowe144 has pointed oui that ehhnacm' Jut to chlorinated naphthalenes has dec leased hciauscthese compounds have been used less since the Second World War. However, Weber in ivmt14* reported an outbreak of chloracne in a company angaged in the production of electric cods dial ware coaled with a mixture of chlorinated naph thalenes with the tradename "Nibrenwax " Detailed examination by teveral German investigalors indicated that the capacily of 2.4.5Irichlorophenol as well as 2,4,5-T{2.4.S-trrthlotophenoxyacetic acid)'4*1**4 to produce chhacne was due to con lamination with 2,3.7.R.ieirathlnrodibenzofuran or 2.3,7 .B-teltachlnrndibenzodioxin. Schwartz el al.l>> indicated lhai lltc must potent chloracne producing agents were ihc lImh tnafed naphthalenes, chlorodiphcnyls. and Jtlorodiphenylokides. Persons whu wurked with chlorinated naphthalenes usually developed ,io:e after a month or more of exposure. Lighl microscopic observations made of die chloracne lesions of humans varied somewha*. depending primadly on die length of mn; tlw lesions had existed at the time of examtmtuon The earliest change encountered consisted oi acsnthotic widening of the external root sheath ot the hair follicle. Above a layer of basal cells <. to 10 layers of large round cells were obvuived. followed by a layer of 2 to 3 cell rowsoT granular cells. In later skin biopsies, folding of the h.ivd layei occurred and comedo formation was observed, resulting in dilatation of the follicle and atrophy of the epithelium surrounding lire com edo. The contents of the follicles consisted predominantly of keratinous material radio than sebum. The sebaceous glands also undetwent changes. Within the alveoli of the glands the basophil cells increased in number and hasal ceil hyperplasia was noted. The ducts of the schai.eom glands were filled with solid mass of husoi-in: epithelial calls. The sebaceous glands gradually 4M CRC CHtktl XevtnM to fwiwlgp P "HONS 207668 dijjppearcd and in later aecretiuns were only be invesiigaied In chloracne paiicnis who manifest ft if, <>jmi/cd js utlid jtrntrtitioiis of the comedo hy per pigments lion- CJ wall- Aside town atrophy of I he aebiceoui glandi, In none of the human cate reports on chloracne cc liyicijitavui has also (teen described and many large have the mucous membranes been studied to any ir sebaceous glandi forming huge cysts (riled pri great extent. Vaginal smears in women and tpuia, iii marily with kcraiinous material have been corneal, and nasal scrapings may give helpful ii- observed. Hyperkcrainail and acanthosis of the information in this respect. Since In animals it, surrounding epidermis has usually accompanied exposed to polychlorinated polycyclic compound* fi- these lesions. Foreign body granulniniU which are low vitamin A levels have been observed In the lie thought to result front rupture of the follicular liver, it is possible lhat the compounds may affect cysts have been found in live skin.14 7,1 Sl In the vitamin A absorption and storage, and adequate Jt Japanese poisoning incident (Yutho), the predom utilization in the liver and elsewhare in the body. VC inant skin lesion was marked hyperkeratosis of the If the leiion it produced by inadequate utMisaUon lie u epidermis, cystic dilatation of the hair follicles, of vitamin A, then administration of vitamin A per snd an increase in the melanin pigment in the basal ae may not necessarily alter the course of the iy at cells of the epidermis. Foreign body granulomala were also observed. The contents of the cysts disease. Protein deficiency may also mimic hypovitaminosh A in that it affects transport of vitamin -li consisted partially of keratin. In a stillborn child A. The liver stores 90% of the available vitamin A lt- the hyperkeratosis was even more pronounced, and these stores may be afTected In liver disease. accompanied by atrophy of the epidermis; cystic The mlcroicopic appesrance of the akin lesions .5o dilation of hair follicles was observed, especially in resembles in some respects observations made in ne the skin of the head. According to earlier skin of patients suffering from vitamin A >r- reports' * ' some differences exisl in the micro deficiency. In vitamin A deficiency, hyperkeratosis li- scope changes observed in the skin following of the hair follicles is observed. Multiple firm exposure to the various chlorinated cyclic com papules may develop at a result of the keratin ut pounds that can produce the lesion. It is. of plugs in the sebaceous glands. Dryness and .ir- course, possible that these histopathologic vari tcaliness as well as furunculosis of the skin may ro- ations arc in reality only differences lhat exist also be present. In contrast, night blindness has between an early, a well-developed, and a not been reported ai a symptom In patients 111 nc subsiding lesion. - suffering from chloracne. Unfortunately, chlor Many theories have been advanced to explain acne outbreaks are often not well investigated and, he the development of chloracne. None is satis because Of problems with litigation, accounts are ul, factory. The concept of mechanical plugging of often only published many years after the out he the hair follicles on skin exposure to polychlor break has occurred if they an published at all. tti. inated polycyclic compounds is not supported by Chloracne is a very persistent skin disease and r the fact that chloracne develops following in (he Japanese outbreak of poisoning with chioroof ingestion of these compounds. It is possible that diphenyls, the skin changes were still present in a to the skin lesions arc caused by a toxic efTecl on the number of patients three years after exposure to :d. epithelium of the skin and its appendages which is the ehiorodiphenyls had been discontinued. What tor manifested in proliferation and lots of normal needs to be investigated is whether this persistence ml function of the cell. Since the chemicals are lipid of the skin disease Is caused only by the fact that /as soluble they could occur in a more concentrated once the lesion has developed it regresses very ml form in sebum and therefore afTect sebaceous slowly, or whether it may be sustained by the in- glands more than sweat glands or salivary glands. chloracne-producing chemicals which have been cil The reports on the Yutho outbreak suggest that absorbed snd stored in the body, particularly the un proliferation of the epithelium of the mucous adipose (issue, and are only very gradually :nt membrjnes may also occur. The increased pigmen excreted- In patients with chloracne, adipose tissue he tation of the skin and mucous membranes may biopsies should be analyzed for the presence of the ell indicate a functional disturbance resulting in offending polycyclic polychlorinated compounds itui increased melanin production. Hyperpigmentalion If adipose tissue Is unobtainable, the sebum of the 111 11 u also observed when the adrenals are damaged, a* ear canal or the content of the akin lesloni may illy in Add item's disease, and adrenal function should represent a good substitute. January 1*74 HONS 207669 Apparently there it no known effective treat ment for chlnracnc and the best control measure U prevention. The manufacture of chlorinated hydrocarbons and the coaling of wires and eon* demurs willt insulating matenali containing them should be done in totally enclosed areas so that the fumes of the insulating substances and of the solvents if any are used do not come in contact with the workers. The workers should be provided with dean overalls and underclothes daily. These should be laundered at the plant and cleaned In such a way (hat no chlorinated hydrocarbons remain on them. Shower baths after work should be compulsory and supervised and there should be sufficient shower facilities provided so that the workers are not unduly delayed after work. Special synthetic wetting agents1 *' seem to remove the material from the skin more effectively than ordinary soaps. The experimental induction of chloracne in men has also been accomplished with 2 appli cations of 10 ug 2,3,7,8-fetrachlorodibenzo-p dioxin'*1 and with Halowax 1014, pentschloronaphthalene, and hexachloronaphthalene.'41 Experimental work In animals has shown that most species do not develop the same typa of dermatitis. The rabbit's skin,lls particularly the inner aspect of the ear lobe, provides a good test medium for the development of seneform derma titis. The rabbits, in addition to the development of hyperkeratosis of the ear. will also show a hepetoioxic effect.'1* Microscopic examination of the changes In the rabbit ear were described by Jones and Kiirek.' * Inagsmi and Koga114 were able to produce hyperkeraiocis In hairless mice that were fed rice oil contaminated with Ksmedor 400. These mice also showed hepatomegaly. Microscopic examination of the akin revealed cystic dilation ami hyperkeratosis of Mir rolUdes and sweat glands. Hairless mke may be another suitable experimental animal for Mating the effect of these chemicals on the akin. X-diteese in A nirmb Chiorscne-type lesions do not usually occur in other species excepi for (he rabbit and the hairless mouse, already mentioned. However, in 1947 Olafson' " described a disease In cattle which was designated as X-dismse or hyperkeratosis. Symptomatology included excessive lecrimation. diarrhea, polyuria, marked salivation, and discharp from the nostrils. Tfcs animals developed a chronic cough, poor appetite, numerous toil maculae in the buccal mucosa;in addition,hypef keratosis of the skin developed. The skin was hj?d. and fissured particularly across the wither jua sides of the neck. Many wart-like proliferations on the lips, longue, and hard palate were also observ ed. The abomasum was greatly inflamed, swollen, and edematous with many superficial ulcers.1 ** In the cows that had X-disease, a low vitamin A plasma level was also observed, which again indi cate* that eitemin A may play a role in the etiology of these various disease entities The disease was first thought to represent a virus infection and other factors were also suspeod. After numerous investigations it was finally established that X-disease could be produced m cattle by the ingestion of highly chlorinated naphthalenes, and also by petroleum products such as crankcase oil. Bell*11 teued the tbrltty of the various compounds of the chlorinated naphthalene group to produce X*diseise. He demonstrated In his studies that dicMorinsfed and trichlorinated naphthalenes did not result in X-disease. while tetrachloronsphthilenc had an effect and the higher chlorinated naphthalenes such as peniachloronsphthaleoe, hexacltlomnaphthalene, hepiachloronaphihalenc, and octachloronaphthalene caused severe disease Octachioronaphthalene was less toxic than 1>c \aand heptachloronaphthalene. Wegener'5* repoit ed the occurrence of hyperkeratosis in cynic m Germany between 1946 and 1948. The cuom this outbreak was traced to a wood prcNcnatiu which was used in postwar Germany. The author proved experimentally that animals housed m cabins painted with the wood preservative develop ed typical X'disease. The poisonous ingredient in the wood preservative appeared only in certain lot> of this product and }tier lots did not contain it The poisonous ingredient was never identified by chemical analysis. Animal experiments were conducted with the toxic ingredients of rice bran oil that produced Yusho in Japan. These toxic substances consisted of a mixture of chlorinated biphenyls containin' 48% chlorine, with a trace of 001% of naphth; lenes-1*' When mice were exposed to them (or to 4 months, they 'nowed eczematous changes t the i'km around the eyelids, erosion, uiceratic and perforation of (he ear laps, loss of hair. an erosion and ulceration of the skin around it neck, forelegs, and aides of the chest. The autho 441 CUC CWifrW Snim fc Tbafeefagr HONS 2076 70 Inml. mid tu on rtwrv- uilrn, !* In nn A indiii the I I 1 ' rni a spcti"inally :cd in inj led duels lily nf muled c. Ht rd and ill in ad an iilcnes ancl ISCJiC. ItexaepoitUlc in use of tvauve author sed in tvelopicni in un lots tain ii. Tied by ill) l he nducetl misled iaining iphlhan foi 3 ngej of era l ion nr, and md ihe authors ...iii-d ilia). hi ilic end of (lie ex/vrniienl. some .lunis.-'- in tLie skin tnnil.it in those hi llic csjviiincnijl nuec were seen m some tif the toniml mice alto, which makes i( difficult li> infer pret 'heir findnip. |n h'57 a disease tlul will he discussed in pcjiei (ifinil later in this arlicle occurred in a large number of chickens. Ii was lerined chick edenu disease and was found lo be caused by ceiiatn wen.- fa'. Allen and Caisicm,1 ** fed litis substance iu monkeys and found that, among ulhcr changes which will he discussed tales. Ihe monkeys developed a generalized alopecia and subcutaneous edema one lo I wo months before death. School lie cl il.1*' produced hyperkcralosis of the skin in ran by feeding ihcm hcxachlotimaphliiafcne. Young swine'*1 and sheep'*'* did not develop skin lesions as a rcsnli of exposure lo clilorinalcd naphthalenes. H*nm oniat t Dermal ins and Irritation of lire Skin As was poinicd oul earlier, tlic hyperpigmenla- nun oltseivcd in palienis will) cldoracne as well as the reaction described as cable rash could also be the result of a photosensitive reaction. This photoseinmvc reaction is the predominant lesion in the -Un produced by a number of germicides, namely, huliiouo! (Figure 2). leirachlorosaiicylanilide (TCSA). and a number of other halogenated phenolic coinpounds including fentidor. Patients who develop pitotodermaiith complain initially of i burning sensation, and this it followed by idling. A rash develops which in (he beginning is ciyihematous and associated with varying degrees of edenu- As ii subsides, pigmentation is observ ed Wilkinson' *s was the first one to describe cues where patients developed a photo allergy which was due lo the incorporation of TCSA into popular soaps. He observed that the outbreak of dfunatiiis was confined lo the areas that usually tic exposed to light Most of the cases reported hum England were transient and as soon as exposure to the compound was discontinued the traction subsided. JUlson and Baughman'** and Jillwn.1*1 on the other hand, reported that (heir cues were more persistent. Wilti and Kligman'** demonstrated that the l*wtocontact allergy of the skin with these various footpounds was simply a contact sensitization. The reason that allergic reactions of the skin developed only after exposure lo the sun was due tu ihe photodecompostlion of the various products lo which the skin had been exposed. If various bmmlnaied and chlorinated saticylanliidei were irradiated in viito in an ippioprjaie solvent, lire resulting breakdown products caused an aller gic reaction on contact, without exposure to UV light. These studies were performed because of information provided in earlier studies by Coxon et al.,1** who had irradiated (nominated and chlorinated sallcylanilides and found that Iheae compounds gave off halogens. With 3.5 substituted sallcylanilides, the halogen substituted on the 3 position was lost. Telrachloroullcylanilide photo sensitized subjects reacted lo 3',4', 5-lrichk>fouHcyianilide and to 3', d'-dlchkrrosalicylanded*. A reaction to non halogensted tilicylanHide was not observed. Halogens on the anilide ring wm un affected by UV radiation, while Ihe halogens on the talicyl ring were replaced stepwise by hydro gen upon irradiation. Among the patients who developed photosensi tivity to teirachioroullcylamlide reported by Wilkinson,1*1 a group of palienis in a shop of a factory was also included. This same outbreak was reported by Jones,' '* who found that among 106 employees who used the tame soap very frequent ly, 31% were affected by a dermatitis due to contact with tetrachJorosalicylanilide present in (he soap. Additional cases of photodermatltis due to (he exposure to tetrachlorosalicylanilide were reported by Calnan et a).,1 " who pointed out that pronounced edema of the eyelids was also observ ed when involvement was severe. Some cases were persistent or developed acute relapses on further exposure to sunlight without further exposure lo ihe offending chemical. Frenk111 reported en additional nine cases of acute contact dermatitis of. the hands due (o tetrachJorosilicyltnUtde. Epstein'71 described a photoalle'gic reaction in Iwo patients to IribromotiJicylamlide (TBS). With patch lesls there patients showed a cross reaction lo TCSA. A similar cross reaction pattern to a number of halogenated salicyianiltdes was (Ho observed by Osmuntben1 in patients who had developed photodermatitis following exposure to Irtbromocalicylanilide. Because of the photorensitivity reactions and the cross reactions that were ait > found with bithionol, which it similar in st jciure to tatrachlorosalicyianilide (Figure 2), the U.S. Food and Drug Administration, on October 24, 1967, with drew ail new drug applications of all drugs that jMUarr 1*74 449 m MONS 207671 eonuined hilluonol in tire notice ol withdrawal of approval in ihc Federal Register, tit# ConitniMuiHicr of lit# Fod and Drug Administra tion pointed out tlui blthionof niay in tome instances cause a very persistent pholoseitsitizalion and severe dermatitis may occur wi|h expoture lo nittliglu without furiher conlaci with llte senaitizmg articles, and tlui bithronol may produce croa phniosciisiiizaiinn with other commonly used chemicals such at ccrrain halogcnned talkylanitides and hcxacliloropltene. Hcxacltlorophcnc preparations may alto cauae irriiaiiun of the ikin if these preparations are used frequently. A true allergic reaction to hexachktro* phene it apparently rare.1 T* The more severe reactions were usually confined to the skin of the scrotum and consisted of t primary irritant conlaci dermatitis.1 '*'TT The cases reported by Baker el a!.11* also included 33-month-old Caucasian rentals who developed rath around the buttocks, vulva, and upper thighs following the use of a hcxachlorophcne preparation In her bath water. We observed severe ikin irritation and ulcera tion in rats (hat were exposed dermally lo a 3% hcxachlorophene solution in propylene glycol ora detergent. The skin reaction was not at severe when (he hcxachlorophene concentration was reduced to 1.5%."* Boutwell et al.1 '* reported considerable skin irritation in mice that had 5 mg/mouse applied twice weekly for 21 weeks. Harber et a)."* were able lo induce contact photosensitivity with TCSA or TBS. Thaae authors also found that cross contact sensitivity and photosensitivity to hexachlorophene existed in guinea pip with primary photoaensiUvily to TCSA and TBS. Photosensitivity lo Irichlorocarbanilide (TCC) was not produced in guinea pip. The guinea pig may represent a suitable animal species lo aludy contact and photoaentitrvity of these and related compounds. After exposure to the photosensitizing agents has been discontinued, humans may still be sensitive to sunlight, a fact that needs further study. To a certain extent this could be due to a persistence of the chemical compound on or in the skin. Dichlorophene used as an antimicrobial pre jrvlive may also cause contact deimetitis. Crossaensitivily to hexachlorophene war not demonstrated.1 ** Chick Edema Simpson el al-1 * * and Sanger et al,'*J dcstub i ed a new highly fatal disease in chickens The fnu t outbreak occurred in 1957 in Georgia. Laip numbers of birds from approximalely three wceki t of age to (dull laying hens were affected. Clinical . sfpts consisted of dyspnea, reduced body weight t. pin. stunting, subcutaneous edema, paleness, and (. sudden death. In young chickens, psptng was the t. first noticeable sip. This was followed by j (, waddling, unsteady pit. Gross inspection of the | birds revealed a pale heart, fluid was noticed in the n pericardial sac, and the liven were pale, mottled, and had an irregular panular surface. In advanced a tlapa of the tiacaae the chickens had large h distended abdomens that were filled with fluid 1- Ecchymotk hemorrhages were present in the skin d beneath the wmp, on the lep, and over ihc Q t bone in few chkkens. The kidneys were pale and u swollen. Foliowring the first oulbreak.no iddiiiun- i at cues were reported until late summer and early p fall of 1957, when extensive losses of chickens and s turkeys occurred throughout the southeastern pan t of the U.S. Suspicion soon developed that losses were caused by the consumption of rations con- c taming certain lots of animal fat.111 The animal r fats contained tallows and peases obtained as ' by-products fiom hides preserved with chloro- phenols prior to leather manufacture.11 ' This material was excluded from commerced poultry rations and no additional cases occurred i where animal fats were responsible. Simpson c: .t tl.1"1 came to the conclusion that the edema m ; the chickens was the result of vascular changes a that teemed to be the primary lesion. The vascular c lesion, or endolheliosis, consisted of proliferation f- and hypertrophy of the endothelial cells of b arterioles and small arteries in most tissues. Pro- s nounced prolifertlive changes in endothelium of h the glomerular capillaries were observed, but no f. ripriftcam changes were noticed in the tubular 2 epithelium of the kidney. Necrosis of the paren chyma) cells of the liver was alto noted. In some reports it was pointed out tlui <- exposure lo toxk fit resulted in 1 decrease ol lire h total serum protein and a shift on the albumin ' >> globulin ratio with a decrease in the albumin t fry.lion This could, of course, lead to edema n formation, particularly when combined with nr.<! e functioning glomerulit capillaries. It has also been " suggested that tncieaeed permesbdily uf tin- or- b diovasculsr bed contributed to the edema forma > 444 CHC Critkwl Rrtitm In Toxboiot)' m HONS 207672 ribfirsl irpc cki ileal iglil . nd ; (lie J i ,..;In ciitici studies (tic scrum protein ..xuYiioatinii did itni differ from Dial of (tic ,itiln'ls' "* ' *' and iiurcascd permeability of llie J(juivasaibr bed was advanced ai (he sole cause ,,i the fluid accninuUiion. In addition, pulmonary c^iin was observed in diseased chickens and perivascular lymphocytic infiltration as well as ejeeiia of I he cardiac muscle with interstitial y a ! I, pinny tic inTif (ra I ion |i:ts been noted1** (lie | flection microscopic examination of (he cardiac (lit . ted, iced *rt* uid. tkln *0 and ion:arly and part rases conimal ' i as | muscle revealed degeneration and loss of mito chondria.'** l( is possible dial die heart may play 1 poultry role in (he development of chick edema. Sui a liepaiorenal syndrome could be the under hint cause as well. Studies that lnves(l|a(e the development uf the lesion more closely are needed in elucidate these problems. The (ype of diet also influenced the development of the disease. It ..tuld be brought on more rapidly with semipurified diet ihan with a natural grain ration; indium chloride in the diet was a prerequisite for the development of (he syndrome.1 *' If low raisons of llie toxic fat were fed (o chickens (hey did nui develop chick edema but ^production and liaictiabiliry were drastically icduccd. oro- j Toxic fat was lurt the only product that caused the L-inck edema syndrome. Chlorinated biphenyls rcial rimed llie disease in chickens1 ** and in Bengalese irred finches.16 A mixture of peniachloronaphihalene n ct and liexadiloroniphdialenc fed to chickens also ia in mulled in chick edema disease.1** Cantrell ei inge* il announced in 1967 that one of the toxic :uiar compounds capable of producing chick edema uion found in toxic fat was 1.2,3,7,8.9-hexachlorodi i of benr.o-p-dioxin. Then Iliggenbotham et si.1* Pro showed that llie chick edema factor was produced m or by two principal compounds isolated from toxic n no fils: 2.3.7-irichlorodibenzo-p-dioxjn. and bular 2 J,7.8-ietraclilorodibenzo-p-dioxin. They alio aren- tuggested fils and fatly acids that contain commercial chiorophenola as possible sources of hat contamination. When crude fata and tallows are if Hie heated to produce Tatty acids, chlorophcnol in lo residues may be converted to a chick edema uinm dema mat- factor. Whether these last three compounds mentioned arc llie only ones that produce chick edema disease is questionable unleu the chfori- been : carDrm- nued naphthalenes as welt as chlorinated biphenyls that produced clock edema were also cuniaminaied with them. Another outbreak oT duck edema diaeate occurred in I969.*1 In this incident millions uf birds in Norlh Carolina were involved. The outbreak was traced to a vegetable oil refinery where feed fat became contaminated with "dilurophenols." The company formulaied (lie clilorophcnots as antimicrobial water treat ment products. An underground pipeline from the "pesticide" plant led to traps used to collect by-product fatty acids held as feed fait. Flick et si.'** have recently shown that pyro lytic products of 2,3,4,6-ie(rachk>rophcnnl were toxic lo chicks and produced chick edema if SO ppb were led for 21 days. The product fed consisted of 53% hexa-, 45% hepta-, and 25% octachloro-p-dibenzudioxln. The pyrolytic product of pentachtorophcnol ooiuiaied of a mixture oT 5% hexa-, 62% hepta-, and 33% ociach)orodibenzo-p-dioxin that produced mild edema and increased mortality when fed a diet containing I ppm (200 pg total ingestion) of the material for 21 days. When a mixture of iri- and tetrachlorodibenzo-p-dioxin was fed lo the animals at a dietary concentration oT 0.01 ppm (a calcula ted intake level of 1.9 fig). the birds developed edema and 83% oT them died. Further studies with pure compuunds are ncceuary to confirm these findings. The effect of PCBs on the chicken was again demons) rated when PCBi that were used in the U.S. as heal transfer fluids m food processing plants contaminated large quantities of fish meal that was processed for animal feed in a plant in Wilmington, North Carolina.'*' From about April 1971 until July 1971, KBs leaked into the fish meat and 16 thousand tons of contaminated fish meal wara distributed lo more than 60 companies in 10 slates. One of the purchasers of the fish meal. The Holly Farms, the nation's largest poultry producer, had to slaughter 77,000 fowls after discovering contamination of the chickens with PCBs. Some of the levels found in the adipose tissue oT the animals were as high as 40 ppm. The Holly Farms first discovered that the chicken meal was contaminated because the hatchability or their eggs was drastically reduced to 18% of normal. In February and March of 1968 an outbreak of chick Hems also occurred in Japan, wlterc more tlian 400,000 chickens reportedly died. The diabase was traced lo the contamination uf animal feed with Kanechlor 400. a Japanese PCB. The chick edema disease occurred simultaneously with an outbreak of polychlorinated biphenyl poisoning bi people that was due to the contamination of Janaary i *74 465 HOHS 207673 rice tiran oil, alto cniilanriuatcd with Kanechlor 400 and produced by thcunie company1" Metcalfe1*1 suggested litat chick edema factor could he minimized if fleshings grease from hides preserved with technical peniachlorophenoli containiii| chlorinated dibeniodioxlns war* no lonter used Tor food products. It was also pointed out dial the use oT glue emulsions containing chloroplienois in rendering operations is another possible source of the toxic factor. Whether elimination of tliese sources from animal food products wtil prevent chick edemi epidemics mmains to be teen, since polychlorinated biphenyls that may be con taminated with chlorinated dlbenxofurans have also produced chick edema disease. Yusho During the summer of 1968. 13 cases of chloracne were observed in the western part of Japan, centering around Fukuoka prefecture. The incidence oT the disease seemed to be femfliat and it became apparent that all patients had used the same brand or cooking oil (canned Kenemi rice oil). These findings were picked up by the news papers and a strong demand wu voiced to elucidate the cause. Therefore, a Yunho study group was established by Kyushu University.1** The term Yusho stands for rice oil disease After extensive investigations it was found that several illnesses occurred in a number of households that had used common cans of rice bran oil. When I total number of 89 males and 100 females that suffered from Yunho ware examined, a variety of symptoms ware found in these patients. The incidence of the different symptoms that varied a great deal is given in Table 2. The epidemic outbreak was finally, after very exteniive investigation, pinpointed to Kanemi oil, manufac tured on February S, 1968. Chemical analytis of this oil established that It was contaminated with 2,000 ppm of Kanechlor 400, a chlorobiphenyl that contains 48% chlorine. It was also demonstra ted that most of the components of the chlorobtphenyls were stored in adipose tissue of the patients for quite a long time and pasted through the piaccnU into the fetus of a few woman who happened :o be pregnant.'** The amount of oil necessary to produce disease was found to be at least 2.S I. The younger the patient, the higher the morbidity rate with a smaller total amount of ingested oil. The average amount of chlorobiphenyl consumed by individual patients was calculated to be a total nf 2 g (Various reports written on this subject different accounts of what actually was consumed and the amount can at beat only be estimated, When the rice bran oil was heated nndd reduced pressure. Kanechlor leaked from the old pipes in which holes were subsequently discover The Yusho outbreak apread not only ova Fukuoka. Japan, but also over 20 oifor prefectures in the western part of Japan. A total 1.057 poisoning cases resulted according to the latest tabulation (August 1971. Ministry of Welfare).1** The most common initial syiiipmm experienced by 136 patients with Yuslm was Increaied eye discharge and swelling of the u^pm eyelids (38.3%) followed by acneform eruption, follicular accentuation (33.1%), and prgmcniainm of the skin (9.6%) Itching or the skin and "stiffening" or the soles of the feet and llic palms of the hands were also observed. Pigmented mucus membranes were noticed in many of the patients, a small percentage of them developed jaundice many complained of a feeling of weakness ami headaches; occasionally vomiting, diarrhea, and fever alto occurred. Various neurologRai symptoms manifested themselves such as transient visual disturbances, numbness and spasms in i!ic limbs, and hearing difficulties. Must patient] showed acnelike skin eruptions that resembled tit chloracne observed after occupational exposure chlorinated naphthalenes or chlorinated biphenyls When the age and sex specific incidence rates s\ cic analyzed il was found that a significant sex difference was not present, but a lower risk for both males and females m the age groups over 60 years was noted. The proportion of severe cjscs among those age 13 to 29 was significantly laiga than that of other age groups. During the Yusho epidemic, II women wni, Yusho and 2 wives whore husbands had Yusho. but who themselves did not show the disc-use. delivered 10 livebom and 2 stillborn babies. Nine of these babies had unusually greyish, dark brown atained skin, and dark pigmentation of the gingiva and nails was noted in five of them. Most infants also had heavy eye discharge. Histological exami nation of a stillborn fetus riiowed a marked hyperkeratosis and atrophy of the epidermis .mil cystic dilation of hair follicles, especially m those of the heed. A marked increase of inekmin pigment in the basal cells of the epidermis was also noted. Moat faunas that were bom were smaller 464 OtC Criikwl Xevfcwt Si Toxkviotr MONS 207676 give .intcd iicd.) mdet c old ered. OVCI oilier lal of i the y nT plom was jppet ilinn. ill IOI1 and Mitts uncus ictus; idicc. s and . and ogieal isiem n the licntj J 1 In ure iu cnyls. i were t lex >k for rer 60 Cates larger ' i with 'uslio. tscasc. . Nine hrown tingiva nfunu :xaminarked us and those id an in las also ^mailer TAHLLi2 Symptoms (compbintt) of Patient* (IS male*, too Female*, a* of October It, tM>* Symptom* Male* % Femela* * Rlivlwaiif of nail* lll.nl; ipoif in all pore* i-.M-CMivr itt-callnp in point* . Acnclikc ikin ft up Horn Red upon on limb* Hchinf Chance in nkni color Swelling f hand* and feel Hardening of back* and hand* Figmenlalktn (if inucnaa membrane* Sebum (awn uccrelion In eye*) Hyperemia of tmicoua membrane* la eye* Temporary failinc of eyetfehl Jaundice Swelling nr upper eyelid* Scn*c of wcakncii Numbnc** of hand* and feel lever Ilcaime difficulty Spusmt of hand* and f*e( Head whet Vomiting Diarrliea 13.1 *4.0 SO .6 17.4 20.2 42.7 75.3 20.2 14.7 56.2 HI 70.1 56.2 11.2 71,9 51.4 32.6 16.9 11.0 7.9 10.3 23.6 19.1 75.0 56.0 SS0 <2.0 16.0 520 710 41.0 29.0 47.0 <1.0 71.0 SS.O 11.0 74.0 52.0 39.0 19.0 19.0 <.0 39.0 21.0 17.0 `Reprinted with permitsioti from Kuralsune el al.`* ' than I lie national siandardsand four of them were small for gestational age babies. As they grew older their skin gradually became lighter over a period of three months Okumura and Katsuki1 *1 studied a group of 24 patients whose main complaint had been acneform skm eruptions and who had been diagnosed as definitely having Yushc>. Eighteen of these were adults and six were children Irrespective of the seriousness of the disease in all patients, the initial symptoms were eya discharge anociated with edema of the eyelids, defects in vision, generalized fitiguc, anorexia followed by comedo and acneform eruptions of the face, and pigmentation and flattening of the nails. The principal subjective symptoms were fatigue numbness of the lower limbs, and inlolcrar e for atAc, which is a Japanese alcoholic beverage made from fermented rice Three of twelve females had an irregular menstrual cycle Seven of nine patients with serious disease showed a low grade fever. Most laboratory tests conducted on these patients were normal, including serum enzyme values. A slight elevation in alkaline phosphatase was observed in over half of the seriously ill groups. An increase in the ] globulin fraction of the serum protein was noted with a concomitant reduction of the albumin fraction, but the total serum proteins were within the normal range. These same 24 patients had abnormally elevated aerum tri glycerides ranging from 200 to 600 mg % In 12 of 24 cues, while the total serum cholesterol remained unchanged, phospholipids tended to be somewhat lower.1** Preliminary animal experi ments in rabbits conducted by (he same authors showed that lipemia was consislently induced within several days. The levels returned to their previously normal values several weeks after expo sure had been discontinued. The serum concentration of triglyceride is usually elevated in humans in glycogen storage disease (Von Gierke's disease). An increase in plasma triglyceride concentration is also observed in essentia/ hyperjlyctridemm, an apparent con- Janeery 1974 4*7 HONS 207475 jyiHij! anoniu|\ f:.*scuii;il liypcrglyccridenriii has lu'i'ti utisetveJ tn ol j iiinnhci of families, Imii ilu' mode ill transmission is not known, The course of this disease is generally benign and is consistent with a normal lifespan1 ** Observation of the familial eases shows that llte mere elevation of lr(glyceride dues not in iisclf increase the risk of arteriosclerosis. Another iiMVicilmy fact in tilts poisoning epidemic w:is Hut nut of 2.1 cases Willi obvious Yudin. 10 showed sy mp I inns or signs of a sensory iicliiopjlliy illm included numbness, pain and liy poestbcsia, and. in addition. I case also showed aicllc*iii 5fl#',B 1 This effect on the sensory nerves would nor be immediately obvious in animal eaperimeors and probably sliould be investigated furl her. Samples of sputa and adipose I issue collected from patterns who suffered from Yusho contained chlorinated biphenyls on chemical analysis.1*4 The concentration in adipose (issue was much higher than in sputa. Recently Kuratsune el al.1** reported on the present status of the patients who suffered from Yusho. Many of tle patients were, still suffering from the illness. When tire findings ntade in 159 patients in 1970 were compared with those observed in 1969, it was found that half of tiie patients had improved clinically, while the remaining half showed no such improvement and more than 10% of the patients had become worse. Immunosuppression Several reports have indicated that KBs and possibly some of the chlorinated dibenzodioxin* and furans may alter the immune response in animals. Alrophy of the thymus and reduced total white count were observed in rabbits following the dermal exposure to f*CB fractions containing letraand pentachlorodlbenzofuran.** In fish, exposure to Aroclor 1254 for 14 to 45 days ted to emaciation and funguslike skin lekons.** However, no attempt was made to isolate organisms from these lesions. When I (May-old mallard ducklings were fed diets containing PCBs (Aroclor 1254) and were subsequently injected interperitoneally with a duck hepatitis virus, significantly more ducklings died than when they received the virus alone.,# * In guinea pigs, subletItti dosage levels of 2,3,73-tetrachJorodibenzodtoxin produced severe atrophy of the thymus with destruction of lymphocytes, lymphoid depletion in spleen and lymph nodes, as well as extensive hemorrhages into various mpim including the adrenals. Reduced thymus and spleen weights were also observed In rats Hut received a single dote of 25 pg/kg TCDD and in mice the cell mediated immunity was found to be suppressed.1** The various findings listed Iwrc du not conclusively prove (hat the Immune response is specifically altered by these compounds as we encounter It fallowing radiation exposure or therapy with alkylating agents. In most studies relatively high doses of the chemical compounds were given, resulting in general toxicity accompanied by weight lues. Weight loss in itself may lead to atrophy of She spleen. In humans maintained on a kw eatoric intake with negative nitrogen balance, a genera) reduction of lymphoid tissue results which Is not specifically related to the immune response and vitamin A deficiency is 1*0 accompanied by a reduction in lymphatic tissue. In this respect it it alto of interest that when monkeys were fed "toxic fat" containing polycydic polychlorinated hydrocarbons. The germinal centers in the lymph node* and spleen as well as islands of hematopoietic cells in bone marrow were extremely sparse.*** This is more extensively discussed in connection with chick edema. The fact that a combination of duck hepatitis virus and PCBs is more toxic to 10day>old ducklings than the virus alone suggests a reduced immune responee but could also merely represent an additive damaging effect on the liver, made more susceptible to infection by the toxic effects of KBs. So far no convincing evidence of "waiting disease" as a result of exposure to chlorinated polycyclic compounds has been reported in rodents or any other species in the literature. Atrophy of the thymus in newborn rodents seduces the capacity of these animals to produce serum antibodies, resulting in westing disease, also referred to as runt disease or homologous disease. Such animats are then incapable of rejecting foreign skin grafts,*** which probably represents the moei convincing evidence for ftomologoti* disease. A shift in llte immune response could Influence the development and growth of cancer and alter the defense mechanism in infections. In order to prove that KB* as well as chlorinated dlbenzodloxins and furans do indeed Influence the immune response, the effect of these substsnees on newborn animals of several specks should be tested with adequate parameters such as skin n a P ip rc Bt r. at cli ><> fr: fl* lb' sir Mu Sin hei (A act aci br>i sub wo acu i jt: II in. i pity tntt noli rein stud 73 . of t 1 first and rirtur: (Yiisl not ri Ar us < purpl porpi ! ulien j diet. , put so j When j in tin : becin 4M CMC Oitretf Mrrirwt in Toxtrviagy MOMS 207*76 and I It.11 J in i> he e do nose t we m id tel imds icily tsclf nsrti stive hold d to :y is wlic that ninp Tlic tn as >rme note hick juck 10jesis rrely iver, oxic :e of : to h*en the ienls duce also ease. :iing semi gons ould inccr S. 'll it, ted e (he meet d be (kin ^jriing and the incjsnting of Hie development of stilibndies. Whetlter the oilier chlorinated polycylie compounds In this article also afTect the immune response hat nol been suggested or reported. Biorliemical Effects and Chemical Toxicology forphyna Porphyria cutanea tarda in humans is an acquired defect in hepatic porphyrin metabolism, cluraclerized by uroporphorinuria, photosensitiviiy ss manifested by blisters, and mechanical fragility of the akin. Tlie hepatic porphyria (Porphyria ciilama tarda) which is responsible for the increase in porphyrins and the skin photosen sitivity can be produced experimentally by number of drugs: all of these have the ability to stimulate the activity of the initial enzyme in the heme synthesis S-aminolevulinic acid synthetase (ALA). In addition to this acquired porphyria, an scute intermittent porphyria erthropoietici and an acute intermiiieitl porphyria liepatica, which are both hereditary, exist together with i number of subgroups which will not be considered here. In 1964, Bleiberg et al.' reported studies of 29 workers in a 2.4-D (2.4-dichJoTophenoxyacetk acid) and a 2.4,5-T (trichlorophenoxyacetic acid) factory. Many of the workers had chloracne and I! had abnormal excretion of urinary uioporpltyrins. Many of the workers with uroporpliynnuria had hirsutism, hyperpigmentition, and increased akin fragility. Liver dysfunction was noleri in iwo hotpilalized patients. Poland et at.1 reinvestigated ihe workers of the same plant studied by Bleiberg et ai.3 They studied a total of 73 male employees; chJoracnc was found in 80% of the workers. In this study six years after the first report, clinical porphyria was not observad and only one worker had persistent uroporphyimuru. In the Japanese poisoning epidemic (Vutito), increased uroporphyrin excretion was noi reported. Apparently polychlorinated biphenyls, at well as tetrachlnrodibenzodioxins. produce hepatic porphyria in animals. Experimental hepatic porphyria was observed in Sherman strain rats when the rets were exposed to Aroclor 1254 in the diet. The porphyria resembled hexachiorobenzene poisoning and human porphyria cutanea tarda,1* When female rats were fed 100 ppm Aroclor 1254 in the diet (approximately 7.9 mg/kg/day) they became porphyric after a delay of about 2 to 4 months. At this time the urinary porphyrin excretion increased rapidly. Uroporphyrin excretion was elevated to a maximum of 550-fold, prophobilinogen 27-fold, and 6-aminolevulinic acid 18-fold. The major porphyrins found in urine from Aroclor treated rets were 8-cerboxyporphy rin (73%) and 7-carboxyporphyrin (16%), Substantial increases In amounts of 5 and 6 carboxcylic porphyrins were also observed. A 10-fold increase in the excretion of coproporphy rins was observed, but the increase In uroporphyrin excretion was 1,400-fold. In control rat urines, coproporohyrin (81%) repremted the major porphyrin fraction. A good correlation between urinary porphyrins and liver porphyrins wis also observed. Only a trace of porphyrin was found in liver samples In control rats. When feeding rats 100. ppm Aroclor 1254, 6-aminolevulinic acid synthetase was not increased in tire first month of exposure. After this time 6-smtnolevulinic acid synthetase was elevated 3to 6-fold in porphyric rats but was normal in nonporphyric Aroclor treated rats. Cytochrome P-450 and microsomal heme were also increased throughout the study but the increase was maxima] in one week despite the absence of an increase of the rate limiting enzyme in heme synthesis 6-iminolevulinic acid synthetase. When a large single dose of Aroclor 1254 (500 mg/kg) was given, a 4-fold induction of 6-aminolevulinlc acid synthetase could be demonstrated in the liver 5 hr after dosing the animals. These results suggested that 5-aminolevuiinjc acid synthetase induction occurred when porphyria hid developed if lower levels of Aroclor 1254 were fed over i period of time, but when high doses of Aroclor were given it was induced immediately. Vos end Koernan*' reported that chemical porphyria was produced by the two European PCB products, Plienodor DF6 and Clophen A60. u well as the American product, Aroclor 1260. The porphyrogenic action was observed in chickens, quad, and rats. It has not been estiblirited so far whether only certain isomers of the polychlori nated biphenyls or contaimnstion with a chlorinated dibenzofuran Is responsible for the production of hepatic porphyria cutanea tarda. Poland and Clover1 ** used fertile chicken eggs it i developmental stage of 15 to 20 days and injected them with 25 sd of the Mogtmiad dibrmo-p-dioxin]. They found that 2,3,7,8-tai ri ch]orodibenzo-p-dloxin produced a daae-mlated HONS 207677 increase in ALA synthetase activity. The lowest dote tested, 4.66*1 O''* mol/egg (1.5 ng) dmbled the enzyme activity. Tliete rendu were statistically significant. Studies with other lialogenated dibemo-pdioxins give the following results: the 2,3.7-utcliloro and 2,3,6-iribnimo isomers are puteiX inducers of ALA synthetase while 2.3-didrloro and 2.7-dichloro, 2.8-dichloro, 1,3,6,8-letrachloro, and 1,2,.7.4-tetrachloro Isomers all failed to Induce ALA synthetase at doses up t 2.5 ing/egg. Of the limited number of haiogcnaied dlbcnro-p.dioxlns that have been tested for fetal toxicity and the ability to produce chloracne. those that are toxic at low dotes also induce ALA synthetase. Whether some of the poly neurotic symptoms reported in Yurfio and also rn severe exposure to technical 2,4,5-T or technical irichloropitenol that were heavily contaminated with letrachlorodibenzodioxin are caused by an effect on the porphyrin metabolism merits investigation since neurotoxic effects have been reported in acute intermittent porphyria.1 *7 The dioxins that induce ALA synthetase in the liver of the chick embryo may also induce it in the mammalian system. Woods14 was not able to induce 5-aminolevulinic acid synthetaae (ALA synthetase) in the liver of male rats up to 30 days after oral doses of 2,3,7,8-tetnchlorodibenzodioxin. Guinea pigs and mice also did not rftow an increase of ALA synthetase following Ihe admini stration of 2,3,7,8-teirachloTodibemodioxin. How ever, it was possible to demonstaie an Increase in ALA synthetase and uroporphyrins in the liver in Csi black male mice given 25 srg/kg/week for 4 weeks.1** It Is possible that ALA aynthetaae induction varies in time with age and sex and may show species variation. Induction in mammals may require longer dosing, since, for instance, porphyria and induction of ALA synthetase with moderate doses of Aroclor 1254 did not develop until 2 to 4 months after dosing was started.1** Female animals and the embryo ray be more susceptible than adull male animals. Effect on Other Enzymes and on Vitamin A A number of Investigators have shown that site polychlorinated biphenylt induce liver microsomal enzymes.110 ' 1 Norback and Allen1" correlated an increase of hepatic microsomal enzyme activity with smooth endoplasmic reticu lum proliferation. Ultarsi at al.111 studied the effect of four PCD mixtures, namely. Arnclm 1242. 1248. 1254, ind 1260 The animals wenfed the nttterisl for 4 weeks and dietary lev civ employed were 500, 50. 5 and 0.5 ppm At higher dietary levels an Increea in liver weight* was observed. Elevation of triglycerides in lire liver seen at 500 ppm with a maximum effect occurring at approximately 50% chlorine conic nr An increase in cytochrome P-450 was observed :rt the 50 and 500 ppm dietary levels of all PCIi nuxiurc*. An Induction of the pentobarbital hydroxylainm and an increase in methylase activity were observed at the higher dietary levels. Induction <>i pentobarbital hydroxyla tion increased with Increasing chlorine content of the PCBs Nitm. reductase activity was markedly stimulated with Increasing doset of PCB and the induction was seen at levels as low as 0.5 ppm for all mixtures ft it difficult to evaluate the enzyme induction at these low dietary levels since the control animal food can be contaminated at limes with PCBs. ' Two types of inducers of drug-metabolizing enzymes of the liver have been reported: one group to which phenobarbftal belongs resulted in increased PASO content of the liver and induction of a varitty of pathways including benzpyrene hydroxylase and dcmethylation ofethylniurplnne The polycylic hydrocarbons such as 3,4-benzpyrene belong to the second group, which resulted in tiw formation of an abnormal vvi. chrome. P-448, and increased benzpyicne hydroxylation, but not ethy(morphine dcmcil'V l>tion, Alvares el at.1'3 reported altar PfU trc.i ment produced an increase in cytochrome IM4N. an increase in benzpyrene hydroxylation. and N-demethylation. Since the PCBs represent mixtures, it is possible that the type of induction may depend upon the Isomers present. The various effects of PCBs on liver microsomal enzymes can be blocked by the prior administration of ad mu mycin D. When the effect of a number of PCBs on the beef heart mitochrondrial enzyme system was atudied, it was found thar they inhibited NADIIoxidase, tuccinoxidase, and cytochrome oxidase activity. It has not been established whether these same effects will occur in vivo. Benthc ci :ii: 1 * Induced liver microsomal enzymes by uuraperitoneal injection of a single dose of Aroclor 1248 or 1232. Most of the microsomal enzymes hsve been Studied In the ret, but they ere effected in other ^eda a well- Altai et el.111 reported liver 4Ti CPC Crttkel Kerin* fa Toiicolatr MONS 207678 Of I ,f ! Is u'l I :is \n hf cs. on re of iili nv nil fas ll al ml ng IIC in on DC 1C. at di lo ne la in IS. nd in on >us an lo on 'li ne CSC I* rator :cn tier ver hypcniupliy iftei feeding polycliloiiiiaied biphenyls (Aroclur 1248) or polychlorinated in phenyl (Amclw 5460) for three months lo jiii.-iics The increase in liver size was attributed io i lie increase in tmoolh endoplatniic reticulum and induction of microsomal enzymes was observed. Proof of the induction of microsomal enzymes in birds is the finding nude by Lincet and Pcakall1 " thai livers obtained from American kestrels exposed to cither Amclot 1254 or Arocior t202 tllowed an increase in the in vilro breakdown of ncsiradiol |o a more polar metabolite. This effect on sex hormones is also emphasised by Plitnnow et al.*'17 Arocior 1254 given lo Yorkshire hoars reduced the urinary excretion of gonadal steroids. Both (lie polychlorinated biphenyl! as well as the chlorinated napliihalciici affect the vitamin A content of the livrr. This was particularly studied with the chlorinated naphthalenes since hyperker atosis and other abnormal keratin formation of the skin is often accompanied by vitamin A deficiency. Low vitamin A plasma levels were observed in cattle with X-disease and young swine.1 *1,141 A decrease in vitamin A storage in (tie Jiver of Japanese quail and rats that were fed Arocior 1242 was also demonstrated.11 * Since vitamin A deficiency affects reproduction, this aspect should he further investigated to determine whether the effect of PCBs on reproduction it secondary to vitamin A deficiency.1 11 Ecobiciion snd Johnstone110 studied the effect of various PCI! isomers on the hepatic enzyme system. Biphenyl itself did nor cause induction of hepatic drug metabolizing enzymes and neither did monochlorobipltenyl (4-Ct). Substitution of chloro groups at the three and four positions of the dtand telrachlorohiplienyls caused induction of microsomal monooxygenaset. Induction of micro somal monooxygenases occurred also when hexaand ocrschlorobiphenyls were given to the rata. Niiroreductase and carboxyies(erase activities were not affected wiiile all compounds produced marked induction of the eulfobromophthalein conjugating system. Terphertyli as well as chlori nated terphenyis increased the pentobarbital meta bolism in the rat with corresponding increase in smooth endoplasmic reticulum which suggested microsomal enzyme induction.111 ,311 Lucler et at.131 reported that a tingle oral dote of 5 or 25 Mg/kg body weight of leirwchtamdlbentodtoxin induced cytochrome P-450 gnifi- cantly in male rata for a period of up to 28 days. Cytochrome b-5 content was also Increased. Hydroxylation of aniline was induced and amido pyrine demelhylalion rates were decreased. Micro somal proline was increased by approximately 15%, The induction of UDPgiucuronyltransferasc was moat striking. Oxidative phosphorylation rates in rat liver mitochondria using succinate as a substrate were unchanged. The enzyme induction was observed at early as one day -- the peak induction was reached between three and Sen days after treatment. The enzyme induction was accom panied by an increase in smooth endoplasmic reticulum and a mild increase in rough endo plasmic reticulum,11* These changes were Aral observed three days after the injection of the telrachlorndibenzndioxin. Thus, enlargement of the liver during exposure to various chlorinated compounds it accompanied by an increase in smooth endoplasmic reticulum. Furthermore, if "inchisioni" or "hyaline bodies'* re teen within the cytoplasm with the light microscope, concentrically arranged membranes that often surround lipid vacuoles can be teen with the electron microscope (Figure 6). Hexachlorophene, on the other hand, does not increase the size of the liver.'1 * The livers show no uitraatructural changes and probably microsomal enzymes are not induced. Whether other compounds listed in Figure 2 affect liver microsomal enzymes is not known. Hexachlorophene has an elTect on a variety of other enzyme systems than those mentioned thus far. Cammer and Moore11* and Caldwell et al.11* have shown that hexachlorophene uncoupled oxidative phorphorylation, Increased adenosine triphos phatase activity, but did not affect the activities of electron transport enzymes when compared to controls. Gouid et at.111 demonstrated that hexa chlorophene inhibited tucclnoxidase activity of rat heart, liver, and kidney preparations. According to unpublished material cited by Gluck,111 hexa chlorophene la conjugated to the glucuronide conjugate which is excreted in bile. The proportion of conjugated hexachlorophene to unconjugated hexachlorophene in blood may be more significant in determining the toxicity of hexachlorophene than the hexachlorophene blood levels as they are presently determined. Methemoglobinemia In the early 1960a outbreaks of methemoglob- Jeeverr 1974 471 --------- --------------J m MOMS 207679 incnius were reported from hospital nur series.33''3 10 One outbreak involved i total of 18 infants with a level of methemoglobiit from 4.9 to 30% of the total hemoglobin. An epidemiological investigation traced the outbreak to 3,4,4tricUloeocarbanilidt (TCC) that was used aa a 2% solution in a rinse in the laundering process, after which the laundry a neutralized with an acid and then dried in an autoclave. Twelve of the eighteen eases in this report were prematures who were probably longer exposed to the offending chemical than term Infants since their hospital stay is usually longer. It is also possible that chemicals penetrate more easily through the skin of prema tures. In another outbreak, methemoglobinemia occurred among a group of custodial patients in an institution. These patienls were given soap suds enemas with boiled soap (hat contained 2% Tcc."' It is possible that by heating TCC a breakdown product occurred which produced the methemo globinemia. P-chloroaniline and 3,4-diehloroaniline have been suggested es breakdown products. None of the other compounds discussed in this article have thus fir been shown to produce methemo globinemia. Stonge, Diitfibution, Metoboiitm, and Excretion As we progress further in our study on the distribution, metabolism, excretion, end absorption of compouods such ts chlorinated biphenyls, chlorinated terphenyls, the chlorinated dlbeniodioxins, and chlorinated naphthalenes, we will probably find that since they ire lipid soluble they are stored in quite the same way as the more studied persistent pesticides such as DDT end dieldrin. From the evidence presently available on the chlorinated biphenyls it can be assumed that the turnover of these compounds in the body, pirlicularty of those thit have a high chlorine content, may be slower than some of the persistent pesticides. The turnover rate will vary with the degree of chlorination. Hexachlorophene, on the other hand, is flirty rapidly excreted end presently no information is available on blthionol end related compounds that have e germicidal effecl. All of these compounds are not very well absorbed by the gastrointestinal tract or the Skin. Analysis of various tissues of rats given a single dose of the PCB Arodor 1254"3 showed the highest concentration of PCB in edlpoee thsue and the least in plasma. Drain, liver, srnl kidney had levels of the same order of magnitude t>ut ware lower. If, for instance, 1,600 mg/kg tindy weight of Aroclor 1254 were given by stomach tube, 24 hr later the mean concentration in adipose lltsue was 1,146 ppm. The concentrations observed in individual animals varied greatly. Similar levels were found in adipose tissue when 3,200 mg/kg of ArOclor 1260 were given by stomach tube to adult female rats. The levels til Arodor 1254 found in the brain showed a mean nf 138 ppm and the values observed for Aroclnr I '(>0 were 145 ppm. AM of the values were calculated on a wet weight beds. The ante concentrations m ppm were found -with both compounds even though only half of the amount of Aroclor 1254 was given. This is probably due to the fact that only > certain amount of Aroclor is absorbed at i given time. When 100 ppm of Aroclor 1254 was fed to male weanling rats for 58 days the mean concen tration of Aroclor derived materials found in the feces varied roughly between II and 35 ppm. After the animals had been removed front the experimental diet for week, the excretion in ihc feces was 8 ppm and after 16 days' recovery an average of 2.5 ppm was found in the feces PCBs at the concentration of about 2 ppm were still observed in the feces 24 days after the animals were removed from the experimental diets. During the entire study the amount of PCB derived material recovered from the urine remained Mow 1 ppm. While PCBs were fed to the rats, the gas chromatogram obtained from the feces was superimposable on the one obtained from the standard Arodor 1254 that was fed to the rats. Once Ihc animals were no longer fed Arodor 1254 the number of peaks observed in the gas chroma tograms that resulted from the analysis of the fecal material decreased. Fewer peaks were also observed in the gas chromatograms obtained for the tissues and from urine. During the SS-day feeding period, a gradual increase in the concen tration of the PCB derivatives was observed m thsues. The highest concentration wis found in adipose tissue; after 58 days it reached mean values of about 500 ppm. The concentration in the liver only tmounted to a mean of 12 pprit at that time. Following e recovery period of 71 days when the enimals were fed plain chow, the concentration of the PCB derived material in adipose tissue had not decrested appreciably and 472 CMC CHtteel Meritwr In Toikrjtofy HONS 207680 still ill awed values in I he neighborhood of about 400 ppm and 4.7 ppm was found in Ihr liver. The concentration in tiie brain never reached very high proportions: it ranged from about 2 to 4 ppm Following a 71-day recovery period the amount in the brim was Mill about 2 ppm. When Aroclor 1254 was fed for 240 days at the dietary level of 100 ppm, the concentration of the PCB derived material observed in adipose tissue had reached values of about 1,000 ppm. If, on the other hand, 100 ppm of DUT is fed to male rats for 2 yean, the concentration in adipoie tissue is about 95 ppm''* In a separate study where 500 ppm of Aroclor 1254 was fed to male rats for 6 months tnd the exposure to PCB containing diets was subsequently discontinued for an additional 10 months, the concentration of the PCBs ranged from 924 to 1,688 ppm in edipoie tissue with an arithmetic mean of 1,192 ppm. The concen trations in the liver ringed from 17.3 ppm to 26.24 ppm with an arithmetic mean of 22.65 ppni1,4 Further studies are necessary to determine whether PCBs are stored in tissues to a certain point when a steady state is reached and the concentration does not increase further. The given concentration at which this equilibrium is reached in adipose tissue would vary with the amount fed to the animals and the type of compound. The effect of itirvition or illness when the amount of adipose tissue in the mammalian body it suddanly reduced also needs to be investigated further. Would this lead so increased excretion of the material or to redistribution or both? Dahlgren el al."1 who determined PCB tissue levels in pheasants, found that a concentration of 300 to 400 ppm of the PCB derived material was usually found in the brains of birds that had died, while live birds that were fed the same amount of PCBs had much lower concentrations in (heir brain tissue. Studies in bobwhite quail showed that relative peak heights and chromatogram for the tissue extracts were similar to the Aroclor standard after partitioning with acetonitrile-hexane when the birds were given a single capsule of Aroclor and their carcasses were analyzed toon after exposure. This suggests absorption of ill components of Aroclor. The pstrointestinal tract, liver, akin, wings, and feet had been removed prior to analysis. When bobwhite quail were fad Aroclor 1254 for 14 days and then allowed to recover, tta pt chromatograms showed gradual reduction in (he number of peaks and after 42 days on a control diet only 4 major PCB components remain, which suggested selective excretion and/or metabolism of some of the components of the Aroclor tested,"* In a reproduction study. Heath el at.** analyzed 2 second-year eggs of mallards fed 25 ppm Aroclor 1254 in their diet for 2 years and found 56 and 33 ppm PCB by wet weight. The gas chromatogram showed a pronounced dimin ution of the early peaks when it was compared to the standard, suggesting again a aclecUvs reduction of the components with lower chlorine concen trations. This is also evident in amriroauweiital samples. Lower orpnism such as mussels aad ftah contained a higher concentration of PCBs with lower chlorination than bkds11 In mammals, excretion of PCBs in milk and transplacental paeaage is also observed. Feeding cows 200 mg/day of Aroclor 1254 for 60 days'* resulted in higher PCB levels in milk fat than in body fat. At 30 days' axposure, 45.6 ppm was found in mBk fat and 21.9 ppm in body fat; at 60 days' exposure, 66.7 ppm was found in milk fat and 44 ppm in body fat. However, once exposure to PCBs was discontinued, the concentration in milk fat tank below that of body fat. Transplacental passage of PCBs also occurred."''"* Sherman strain rats were given Aroclor 1254 in peanut oil by Momtch tube during days 7 to 15 of gestation. A mean of 20 60 ppm PCB derived material was found in their milk when 10 mg/kg/diy was given to the dams during pregnancy, while those given 50 mg/kg/day axcreted a mean of 66.34 ppm in their milk on a wet weight basis. Fetuses taken by Caesarean section on day 20 of pregnancy contained a mean of 0.63 ppm and 1.38 ppm of PCBs, respectively. With a fivefold difference In dosage, the concen tration In fetuaes showed a twofold difference and in milk a threefold difference."* The hyperpigmented babies observed in the Yutho Incidents In Japan represent additional circumstantial evidence of transplacental passage of PCBs. Polychlorinated biphenyls have also been de tected in human adipose tissue. Biros et al.'** analyzed two human adipose tissue temples which contained 200 and 600 ppm PCBs ranging from pentachlorobiphenyl to decachlorobiphenyl and Price and Welch'** found 100 ppm PCBs in another human adipoae tissue sample. According Jsnaary lT4 4TJ HONS 207681 It* 1`u.v .mil H.'ltli " II In l> nl tin|M|tti):ilinii 111 I In' t'S ,nt)(jin ) |*]Hll of more I'Uls in their jJijxrc ltic An*i*m; ITU sources Hicy discussed tltc fixwl chain and bouse dust wind*, from residences in southwestern Michigan, con mined up to I HO ppm PCBs. Polychlorinated biphenyls were found in .lit of ft.17 samples of human adipose tissue dial were collected from the general |Ht|Hii;ilinn of I he I IS at purl of the Ihiiiiiui Monitoring Survey Of rite samples ana lyzed. S9t- cottlaincd mote Ilian ' ppm.'41 PCBs luivc also been found in human adipose tissues of the general population of Scandinavia*4 * and Japan,544141 and in human milk and human adipose tissue of tlie general population of Germany.*4 * In general, low concentrations of PTBs were stored in adipose tissue in a certain proportion of the general population. This Is probably more prevalent in (he highly industrial ized countries and almost nonexistent in under* developed countries unless most of the food was imported from polluted areas. These low levels of PCBs are liarinless. as far as we know at the moment, however, the occasional levels found in the 100 to 1,000 pptn range give rise to concern. It would be of interest to establish whellier average adipose tissue levels increase with age and whether those in certain professions such as painters, printers, textile workers, workers in plastic fac tories, and (hose engaged in the manufacture of transformers and capacitors hid higher PCB. levels thin the general population. Reports on the absorption, storage, and metab olism of chlorinated naphthalenes, terphenyls, dichlorophene, bilhronols, fenliclor, trlclotan, and tetrichiorotalicylaniide are almost nonexistent In the literature. Some unpublished results on some of the compounds used as germkidet are men tioned in the summary minutes of the Over the Counter Drug Panel on antimicrobials. Copies can be obtained by writing to Dr. Mary Bruch, Bureau of Drugs, Rockville, Md. Approximately 6% of irictomn. also called Irgaian DPI00 and CH 3565 (2,4,4`-trichloro-2'hydmxydiphcnyl ether), is absorbed when It is applied dermally in a soap solution. The halfllfe of the chemical was 20 hr when applied dermally and 10 hr when given intravenously. Humans excrete 65% in urine and 20% in feces after intravenous injection. It is excreted either as a free compound or as a glucuronide. In dogs nearly 100% of the injected material is recovered as the ghicnroniilc or sulfate in nnin and feces over a 5-day period It Iras been ilmuii that TCC is absorbed through the skin. Tissue levels of the chlorinated Itr/ihcti i / Arodor 5460 were determined in codfish.54'' h* this study it was found that Arodor S4t>0 w;i* apparently poorly absorbed from the gastrointesti nal tract; It was stored m all tissues that were analyzed. Tire highest concentration was fmtml m the liver and following a single dose of I g some nl it was Hill present in Ihe codfish after 70 day ., m Indication of slow excretion. The authors pointed out that with the method used to determine the chlorinated terphenyb, it would be difficult i distinguish chlorinated biphenyls from chlorinated terphenyis A comparison of the chromatograms of the standard with those obtained from clilonn tad terphenyis extracted front tissues revealed more of the earlier peaks in material obtained from tissues and fewer of the later peaks. A variation in the intensity of (he peaks was also observed. There seems to be a change in the chlorinated terphenyis within the living organism, however, pari of this could also have been Intro duced as an artefact by the analytical methods in by differential absorption from (he gastrointestinal tract. Calves borr. to cows with X-disease develop hyperkeratosis and it was found that IrigliK chlorinated naphthalenes were excreted in milk.1 ** This suggests that chlorinated naphtha lenes may vwll follow similar storage and excretion patterns as PC Be. Chlorinated dibenzodioxins and {titans arc probably also stored in tissues of animals and nten who are exposed to them, but again not mm.1) in formation is presently available. Firestone et al.*41 fad the untaponifiable fraction tsolmod from "toxic fat" or 3% ''toxic fat" itself to young cockerels. The "toxic fat" contained whloiodibenzodioxins with 2 to 8 chlorine atoms. When the excreti were analyzed it was found that a certain percentage of the hexachlorndihen/odioxins had been absorbed from the gasirointt-siInal tract, while all of tire oc-iachlurodilvn/**dioxins remained in the gastrointestinal tract, ami only very little of the Itepiichlurodiben/.odtoMn was absorbed. Because of the presence of into faring components, the chick tissues and exucu could not be examined for the lower chlorinated dioxins. In the orpns. the highest concentration wts found in the liver and an appreciable amount (7( OtC Critical Review* to ToxtcvkjfY -HONS 207682 js .ilto found in the hone aiul skin, while the .uli.-i mpans amiyincd very few dioxins. Some nidi-mv for a possible metabolism or hexa- and h,-pi klilmodihetizodioxiii by tlie chick was also observed. Zilko and Wildish94 * fed di-. trl-, tetra-, ind ociaehlorodtbcnzofuran to fssli and detected only oeiwhlurodibemtofuraii in muscle and pit uf dead fish- One problem with metabolism, storage, and distribution studies of chlorinated dibenzodioxini and furjns is that not all of the com pounds are readily available and, if they are available, any long-term feeding studies in Urge animals are expensive. The very toxic dioxin compounds can be given only at auch low levels that they escape detection easily. In our labora tory we detected a chlorinated dibenzofuran in the unite of rats Ted Aroclor 1254. Wc found this same dtlonitaifd dibenzofuran in the Aroclor ilielf4 as a contaminant. Williams cl a!.94* fed rats a total of 22.7 and 120 7 #ig of ociachlotodibenxodioxin. They re covered a great deal of the compound in the faces. Very small amounts were recovered in the liver at both dosage levels and in adipose tissue at the higher dosage level. Further studies are needed to establish the absorption, excretion, and storage of these compounds, and ilieir persistence. Studies with labeled material tnay eventually give us some information on their metabolism. Piper et al.9** gave a single oral dose of CM labeled 2,3,7,8letrachlorodtbenzo-p-dioxin to rats and found that about half of the radioactive material was re< covered within 21 days in the feces and only 13% in urine and 3% in expired air. Most of the remaining radioactive material was found in the liver. In another study9*1 100 pg of Cl1*octachlorodibenzodioxiu was given daily to rats by stomach lube for 21 days. Lets than 0.3% of the material was absorbed. Subsequently the rats received a control diet for 7 weeks and Mill showed 20% of the absorbed material found at 21 days in (heir tissues. The in vivo metabolites of the various com pounds under discussion have in most instances not been identified. Block and Cornish9*9 studied the urinary excretion of biphenyl and 4chlorobiphenyl in rabbits. Biphenylglucosiduronic acid and 4-hydroxybiphenyt were isolated from urine of rabbits fed biphenyls. The rabbits fed the a -c h t o r ob i ph e n y I excreted 4-(pchlorophenyl)-phenol and 4-chlorobiphenyl glu- cotidumnide. Twiee as much 4-chlombiplicnyl as biphenyl was exacted aa (he ghicnsidutontc acid derivative. It is possible that other low chlorinated biphenyls and naphthalenes are exereted in urine in a amilar manner; whether they would be delected by preaent gas chromatographic methods would have to be checked by combining the identification of radio labeled material with gas chromatography. Hutzinger et al.9** found evidence that some chlurinatcd biphenyls are hydroxylated by come species. Apparently trout Is not able to hydroxylate 4-chlorobiphenyl, 4,4'-dichlorobiphenyl, and 2,2',5,5'-tetrachk>ro6iphenyl, while the rat and the pigeon both formed ntonohydroxychlorobiphenyi from 4-chkrrobiphettyl. In addition, the rat also produced dihydrochloroblphenyi. Large quant Mies of unchanged material were excreled by Ihe rat when 4,4'-dichlorobi phenyl and 2,2\5,5`tetrachloroblphenyl were given to them and only a monohydroxyderhrattve was identified In their urine. Pigeons followed a similar pattern but no dihydrochloroblphenyi was observed in the bird excreta fed 4-chioroblphenyl. No hydroxy metab olites could be identified in the excreta of the three species when they were given 2,2',4,4'S,5'hexadilorobiphenyl. No evidence of reductive dechlorinition was observed in eny species. Only very few studies have been conducted with radiolabeled hcxachbmphenr to study its absorption, distribution, and metaboltsm. Wit and Van Genderen914 gave rats and rabbits hexachiorophcne-C14 (2,2'-nlhylene-Ci4 bis(3,4,6Irichlorophenol). Complete recovery of radio activity in excreta after a single dose of SO or IS mg/kg of radioactive hexachloropltene was ob served In rabbits within 5 days. Roughly one third was excreted in urine and feces, respectively, as unchanged hexachlorophene end the remainder wai excreted In ihe feces as unidentified metab olites. The me laboil let could not be extracted from the feces with ethanol. The biological halflife calculated from the experiments) results was found to be approximately 17.8 hr. In rats, about two thirds of the radioactivity was recovered in feces within seven days and only very little in urine. A complete recovery of ra .inactivity was not obtained in these studies u ihin seven days. Five cows given a angle dose of 15 mg/kg body weight of hexachlorophene excreted about half to two thirds in thalr facet within five days. Very little was excreted in urine and none In milk. January 1*74 475 HONS 207663 Metabolites were m>t identified and ikmc levels were ikh determined In this study- Tire mliun idl'd a study by Kuk.1,s who did determine lii"i,ii hioii>t>tii,iii' in various organs of rats. i .imdl i-i al.,,*<* studied the absorption off-14 labeled Itexacltlcirophene on tbc tail skin of Mte rat. Hexaeltloroplienc in a delctpcnl uditiioii was absorbed by the intact skin of tbc tail at Hie average rale of 1.7 fig/vm1 of exposed surface aica/ln. Iiiimcdialdy alter tbc iiiniclioit of second dcpcc burns, tbc absorption rate increased 2.5 times iliai of lire noinisl skin surface. After 24 br die absorption rate dropped to normal or sub normal levels. Immediately after traumatizing the tail, the absorption rale of hexachlorophene was even greater than following the infliction of a bum wound. ManowiU and Johnston11' found that hexadilorophene was deposited on the skin when used in various formulations and could be extracted from Hie skin by soaking the'exposed areas (hands and forearms) in alcohol The amount recovered increased in proportion to the amount applied. These auihors were unable to confirm earlier investigations which soggested that the quantity of hcxui'hloroplieiie retained by the skin reached a plateau level after a numher of waitings and remained relatively constant thereafter. Arms were washed and rinsed every 20 min. When extractions wills alcohol were made on different arms after different numbers of washings, It was found that the amount of hexachlorophene recovered In creased with the number of washings and no plateau was reached. A greater amount of hexachloropliene was also deposited on the skin with increasing length of lime of a single washing. Since we now know that hexachlorophene Is also ab sorbed from the intact skin and some of It If probably also lost from the surface of the skin, the time interval between the washings plays a role in the amount of hexachlorophene found to accumu late on the skin. Also of interest is that bathing water containing 4 mg/I of hexachlorophene de posited approximately the same amount of the chemical on the skin as a single washing with a 2% hexachlorophene soap. Since hexachlorophene was used very exten sivcly and often unknowingly by the general population, wc investigated whclb'i it was present in the blood nf a few random samples of the general population. Wc found that the arithmetic mean concentration of hexachlorophene in adults was 0.028 ppm Oig/g) of whole blood of a total of 14 samples. In infants who were washed with hexachlorophene in the nursery the amount nr licxacliloropbcne found in whole blood after die infante hail anywhere from 2 to 14 wadies averaged 0.10*1 pptn (jig/g whole blood), blood obtained from the cord at the time of birth contained an arithmetic mean of 0.022 ppm hexaclitorophcnc.1 ` *-11 * 111 the rats that were used in a reproduction study and were fed 100 ppm hexachlorophene in (he dial for 258 days, the hexachlorophene blood levels averager) 1.21 ppm, while the blood levels of rats fed 500 ppm of hexachlorophene for 55 Jays who had severe symptoms of poisoning amounted to 8.5 ppm.1 At the time we reported these blood levels, our recovery rale was only about 75%. All blood levels reported at that time were determined in whole blood. (II Is assumed at the moment that blood levels are equivalent to half the amount of that found when plasma is ana lyzed.) A study undertaken at the request of Sterling Laboratones'11 Illustrated that newborn rhesus monkeys foOowing total body bathing with a 3% detergent solution for 1 week had blood levels of about 1.5(/g/ml (ppm). In subsequent months a plateau affect was observed and at 90 days the blood level was equivalent to 1.1 s/g/ml These monkeys showed status spongiosus or rl*- white matter of the brain, this indicates that at least in the monkey, blood levels of about 1.5 fig/ml whole blood were consistent with fluid accumulation In the myelin sheath of the central nervous system. Lockhart111 cited unpublished studies by duck, who found that hexachlorophene blood levels in term infants after washing with a 3% hexachlorophene detergent solution were some what higher than those reported by us with a mean concentration of 0.345 ppm (jig/g whole blood). This difference could have been due to a differ ence In washing procedure and a better recovery rate in the analytical method In Gluck's study, or both. - Premature infants apparently absorbed hexa- chlorophene more readily. In IS premature infants that were washed with a 3% hexachlorophene detetgent solution, the average blood concen tration amounted to 0.475 ppm Olg/g)-1*1 Abbott et al.1*1 using a different washing regimen found a mean hexachlorophene blood concentration 12 lo 18 hr ader the Initial wadi of 0.154 ppm (jig/g CPC L'l.ir*'fmr- hi 7 xrnl-rr HONS 207604 ,.f nli of he lies khJ rlh pin ion in >od . of ay! led etc pul ere the lalf iniof orn iod enl 90 ml. (lie ai 1.5 end (ral by cod 3% meican ad). Ter re ry . or exisms iene cenhoii iund in */ whole blood land oil the 7 in Oih day of life. 74 io 4S In ai'iei llic second or ilind application of Jk-m. hl.'-.-rb.-pf. :'-.r --iv M.W >v' - -- .\ -ii d : f;'": t-C > iVunJ in ivu! blood of U< inl'jim was 0.0Id p|Mlt Ipg/g;). VVlicii hexachlorophene was applied only on the day of delivery, the hexschlorophene blood levels grsdu- ally decreased from a mean of 0.145 ppm on the Tint day to t mean of 0.057 ppm on the ninth day, following an increase io 0.194 ppm on the fourth day. The authors suggested that premature infants may absorb more hexachlorophene through their skin since in one infant 29 weeks of gestation and weighing 1.42 kg the blood level was 1.17 ppm 24 hr after the initial wasli and 0.655 ppm at 40 hours. Alder et al.*** reported mein hexschlorophene blood levels of 0.18 ppm in infints who were dusted with a powder containing 033% hexachlorophene. Adults using hexachlorophene preparations also absorbed It via the skin.131 The blood levels obtained In adults that exposed their total body surface to a 3% hexachlorophene preparation twice daily for 60 days, 5 min each time, showed blood levels of approximately 0.68 ppm (pg/nil whole blood) with a range of 0.25 to 1.08 ppm Org/nil whole blood). If only hand washing was employed for a period of 28 days, 4 times daily, a man blood level of 0.07 ppm was reached and in a face hand washing study the mean blood level after 28 days was 0.196 ug/nil. Mood levels obtained in a patient with burns who died between 24 and 48 lir after the last hexachlorophene application were 2.2 ppm Og/g whole hlood). The blood levels that have been reported all indicate that following dermal hexa- chloropltcne exposure the chemical can be round in blood, and from information through animal studies as well as some human experience, levels of approximately 1 5 to 2 ppm 0g/g)in whole blood in humans are presently considered to represent toxic levels If the anslysis Tor blood was performed witii the methodi mentioned elsewhere in that article. Probably a better way of assessing a toxic doie would be to measure the amount of hexachlor ophene in the brain, but Tor obvious reiaons that is not possible. Female rats fed hexachlorophene A the rate of 500 ppm (25 mg/kg body weight/day) for 55 days with very definite lesions hid about 4 ppm hexachlorophene in their brain and 3.91 ppm in the liver. Squirrel monkeys given 5 mg/kg body weight/day of hexachloropticne for .IX day* showed i mean hexachlnmplttw ooivent ration in :lv 1- or O |'p;li and >> .',!-!w liu.ii,- th,i lUAJCtilo'Ophi'tir tOMlYlltlUtuni w;ii ,t'4 llto recovery rate in this study was only 75%. These squirrel monkeys had no overt clinical signs of toxicity, but showed mild status spongioaus of the white matter of the brain. EJcctroencephalo- graphic changes hed also been observed in these pnmstes. Ultamer et al,*** found hexachloro phene brain levels of about I ppm in newborn rats with signs of central nervous system toxicity. In I suspected case of human hcxacMwophene poisoning in a child, the hextchloropbane brain level was about 2.2 ppm.1*' Adult rats dying after the administration of a single dose of hexachloro phene diowed concentrations of 6 to 9 ppm of hexachlorophene in the brain.*** Some variation of (he concentration or hexachlorophene In the brain which can be related to neuroloxic symptom* must be expected in dilTerent species and perhaps also with age and sex. Whether the fluid accumulation which may occur in the myelin sheaths after repeated exposure to hexachloro phene would alter hexachlorophene brain levelt hit thus far not been reported. With present methods, we are probably only determining free hexachlorophene while the conju gated portion remains undetected. The toxicity of hexschlorophene will vary with the ability of the organism Io conjugate, metabolize, and excrete this material. We Tound in rats that hexachlorophene is excreted in milk of dams fed hexachlorophene at the rate of 20 and 100 ppm in the diet (2-3-1.1 mg/kg/day and 11.8-5.5 mg/kg/day, respectively, for 241 days). Electron capture, gas-liquid chroma tographic determinations revealed levels of 0.07 and 0.33 ppm In the milk. Since hexachlorophene did not seem to accumulate to a great extent similar values would probably have been obtained, had the lemls In milk in these animals been measured earlier. Although we now have some information on the absorption and distribution as well at excretion of hexachlorophene, the possible storage of the maierial as well as Its metabolism needs further investgation. Since hexachlorophene was found in cord blood of infants, it can be assumed that transplacental passage also occurred. Morphologic Effect on the Urn Some of the chlorinated naphthalenes, chlon- Imauy 1974 4T7 I MOMS 207685 dated ililico/i'dmxins. chlorinated dihcnxufiirans, (lie chlorinated biphenyls, as well n the cliiori- nalcd terphcnyis have an effect on ihe liver In various species, including human*. Bennett et al.1*1 compared the toxicity of differently chlori nated naphthalene! and a biphenyl that contained 65% chlorine. Tri-chloronaplillialent apparently was lets hepaiotoxic than Ihe higher chlorinated compounds and Ihe chlorinated diphenyl wat the most toxic or alt. Tire studies were done in ratt and the expoiure wai by feeding a* well ae Inhalation. The findings described in the liver consisted of liver necrosis, fat accumulation, and Ihe presence of hyaline bodies in the cytoplasm of the liver cells. A marked depcdl of finely divided granular yellow or yellow-brown pigment was observed in Kupifer cells, particularly in thoae animals that had been expoeed to the chlorinated biphenyls. This pigment apparently did not stain characteristically for either hemosiderin or hemofuchsin- Of course, ail livers were enlarged microscopically; in addition, theae authors described proliferative changes in occasional bile duels, parliculsrly after reeding mixtures of chlori nated naphthalene* and chlorinated biphenyls. The r.untber of bile duels was increased in certain areas and the epithelial cells lining the bile ducts showed mitotic figures. In inhalation studies with low concentrations oT PCBt which eonsitted or either a 16-hr daily exposure to average concentrations of 0.57 nig/m1, or lo 8 hr daily wilh an avenge concen tration of 0.93 mg/m1, neither group or rats appeared ill. The periods of exposure varied from 37 to 143 days. When the animals were sacrificed the livers were pale or slightly yellow and some what mottled. Microscopic examination of the liven revealed similar change! as had been observed after feeding the material, and an increased vecuolatkin of the liver cells. Micro scopic evidence of recovery was not observed in the liven or nts that had been exposed lo Ihe chlorinated diphenyl for a total of 105 days and were removed from exposure for 2 months. Similar findings were made by Miller1*1 who studied effect of a PCB which contained approximately 42% chlorine. Thit author observed varying degrees of liver damage after subcutaneous Injection as well as oral ingestion of the toxic substsnee in guinea pigs, rats, and rabbits. The liver damage consisted of fatty degeneration and atrophy of the central lobular cells. In the rat, hyaline bodies within Ihe cytoplasm or ihe hvn celii were also noted- Most of the liver damage was found in the guines pig, less in the rabbit, and least in the rat. Nfshizumi,*** who studied the effects oi i Japanese biphenyl Kanecior 400 with 40,; chlorine in monkeys and mice, described enlarge ment of liver, fatty changes, granular cytoplasm t>r the hepatocytes, Increases in Ihe site of KupTfcr cells and hepatocytes, and a btown pigment in aoine or these cells. He also studied the livers under the electron microscope and found an increase in smooth endoplasmic reticulum, vari ation in the appearance of mitochondria, and an increaae in the number of microbodies In addition, "myelin figures" were observed In the cytoplasm of hepatocytes. Vos and Koeman** observed liver necrosis in chickens that were dosed wilh European PCBs. while a Monsanto product with 60% chlorine did not produce this efTect. Increased amounts or iron were demonstrated with Perl's iron stain in the livers of all chickens exposed to the European as well as the American PCB sample. Vos and Beemi'* studied the PCB-induced lesions in ilic liver of rabbits, following dermal exposure to Arodot 1260, Phenodor.and Clophen. The microacoplc changes observed in Ihe liver included Tatty degeneration, focal necrosis, cenlrolohular live r cell atrophy, ceroid pigment in Kopfrer colk. periportal fibrosis, and cytoplasmic hyaline degen eration. The Teeding of Arcelor 5460, a chlorinated tcrphenyl, Aroclor 1254, or a chlorinated dibeurop-dioxin, not further specified in the article, led to liver hypertrophy of Sprague-Dawley rats within eight days to three weeks. Ulirastructuial aliciations were similar in the three groups and consisted of numerous multilayered, concentric membrane arriyi and proliferated smooth endo plasmic reticulum. In many instances these mem brane arrays surrounded lipid droplets.116 Rabbits given 300 mg of Aroclor 1242 and 1254 once a week for 14 weeks developed enlarged livers amt the ones given Aroclor 1254 showed midzonal necrosis of the hepatic lobules. The livers of rabbits fed Aroclor 1221 in i similar fashion did not show any histologic changes.171 We conducted studies to compare the toxicity of Aroclor 1254 and Arocior 1260 in Sherman strain rats. The acute efTect of these two mixtures on Iht liver was (tangible- If rats were given a 471 Otc Crtricwl Rfttrwt k Toxkoltmr HONS 207686 Cl u [Si 1 >% Itof et in !fS an ri an In he in Si, lid on he as nc! I IGURI 3. Section uf livci of a rai fed Arodor 1260 for 90 toys. This figure Bluttrilet a mitotic he 4'un' and a hypctihronlalK nurk-ui (snowi) HtEX 300. to : ci ty tingle dose of 10.000 mg/kg in peanut oil by dimethinomphlhilene, and piperonyl :cr stnniaili tube, and were sacrificed the next day, but oxide* 74 (o<2(-bu(oxyethoxy)efhoxy)-4,5- 111. the livcj was essentially normal microscopically. melhyienedioxy-2-propylloluene). They are identi li Doses of 1,000 ppm (72 mg/kg) of ArocJor 1260 cal to (he so-called hyaline bodies that were for 3 months produced only an increase in mitotic described in the earlier literature by Bennett et ed figures (Figure 3) and enlarged hepatocytes. When l.147 and Miller.141 ;o- groups of rats were fed dietary levels of 20, 100, Aside from these genera) changes, extensive, <0 500. and 1,000 ppm of Arodor 1260 and 20. 100, grayjih-while, Ann, glistening areas were noticed lin and 500 ppm Aroclor 1254 for 8 months, (he liver in the livers of a number of the experimental er- changes were pronounced.171 The hepalocytea animats, particularly at the high dietary levels. nd were enlarged in many of the rats of the experi Microscopic examination of these grayish-while he mental groups and lipid accumulation, foamy areas showed that the hepatic parenchyma had lo cytoplasm, and a brown pigment which stained been replaced by ^andular pate siaining epithilial rn- partially positive for hemosiderin were observed in cells that formed ducts and were surrounded by Jls the livers. The pigment waa primarily observed in proliferating fibrous tissue (Figure 5). Larger t a macrophages and Kupffer cells Indusions (Figure leiioni often had extensive fibrosis and alto nd 4) that stained slightly more eosinophilic than the contained collagen and the ducts that were formed rul surrounding cytoplasm were present in the cells. by epithilial pale staining cells were markedly or These inclusions wlih'i Ihe cytoplasm have been dilated and contained necrotic debris or mucus. lid described for many .ompounds such as mire*173 The lesion hit been classified as adenoftbrotis for (dodccachloro-c ctahydro-1,3,4-metheno-2H- the time being. 11 has been described in detail by iiy cyclobuta[cd)pentaiene), DDT174, l,l,l-(lii- Edwards and White174 who observed it in rats lan chloro-2,2-bis(p-chiorophenyl) ethane dkldrin171, that were fed butter yellow (p-dimethylamino-aEorei 1,2,3,4,10,) 0-hexachloro-6,7-epoxy-l,4.4i- benzene). These letions enlarge peripherally by the a a 5,6.7,7,ga-octahydro-) ,4-endo-ex 0-5.8- formation of new glandular structures. Hepatic January IBM 479 HONS 207667 TlCUIlE 4. (Klim or liver rnw ml M Arodw 12S4. A cinder f IncluttoM ii pnnal in the cytnRtaim of one hepe'oeyte (arrow). UcuoHy lndivMiul ceRi comoin only one of Hick formelioruti time. Toloidim blue tiein of ourefba eittbeddaU material X 300. Iittue may be trapped between the glands of (he peripheral area of adenofibrosis. The older central peril of the leiiont sometime! contain pigmented avascular collagen ized atroma with atrophic epi thelial cells lining dilated ducts that appear atropiiic and dead. The glands of actively growing adenofibrosis show atypical epithelium with large pleomorphic nuclei. The epithelium can occasion ally become stratified. Parts of the lesion may be qpite extensive, while in addition, in many areas of the same liver small foci of adenofibrosts exist simultaneously In dote relationship to portal areas. It has therefore been assumed that these lesions arise from bile ducts, but they can alto be primarily confined to the repon of the hepatic twins. The absence of bile ducts around hepatic veins led to the condusion177 that i precursor, other than bile ducts, existed from which adeno fibrosis developed. However some investigators of the lesions have assumed that It represented an atypical bile duct prollfaralion, also referred to u cholangiofibrosit.1 7* In addition to this lesion we also observed small dusters of glandular cells within areas of adenofibrosts as well as adjacent to Mood vessels that ware surrounded by reUnmMy normal appearing hepatic parenchyma with `(.lin ing characteristics of salivary gland tissue. llrcv dusters of cells very closely resembled pancreatic tissue.17* The distinction between idenofibiinss od adenocarcinoma is difficult. The glands ate quite atypical in adenofibrous but it may be helpful to remember that in adenofibrosis.. in contrast to adenocarcinoma, the irregularity observed is limited and duplicated with consider able exactness from lesion to lesion. Mucus is alwiyt present in adenofibrosis, but not always in adenocarcinoma The stroma of adenofibrosis is concentrically arranged, young and cellular a> ihr periphery and old and hyalinized towards the center of the lesion.177 Of eouwe i liver showing adenofibrosis may also contain carcinomatous lesions. So far condusive evidence is lacking m the literature (hat adenofibrosis gives rise lo carci nomas. However.it hat been round to coexist with carcinoma of the liver in rats and some hivcshgatort feel that the lesion is in fict s precursor of cancer.*** Choline deficient rats apparently, in addition to other dunpt in the liver, also develop 4*> CXC (VMrwf JteWew* in Toxicology HONS 207666 I'K-liKI' S. Section Jllwiraics an area of tdtnoflbmit of the H*er of a ral fad Aroctor I2$4. Nnif (Itc epithelial ccUt forming (mall durti. The duel* contain Cdlular debrit and ara Surrounded b) fibroin whieli can berome quite pronounced. H A T X I2S. rely Jillteic jiit: tins arc be . in lily tierIS is s in is is i lie I he ious i ihc ?;>with esli>r of in clop sdeiiofibnisis.1* ' However, llicsc rais were fed peamii niciil and soy meal and therefore rite possibiluy exists that IJvc expcnnientai diet was conuiniiuiicd willi a inyaiioxin. Dcnnct et at,1*7 also deuiihcd Ihc lesion (adcnofibrosis) in rail when a mxtuic of chlorinaicd naphthalenes and chlorodiphenyls was fed to litem. A critical review of the htsiopathopciicsis of (hit lesion was made by Siewail and Snell,177 Ulrraslructuiai changes of llie livers of animals exposed in ihc Aroclor consisted of an increase in smooth endoplasmic reticulum and atypical mitochondria. Many lipid vacuoles were observed, ptiticulaiiy at I lie lugher dietary levels. They were it nmes surrounded by concentrically arranged membranes. The "IncliBtons'' or "hyaline" bodies obierved in the cytoplasm with the light micro scope correspond to these formations when they irt examined under the electron microscope. Their uliristructurtl appearance has led to (heir being referred to as "finger prints" (Figure 6). The rpithelial component of the adenolibrosrs outlined consisted of cubotdal or columnat cells with the free surface lined by microvilli, granular cylopiaim with many ribosomes, and a few endoplasmic membranes. Some of the cells contained a great deal of mucus and resembled goblet cells. TonofiJamcnts and terminal bars were also observed. These various observations suggested that (he epithelial component of adcnofibrosis had all the morphological characteristics of bile duel epithelium. Chickens with chick edema disease and cattle with the X-disease, already mentioned, usually showed changes in the liver which consisted predominantly of M*er cell necrosis and fat ac cumulation. The hepatoloxic effect of chlorinated naphthalenes has also been described in rats.1*7 In s number of reports on the occupational occurrence of chJoracne in workers that were exposed to chlorinated naphthalenes, chlorinated diphenyls, or 2,4,5-T, and technical trichlorophenol, reference is made to the hepatotoxic effect of these compounds. This hepatotoxic effect can manifest itself only in a decreased tolerance for alcohol; it may also develop into hepatic deficiency with acute yellow atrophy and jaundice. Apparently the affect on the bvw was Jentmry I9M All MONS 207669 FIGURE 6. Electron micrograph of i portion of the cytoplasm of i hepalocyts. The Mction shows concentrically arranged arriyi of membranes which surround vacuoles coolaMng lipid Load citrate, urinyl acetate X >4,200. particularly bad following exposure to halo-waxes, which are chlorinated naphthalenes.1*1 Cotier1*1 reported seven cases of ptntachlorlnated naphthalene poisoning in workers who were engaged in manufacturing wire cable during World War H for the navy. Four of the workers develop ed jaundice and two died. Microscopic examina tion of the liver of the two workers (hat died showed complete loss of liver cells in tome areas, the oentrolobular areas were hemorrhagic, and prominent bile duct proliferation wh seen tat the periphery. Another group of compounds that have it least some representatives that cause a hepalotoxk efTeci ere the chlorinated dibenzodioxini and chlorinated dibenzofunns. Single oral doses of 1 to 10 pg/kg 2,3,7,5tetmehlorodibtniodioxin given to rabbits produc ed a hepetoioxic effect. At the higher doeage level the animals died and the lower doses caused serious liver damage.1** Schulz'*1 had reported earlier that tingle oral doles of 20-50 pg/kg body weight resulted in fatal liver necrosis in rabbits. Weanling rabbits were treated with 2,3,7,8tetrachiorodibenzo-p-dioxin by applying a dose of 1.67 pg daily to the Inner aspect of both eart for three days. The total dose given was 7.05 pg/kg body weight. When these rabbits were sacrificed 18 days following the last application or the dioxin, their hveis were significantly larger than those of the controls. Microscopic examination of the liven showed enlarged hepatocytes. Some of the hepatocytes were multinudeated. The cyto plasm was foamy or vacuolated A light brown pigment in some liver cells as well as Kupffer cells was also observed. Inclusions were seen in the cytoplasm in some liver cells, slight interstitial fibrosis was present, and two of three rabbit livers examined showed foci of necroeis that were surrounded by flbraiis.1*4 Studies with "toxic ret," the cause of chick damn disease, showed that it had a necrotizing MI CHCCrirtttl Reviews Sr Tojrlcofatr HONS 207690 A on ilu- li*ri in chitU'di. A* *. iu* lmi'. membrane* within tlie cell is iti'i known. The fact ilic toxic agcm* lexpuniaMe foi thick edcnu that smooth endoplasmic reticulum may become (jittjsr jir cliiurinalcd diben/j'-p-diozini- Toxic hypoaclive was first postulated by Hullerer el j-jn also altered liver morphology in Macacca al1"' imilaHa monkeys.1*' Hepalucyies were enlarged, Fibrosis of the liver as well is necrosis and the niulimuclcjietl and focal necrosis in <lie eeniro- adenofibiosis observed after PCB exposure In rets tubular rone was observed. Many liver cells were should definitely be considered a very serious vacuolated and stained positive foi neutral fal. lesion. Usually adenoftbroais occurs concomitantly In our studies of la's given bcxachlorophrnr. with hepatomas or htpatocarcinomas In rodent vt did utn observe any liver philology wtlh (he been. A recent publication by Khmsre and f: light microscope and no change wills the electron Baba1'*1 showed that hepatomas can be Induced k microscope ai the dosage levels that we in ran with KanecMor 400. Chronic feeding i 4 employed."* Thorpe'1* on ihe olher hand, studies may show that other KBt wfll also induce observed periportal fatty changes in liven of sheep hepatomas In rodents. pven three or four dotes of SO mg/kg hexachloro- pliene and hisiochemicat changes were also observ Neurotoxicity k.m ed in iIk liepaUicyles high and Crowley1** also In some of the reports on cMoracne and also in 1 observed hepaioioxidiy in sheep after giving them hexaclikuopiiene. Ihe Yusho incident, peripheral neuropathy is mentioned is well as psychic alterations'4** as 4 Of iIk otliei germicidei discussed in this article, part of the symptomatology of poisoning, but in only iriclosan (ligasan W-.I00) (2,4,4-trichlofO- moat reports not much emphasis eras put on It. In 2'-h)dioxydrphenyl ether) has been reported to the Yusho epidemic Mure* and Kuroiwa1** per have a toxic effect on the liver at a dose of 125 formed more detailed examinations in 21 cases mg/kg and enlarged Ilie liver of rats ai a dose of 25 admitted consecutively lo Kuyushu University I J1 mg/kg. according > lire summary minutes of Ihe OTC Panel, U-S. Food and Drug Adminisiraiion. It Hospital in the northern part of Japan. Seven of ihe patients complained of numbness or pain in war not staled wlwtiier these were single or ihe peripheral portion of their axtremitles and In IV repeated doses. Oral administration of Iridotan to five cases, hypoestheaia, hypalgesia. and thermo beagle dogs produced Jaundice and severe liver hypesthesia were noted. Slowing as well is reduc damage was observed. The effect that Iheae com tion of Ihe eenaory nerve conduction velocity m y pounds have on the Irvet will have to be studied Ihe redial and aural nerve was observed in a further. / numbei of iheae patients. Headache and peripheral The accumulation uf fat in Ihe live' indicates nervous system symptoms were also reported In e r r that cither fal transport or fal metabolism is impaired; if (he insult ro Ihe liver is remewed, the liver gradually recovers sinless the lesion has poisoning episode which occurred in a Finnish company where paper was impregnated wtlh biphenyl.*1 t J become irreversible. The increase in smooth endo The only chemical of the group of polychlorin t plasmic reticulum as well as ihe "inelusions" will ated, polycydic compounds ditcuued in this i also disappear after a certain amount of lime if report thei has a predominant central neurotoxic r exposuie lo the material Is discontinued.1*1 The effect in animals as well as humns is hexachloro- r significance of the increase in smooih endoplasmic phene. Aduli female ran that were fed 500 ppm iruculum and the hyaline bodies with a concomit hexachlorophene in the diet for 14 weeks (about i ant rise in liver itucroaomal enzyme activity is 30 to IS mg/kg body weight/day) developed leg i present!)' not understood. These changes are con weakness efter 12 to 19 days of exposure. This leg sidered adaptive and beneficial, but in eddition to weakness which was only evident in the hindlegs i metabolizing unwanted chemicals ai a higher rate, progressed to paralysis wilhtn 3 to 5 weeks.1* ' At t the stimulated lives also metabolize] some medica autopsy the brains of the exposed rets were tions more rapidly,1** as well as aubstances enlarged; they weighed an average of 2.6 g while produced by the body such as steroids. Whether Ihe conirul breirn weighed an avenge of 1.99 g. the inclusions within the cytoplasm represent the Microscopic examination of the brains revealed so-called inactive smooih endopiaunic reticulum normal gray matter throughout the brain and the which is deposited in Ihe form of arrays of spinal cord in the few butancea that tha spinal Jamwnr 1934 4U HONS 207691 riGURC 7. Electron micrograph front the while metier of (he brain of tel. Mluslratina status iponfKMui Note the l..r-, empty appurinp rmolei lined by myelin (MY myeUn, A * non), lent citrate, tin nrI icetete X 34.200 cord was examined. The while miller on the contrary teemed widened and showed many cystic spaces thu appeared empty and were lined with sirands of male rial Ihal stained poti live for myelin. The vacuoles were very numerous and varied greatly in size giving the tissue a spongy appearance. The brains in these studies were usually fixed in either Bourn's solution or buffered formalin for light microscopic Study- Brain tissue, particularly that of rodents, may normally show small vacuoles that are considered lo represent an artefact.1*1 These vacuoles are smaller, usually very uniform, and at times contain a homogeneous pink staining material. They are usually separated by a wider area or normal appearing white matter than the vacuoles in stilus spongiosus. Brains that show this artefact do not weigh significantly more than those that do not show it. Electron microscopic examination of brains of rats given hexachiorophene showed tint the vacuolation in the white matter of the brain revealed large vacuoles lined by myelin. The aM>\ tli.it were present appeared usually to be qunc normal, occasionally strands of myeltn had separated and . traversed the vacuoles. The vacuoles were other wise empty except for some electron dense gtanular material (Figure 7). No other alterations were observed in the white matter. This lesion is mu unique for hexachiorophene. other chemicals that have produced status spongiosus as we have called the lesion include triethyltin.1*1 isoniazid,1** the halogenaled bcnzanilide, 2-acetoxy-4>chiorn-1 5diiodobenzanilide,1*1 and rafoxanide.1winch is the 3,$-diiodo-3'-chJoro-4'-(p-chloro-pttciiii\y) salicytsnilide. Trieihyltin seemed to produce the lesion at a much lower dietary level and nunc rapidly than the other chemicals mentioned, jvnticularly hexachiorophene. In an additional study, after feeding rats 500 ppm hexachiorophene in the diet for 10 weeks and then discontinuing the exposure to hcxachloim phene, function began to return in the hind quarters of paralysed rati after 2 weeks and wulun 444 atCOMcaf XrWrwr h Toiicotqy HONS 207692 ti wi-ikt. rlie ruts hull ulmoxl coinpkiely rtciivercJ tiinu:jlly. However. microscopic examination of ilie Duini of these rail ihowed that a Tew cystic ipaces were still present 12 weeks after the exposure to Itexacldorophcpc had been discon tinued. In weanling rats, but not in adult rats, the bn in lesion and symptoms of paralysis could be pioduced with a single oral dose of 100 mg/kf of body weigh I pven by sionuch tube in peanut od.'1* Part of the reason for being able to produce tire brain lesion in the weanling rats with a single dose may be related to the fact that the weanling was able to survive a higher single dose than the adult tal. Olhei species that have also shown paralysis ind other neurological deficits after exposure to hexachlorophene are rabbits,1'1 pigs.1*" and cats.1" Sheep become blind according to (Jdall end Malone.'16 Hamg el al.1" recorded the neuiolopcai deficits produced in cats in more detail. Early symptoms consisted of lassitude, weakness and ataxia of the hindlegs, impaired righting reflex, patellar hyperrefiexia which later developed into Ityporeflexia. urinary retention, and eventually complete flaccid paralysis. The annul nerve functions remained intact except picin initially. In these cals an increase in cysternal ccirluulspiiiul fluid pressure was observed. In our first studies we continued exposure lo liexjcliloroplieiie lor a period of 14 weeks because we had designed ilie experiment to conduct an approximate 00-day feeding study. It is noi necessary lo feed rats for this length of time. As we determined later, microscopic changes can be observed in rals after a feeding period of two weeks when the animals were exposed to levels high enough to produce the lesion. Some vacuolaion can already be observed 3 to 4 days after onset of exposure to hexachlorophene in some rais. When lower dietary levels were fed In a reproduction study.'1 * the dietary level of 20 ppnt (2 4-0.9 mg/kg body weight/day) had no effect on the brain. At the dietary level of 100 ppm (11.8-4.6 mg/kg body weight/day) focal areas of vacuulalion of the while matter of the brain were observed in some of the rats, but they were never extensive. The rats in these lower dietary levels did not show any clinical neurologi cal symptoms. The brain lesion observed in rats was also described in monkeys following Ihe exposure lo hexachlorophene. Newborn monkeys washed with 3% hexachlorophene detergent mluiimi developed a diffuse status spongiusus of the while matter of the brain.,0 Santolucito1"J#, was able to produce Ihe vacuoiation of the white nutter, the so-called status spongiusus, by infecting squirrel monkeys daily subcutaneously with 5 mg hex achlorophene/kg of body weight for 37 days. When monkeys were fed hexachlorophene they did not develop the brain lesion but showed electroencephalographic changes which need further investigation. The monkeys that showed the vacuoiation also demonstrated eleclroenccphalographk changes. Sheep have apparently alto shown status spongiosus of the white matter according to hale and Reid.101 Lampert at al.1*1 were able to produce ihe lesion in mice as well as rats. These authors reported that Ihe morphological changes were also observed In frozen recti ons of brains quenched in liquid nitrogen. Trypan blue injected IntraperUoneally did not slain the brain, an Indica tion that the biood brain barrier was intact as far as trypan blue was concerned. The brain changes consistent with stilus spongi osus due lo hexachlorophene exposure have also been described in humans, particularly children. MuJIick'10 examined the brains of six human cases that died from hexachlorophene poisoning. The two adults in this group did not show vacuoiation of the while matter of the brain. They had died less than 48 hr after exposure to hexachlorophene. The four children (two with congenital ichthyosis, (wo with burns) had at least three or more days of exposure to hexachloro phene baths in which a 3% hexachlorophene preparation was used. The final concentration of hexachlorophene in the bath water was not known. AD four children showed status spongiosus of the white matter. The brain weight given in one of the cares, a 12-day-old child, was 425 g. The normal brain weight for a child that age is 382 g. Shuman et al.1*4 In a retrospective blind study were able to demonstrate that Ihe incidence of status spongiosus in the brain stem reticular formation in infants who had died from a variety of causes could be correlated to hexachlorophene exposure. The authors found 21 cases in a series of 250 autopsies of children. Of these cases, 18 were premature infants weighing less than 1,400 g and were of less than 30 weeks* gestation. All but 2 of the cases had 3 or more total wishes with 3% hexachlorophene. In the group of cares that were studied were a number of stlllborns who did not JiMwy I9T4 485 NONS 207693 vli,. Ilu- loxui jml of KiurM- had mtt had dermal I,.Aul.i.Miii'tiif'-- cmhimiic Shuman ei al. , ................. cmI*. .* i* Lclierer Siwe * dise.se ,u wI.kIi alley had observed stem spungiosos of ilii* (,)!. .tuner. E*h r ,,K ehi,drcn wi,h j | iciei Siwc a disease had had pronounced expo ! MMC in Iwxachloropliene became of the akin I Jcmuiis which had been vigorously scrubbed with a j detergent containing the dtemical. The various i vhtcivaimna show that. in paiienls with abnormal skin as well as the skin of the premature, hexach Ini o phene may be absorbed in sufficient amounts to cause status spimgiosus. The fact that blood levels in premature infants washed with hrxacldnmphcne lend to be higher than in mature infants also substantiates Iheae findings. Almost identical observations were made by Powell et al.'** In premature infants weighing less than 1.400 g, 7 of 13 that hed 4 or more exposures to hcXaclilorophene wadies showed status spongiosiis of tiic myelinated tracts of the brain stem. If this group was further subdivided according to the number of exposures, 6 of 9 in fan is that had 9 or moie exposures to hexacldoropliene washes showed the lesion, and I of 4 that was washed 4 to ft times. Four stillboms and 1.3 infants washed 0 to 3 times with hexschloropliene and weighing less than 1,400 g did not show the spongy change. Electron microscopic examina tion sliowed that the spongy change observed In the premature infants was due to a split in the myelin sheaths. in August of 1972 the press reported a number of deaths of infinii in France that occurred because a baby powder to which hexachJorophene had accidentally been added al a concentration of 6% was used on the infants. The final death toll amounted lo 41 infants and young children114 (New York News, Dec. 13, 1972). Detailed accounts have not been published of this incident because of ItKgalion problems. As we pointed out earlier, under acute toxicity, the higher the con centration of hexachlorophene it in a preparation applied to (he skin, the more likely it is that skin damage will result. Skin damage also occurred in liic French Incident, tn addition to this, the powder was probably used in many instances where some diaper rash already existed and was then well covered with a diaper, which may have contributed lo increaaed absorption in this area. IThe brains and spinal cord of a few infants from the French poisoning outbreak that were (St CXC Crttkil Affirm lx Toxicviatj -examined microscopically showed status spongio*us of Ihe while matter.14 These various accounts show that status spungiostti can alto be produced in humans and probably represents a nonspecific reaction which can be elicited by a number of chemicals. The lesion status spongiosus of the while matter as far as we now know teems lo represent a Specific or restricted type of brain edema where (he fluid accumulation it confined to the inside of the myelin sheath wtd myelin is not actually damaged or al least breakdown products of myelin have not been observed, The term status spungkrsut or spongy change has been used as a descriptive term and a spengy change of the gray matter may also occur in certain diseases. Status spongtoeus together with other morphological changes can be observed in the brain in a variety of many etlologically different diseases and the reader Is referred to Klatro,1* Ardonato and Lamped,1* and a recent editorial in (he British Medical Journal.1* From clinically observed poisoning caaes as well as animal experiments it is evident that the brain lesion is to great extent reversible. However, in our own studies as well as those by Lamped et al.,1*1 microscopic changes In rodents did no1, completely regrets and the fact that sheep do not recover I heir eyesight after hexachlorophene poisoning'1* indicates that repair may nol always be complete. No reasonable explanilion exists at the moment to indicate why certain chemicals produce this very specific edema confined lo certain structures of the central nervous system. * ! . ! i : , | Gastrointestinal Lemons Only (wo types of compounds under discussion have produced lesions in the gastrointestinal tract. We observed in our ttudy with Sherman strain rats that high tingle oral doses of 3,000 mg/kg Aroclor 1254 and 1260 or more caused ulceration of the gastric and duodenal mucosa (Figure 8). Ulcers in the Gl tract were not observed in the long-term feeding Studies with 500 ppm or less. No other changes in the gastrointestinal tract were observed. Ringer et si,1,1 in their reproduction study with mink, observed that Aroclo' >254 produced liemorrhage in the gist roin It j Inal trad of the offipring. Allen and Norback11* observed hyperplasia and dytpbala of the gastric mucosa in rhesus monkeys. Six of these rhesus monkeys were fed HONS 207694 i lGURl: S. Stciion or the glandut*r portion of the stomach of a rat (Inn a cinfla dot* oT 3,000 nis/Wf of Atoclor 1254. Thii figure illustrates the Focal lott of mens and lha PflHralKMi by inflammatory round calli. H A E X SO. 300 ppm Arodor 1248 and six were given 5,000 ppm Aroclor 5460, a polychlorinated triphenyl. The glandular fnrmationi in (hit letion were highly atypical; peitelration of the muacutirii mucosae and invasion of the tubmucosa by the mucosal epithelium were observed. AUen and CarMent'1' reported an effect of tonic fat on the gastric mucosa in 18 of 27 monkeys, Afececa muJerre, which consisted of hypertrophy of the gastric mucosa and small gastric ulcers within the mucosa. The toxic fat used in these studies, at we now know, contained chlorinated dibenzodioxins. It has also been observed that hemorrhaging occurred into the gastrointestinal tract of rat fetuses who were exposed to 23,74-tetrachlorodibenzo-o-dioxin in utero. It was not stated in the reports whether the gastrointestinal hemorrhages were caused by ulceration of the mucosa.11 *'11 Ulceration of the glandular portion of the adult rat stomach was also observed.113 Patients with chloracne frequently complained of loss of appetite, nausea, vomiting, and abdom inal pain, and Goldmann*11 deacribed a worker with a follicular dermatitis who died six months Her exposure to 23,7,8-tetrachlorod ibenzodioxin. At autopsy a ruptured duodenal ulcer and a gastric ulcer were found. Whether any relation ship existed between the ulcers and the exposure to the poisonous chemical was not mentioned in the paper. It should be established If these gastric lesions alio occur at lower dietary levels and whether the contamination of the chemical* with various chlor inated dibenzodioxins and furans h responsible for (hem. Valuable information might be obtained from retrospective studies of workers following occupational exposure. Whether the ulcers are caused by direct effect of the chemical on the mucosa of the gastrointestinal tract or whether stress plays a primary role in their etiology similar to ulcers that may occur in burned patients Tor instance (Curling's ulcer) is not clear. COMMENT J The acute toxicity of compounds such as the chlorinated biphenyls, the chlorinated naphthabnea, and the chlorinated terphenyb k vary low. On the other h:iml. they arc jnwfly broken down in ihc environment and lie metabolism of the biphenyls with more than four chlorine atoms seems |o be negligible. Because or these character istics, the use, wherever possible, of chlorinated biphenyis that consist of mixtures of compounds with one to four chlorines has (seen proposed. Neil her the chlorinated naphthalenes nor the chlorinated letphenyls have so far been shown to present a pollution problem. Whether ihc various disease entities reviewed in this article are poduced by all the different products mentioned in connection with them is not established with certainly. For instance, lome of ihe compounds thai produce chlorecne nr chick edema have been shown 10 be contaminated with various dtlorinaied dibenaodioxini and cMorhtaied dibenaofurans. The possibility remains that the compounds illustrated In Figure I are contaminaied with these chemicals and the products themselves are not really responsible for the observed symptoms. On the other hand, a number of structurally unrelated compounds may produce ihe same disease entity. Some of the chlorinated dibenzodioxins and the dtlorinaied dibenzofurans are highly toxic. The most toxic is 2,3,7,8-tetrachiorodlbento. dioxins, while the dibenaodioxini without any chlorine and those with eight chlorines are much less toxic. The difference in toxidty of these compounds may be partly caused by the fact that they are very poorly dissolved and absorbed. Intravenous toxicity data are thus far not available on most of these compounds. We also do not know at present if the porphyria which develops after prolonged exposure to a number of technical compounds such as hexichlorobenzene, chlorinated biphenyls, chlorinated dibenzodioxln, and 2,4,$-T is in many Instances due to Ihe contamination of these products with chlorinated dibenzodioxins; of course it may be possible that one or more chlorinated dibenxofursns abo produce porphyria. It has also not been established whether all compounds that produce porphyria in animab will do so In humans as well. In general, the exposure of the various popu lation groups to chlorinated biphenyls and oti a cyciic chlorinated hydrocaibons is at too 1 # a range to induce porphyria. Whether thb can also he said for occupational exposure is not known. When production conditions and general hygiane of the workers are poor, porphyria cutanea tarda can apparently occur? It is also not known if tile combined effect of different porphyria inducing agents is cumulative. Whether the photosensitivity observed in humans with some of the germicides discussed in this paper b related to an effect nn the porphyrin metabolism was not evident from the published literature. So far the mechanism that causes chick edema, chlorecne, X-diiease, and liver disease has not been satisfactorily demonstrated. Chick edema disease has been thought to be caused by proliferation or the vaacubr endothelium. This might explain iIk fluid accumulation obearved in diseased smmals, but so Tar the sequence of events that led to this disease b not. really dear and factors other thin Ihe ambarresunent of die vescuiar system may ptay a tote in its etiology such as a toxic myocarditis, decrease in serum proteins, and renal failure. More detailed study of the pathology of the various disease entities Is needed for a better understanding of the development of the various lesions mentioned. The chlorinated polycyclic hydrocarbons seem to alter the immune response of certain animal apectes and perhaps also of people. These very important findings need to be pursued further. Thus far it has not been established beyond doubi whether this is caused by a general debilii ating elTect on the animal of which atrophy of the lymphoendothelul system may be one manifes tation, or whether thb b a specific alteration, as we know It from the effects of radiation and exposure to certain alkylating agents. In this respect not enough attention has been paid 10 ilic function of the adrenal, and several of the observed symptoms In humans such as the by per pigmentation suggest hypofunction of the ad renal cortex. Depending on the information obtained from these studies, a tolerance level of 3 ppm in some Food products as set by the U.S. Food and Drug Adtninbtration may have to be revised particularly since no long-term low level feeding study has been reported in animab. Many question) have recently been raised about the use of germicides in various toiletries, common soaps, etc. The objection to their use m the specific cue of hexachlorophene is its neuroioxic effect, that can also occur in humans, particularly if Ihe akin b defective in tome way. Of course, limited localized expoeure, such as handwashing or treatment of a small wound, will not produce neurotoxicity. 4tt CHC 001*1 JteWews to Tutkolegy MOMS 207696 Iii nurseries in hospitals. one of the gic.iicM problems ii ihe nl threat of a (U|)hyloaK:ais infccixui of epidemic proportions. In order lo prevent outbreaks of tUptiylocaiciK infectious. i routine was developed in many linspmlt in the U.S. and elsewhere in the early I'H.O's where infants were washed once daily with 2 ,n- hexaclihiroptiene itcicrgciit solution, and in add it ion the diaper area was washed several times a day Tlie detergent was soWnurnily rinsed olT tiiore or leu vigorously, depending upon tlie pliiloaupliy of (lie different nurses. In tome hot pitils the detergent was directly applied to the infant. In others it was firm diluted inwaler. Since (lie lie urotoxic effccl of hexacliiomphene lias become known, many hospitals have changed I heir policies on the use of hcxachloroplienc. Another drawback |o this routine, in addilion lo a possible neumioxic effect, is that hexachioropliene keeps not only the staphylococcus of certain phage types from growing on the skin, bul reduces all Cram-positive bacteria.111 which is highly unde sirable and may lead to disease. Recently Evans el at.111 have pointed out that a marked annual variation exists in the prevalence of several bac teria in tlie uares as well as lire umbilical cord of newborns. A decline of st a pitylococcus aureus was observed in the fall; the prevalence of Etcltcricha coll and streptococcus also varied. Staphylococcus epidermis and Entcrobacler organisms did not show this fluctuation. Any studies of efficacy undertaken in a clinical selling should lake these normal fluctuations into account and it is possible that good general housekeeping practices are ultimately more effective in preventing bacterial disease in newborns than various germicides. Unfortunately, breaks in technic do occur which in many hospitals makes the use of germicides on infanls desirable. The in forma I ion reviewed in this ariick shows lltal lit* toxic affects of the polycyclic poiyctilnr. ittaied compounds are complex. The mere know ledge of an LDfs value In an animat species does not give us a great deal of information about the toxicity of a compound. Toxicity or toxic effects are a relative concept and such household hems as able alt. vitamins, pepper, and allyl propyl disulfide voliiHIzing from onions to name a few are also toxic. Nevertheless, we have to learn not to use chemicals indiscriminately, without knowing i great deal about them, particularly their long-range effects, when they are persistent and poorly metabolized. The Indiscriminate use of these products and discharge into the environment Invariably lead lo Iheir restriction or ban, whidi is accompanied by a great deal of emotionalism, dial orled facia, and often great economic toss. Prudence, better judgment, and a critical approach to ihe usage of chemicals and drugs, particularly an evaluation of whether in specific situations lltey are necessary or not. may help us avoid these situations in the future. Particular attention should be paid to avoiding unnecessary exposure of children to chemicals that we know little about. Children may react differently from adults and as far as long-term exposure Is concerned, they would be Ihe most vulnerable. ACKNOWLEDGMENT My sincere thanks go to my colleagues and former co-workers who have made this review passible, particularly to Drt. R. E. Zehr, J. E. Suggs, V. W. Burse, R. E. Jennings, R. E. Linder, V. E. Sedlak, J. A. Goldstein. E. C- Vlllaneueva. and T. B. Gaines for letting me incorporate some of the unpublished results Into this manuscript. REFERENCES I Poland, A- P.. Smith, D-, Metiti, G., aad fomfck. P., A health survey or workers in 2,4-D and 2.4.J-T plant. ,4red tnnmn. HctUH. 22.316, I97t. 2. BleibWf. J., Wallen, M,, bodkin, H, and Agpiataam, T, *_ InduilrMly aoqexed porphyria, Arch Dtrmuei.. (9, 793, 1964. 3 Laraoa, D. L, Studies show hcxachlorophrnt causes bs .n syndrome,/ Am. Hotp. Anoc. 42, 63, 1961. 4. Schmidt, It. aad Schuli. 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