Document ykqV6ojNYzM1jQDgeV6ByBv7d

SEPTEMBERlOCTOBER 1981 Y VOLUME 36 NUMBER 5 . -- Agricultural Chemical Use and Congenital C l e f t Lip and/or Palate Cadmium and Lead Content of Maternal and Newborn Hair: Relationship to Parity, 3ir:h M'cizht, and Hypertension A Wire Reclamation Incinerator as a Source of Environmental Contamination with Tetrachlorodibenzo-pDioxinsand Tetrachlorodibenzofurans Mortality among Welders, Includinga Group Exposed to Nickel Oxides The University of Akron Study on Air Pollution and Human Health Effects. 1. Methodology, Baseline Data, and Aerometrics The University of Akron Study on Air Pollution and Human Health Effects. 11. Effects on Acute Respiratory Illness Percutaneous Penetration of Benzene and Benzene Contained in Solvents Used in the Rubber Industry Effect of Hydrogen Sulfide on Bacteriai Inactivation in the Rat Lung Brief Communication Volcanoes and Carcinoma of the Thyroid: A Possible Association jane E Gordon Carl M. Shy Guy Hue1 Claude Boudene hlichel A. Ibrahim Daniel 0. Hryhorczuk William A. Withrow Carolyn S.Hesse Val R. Beasley Anthony P. Polednak 21 3 227 228 235 Richard A. Mostardi, Daniel L Ely Nancy R Woebkenberg Barry Richardson, Marcia Jarrett 243 Richard A. Mostardi Nancy R. Woebkenberg, Daniel L E& Mary Conlon, Glenn Atwood 250 Howard I. Maibach Demis Mark An;o 256 Robenf. Rogers jumj Ferin 261 Tak-Min Kung Wingding Ng, james B. Gibson 265 ISSN 0003-9896 Percutaneous Penetration of Benzene and Benzene Contained in Solvents Used in the Rubber Industry HOWARD 1. MAIBACH, M.D. DENNIS MARK ANJO, M.S. University of California Medical Center Department of Dermatology San Francisco, California 94143 ABSTRACT. Penetration of benzene through the skin of the rhesus monkey was determined using 14C-benzene,and quantitatingthe labelled metabolites in urine. The modes of application and amounts of benzene that penetrated the skin (indicated in parentheses) are as follows: (1) a single, direct cutaneous application of liquid benzene (0.172 i 0.139%); (2) a single application of benzenetontaining [036%] solvent (0.0805 i 0.0306%); (3) multiple washes with full-strength benzene (0.848 i 0.0806%); (4) multiple washes with the benzenetontaining [035%j solvent (0.431 i 0.258%); (5)removal of the stratum corneum followed by application o f full-strength benzene (0.909 :0.627%); and (6)application of benzene to the palmar surface (0.651 2 0.482%). Until more complete human data becomes available, benzene penetration in the monkey may be used to estimate penetration in man, both for industrial hygiene ' purposes and general toxicological use. ALTHOUGH BENZENE i s a ubiquitous substance to which most of the population i s exposed daily,' there has been little direct experimental data regarding benzene absorption through the skin. This i s of practical importance since benzene i s present, for example, in small amounts in gasoline (up to 2-3%)and in minute quantitics in water sup- plies.' Various amounts of benzene are present in solvents and mixtures used in manufacturingand other industrial processes; therefore, employees may have skin contact with 25 6 solvents or other mixtures containing benzene in trace amounts, i.e., less than 0.5% by volume. The data contained in studies conducted prior to 1962 investigatingbenzene absorption via the skin are summarized in the DISCUSSION section. Early data do not provide a sufficient factual basis for determining to what extent, if any, benzene is absorbed through skin-either at full strength (loo%),or where present in a small amount in a solvent or other liquid mixture. T h i s report describes skin absorption of full-suength ben- zene and benzene which is present in trace amounts in solvents used in the rubber manuf::turing industry. The purpose of the study was to determne the extent of benzene percutaneous penetration, if any, using full-strength benzene and a rubber solvent containingsmall amounts of benzene. MATERIALS AND METHODS The experimental details are the same as those described by Feldmann and Maiba~h.T~hree rhesus monkeys were studied for each experimental variable. The rhesus monkey was selected for these experiments because it has been shown to.& a reasonable model for percutaneousabsorption where relevance to man i s sought4 While percutaneous penetration i s a complex process, it appears from data derived from research with compounds other than benzene that absorption i n the rhesus monkey bears a marked similarity to absorption in man. Parenteral administration of benzene. A 1.1 5 pg (3.7->3 pCi) dose of "C-benzene was dissolved in propylene glyco! and injected into the thigh of each rhesus monkey. Archives of Environmental HeJlth g .. fianmus administration of benzene. A 50UI(43.95 mg) do= of benzene with an activity of 4.33 Pci W a @pliedto a 12.9cm2 (3.41 mg/cm2) area of the forearm or palm of each monkey and allowed to dry. The treated ym was kept isolated for 4 hr after application of benzene. Duringall cutaneous applications of benzene the mon- keys were transferred outsidc thc Vivarium to minimize inblation and to preserve the controlled atmosphere of the bboratory; all monkeys wore respirators during the appljcabon. One to 3 days prio; to benzene application, the fore- were lightly clipped with an electric clipper. It should be noted that a previous experiment with rhesus monkeys indicated that t h i s procedure does not grossly alter pene- tration.' All monkeys used in this study were trained to s i t in metabolic chairs. The monkeys were sedated with ketamine hydrochloride and appropriately dosed (see above). They were kept in the metabolic chair and were fed food and fruit (each havinga high water content) and water ad libi- dum. Percutaneousadministration of benzene in a rubber sol- bent. X petrolleum distillate mixture contlining 0.26% benzene (15.7 pCii50 pl dose) was preparea. The 50 pI dose was applied over a 12.9-cm2 (12.3 p&lcm2) area and was allowed to evaporate. T h e rubber solvent-a petrolleum distillate-was a mixture of aliphatic and aromatic hydrocarbons with a boiling point range of 104' to 266OF. The major components of this mixture are C4-C, aliphatic hydrocarbons, including butane, isopentane, pentane, hexane, cyclopentane, and heptane. The aromatic portion of the mixture contains benzene, toluene, and xylenes as major components. Palmar exposure to benzene in a rubber solvent I4C-ben- zcne was dissolved in a rubber solvent containing 0.35% benzene to a concentration o f 5.92 pCV50 PI. The solution w a made from a benzene lot with an activity of 54 mCi/mmol; 8.53 pg benzene was added to the solvent containing 158 pg of benzene. This addition, however, did not significantly increase the benzene concentration. Fifty mi of the solvent containing wC-labelled benzene were applied to 12.9 cm2 (12.3 pg/cm2) of the palm and allowed to dry. Exposure of "damaged skin" to full-strength (100%) benzene. An area of the forearm skin was repeatedly stripped with cellophane tape until it glistened. This process removed the stmum corneum to simulate damaged skin. Fifty microliters of "C-labelled benzene (43.9 rng; radioactive dose of 4.25 pCi (4.31 mg/cm2)] were applied to t h e stripped skin m a and allowed to dry. Multiple exposures to full-strength (100%) benzene. One hundred microliters of reagent grade benzene (unlabelled) were applied to a 12.9cm2 area of the monkey's forearm on 10 separate occasions at 15-min intervals during a 2.5-hr period. A total of 68.1 mg/cm2 benzene was applied. Fifteen minutes following the last application, 50 pI of 14Clabelled benzene [43.9 mg; radioactive dose of 4.94 pCi (3.41 mg/cm2)] were applied to the same site and allowed to dry. The application of the "C-labelled benzene was always performed in a ventilated passage open to the outside air with the monkeys wearing respirators. Multiple exposures to benzene in a rubber solvent A r u b her solvent containing 0.35% benzene was utilized, the char- acteristics of which are described above. UC-benzene was dissclved in the rubber solvent (0.35% benzene) to a concentration of 5.92 pCi/ml. T h i s increased the initial amount of benzene (3.16 mg) by 8.52 pg but not significantly (J4C- benzene activity = 54 mCi/mmol). T h e rubber solvent con- taining ''C-labelled benzene WY applied to the monkey's farearm in 100 ul (1.29 cm2) doses on 10 separate occas- ions at 10-min intervals. A total of 5.92 pCi was applied (68.1 mg/cm2 benzene). The application was done in a ventilated passage open to the air with the monkeys wearing respirators. Urine and fecal analysis. Urine and feces were collected for assay of I4C to determine the completeness of excretion and to identify the kinetics involved. Each aliquot of urine was oxidized with chromic acid; the 14C-C0, released was captured and counted. The feces were diluted with water, aliquoted, and oxidated in a manner similar to that of urine.3 Data analysis was performed on a programmable microcomputer. RESULTS Parenteral administration of benzene. The average excretion was 41-04 f 9.41 % o f the injected dose, with individual values of 47.04%, 45.84%, and 30.1 8% (Table 1). Parenteral excretion data were utilized in preparing the correction figure for topical experiments, i.e., to correct to incomplete urinary excretion. No significant counts were found in t h e feces. Cutaneous administration of benzene. The penetration values were 0.1 06%, 0.1 221, and 0.288% of the applied benzene doses, with an average of 0.1 72 f 0.1 39% [standard deviation (SO)] (Table 1). This utilized the correction factor of 2.44, as determined in the parenteral study of benzene. No significant counts were found in the feces. All standard deviation values include the uncertainty contribution of the parenteral injection. Percutaneous administration of benzene in a rubber solvent Usingt h e parenteral correction factor of 2.44, the penetration was 0.0805 +, 0.0306% of the benzene applied dose, or 0.00029% benzene in relation to the solvent applied (Table 1). Palmar exposure to benzene in a rubber solvent Using the parenteral correction factor of 244to correct for incom- plete excretion, the 14C-benzeneand metabolites detected in the urine was 0.651 k 0.482% (N= 3) of the benzene portion (0.35%) of the applied dose, or 0.00208% benzene in relation to the solvent applied (Table 1). Exposure of "damaged skin" to full-strength (100%)benzene. Penetration, corrected for incomplete excretion using the parenteral correction factor of 2.44, was 0.6023%, 1.3868%, and 0.7393% o f the applied dose, or an average of 0.909 k 0.627% o f the applied dose (Table 1). Multiple exposures KO full-strength (100%) benzene. Urinary excretion of the ''C-labelled benzene, corrected for incomp!ete excretion, was an average of 0.838 2 0.806% (N = 3) of the applied dose (Table 1). Multiple exposures to benzene in a rubber solvent. Penetration of the benzene portion of the appticd dose, correc- red for incomplete excretion, was 0.431 -C 0.258% (,V = 3) of the benzene portion (0.35%) of the applied dose, or SePtember/Octobcr 1981 [Vol. 36.(No. 511 251 I 1 ,; I !; , .. I I- I ! ,; i: iT W - -v 9* 0m f-. 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' iIS(Z\J !;!<I:)I4 Full s[rt:rgt 1 '*C:.b~nrefif:ws iniect4.d ir to rl-es s m o r . * eys and i t s lpr :senc:i: wa irlmtified uuantitiitively ;.id kinetally ttirotigi mdysi: 01 the ur ne anc feces. Radio.ictive : Znzerii: r~i1.s pllea t:, t:e forearm of the rnoni:t:ys to c p m u t ( : aisor ;3,:i( n. U s r g ittc correction factor of ;!.44 deterI ,rned i I:he p,;renteriil study the pent:tration was 0.1 7 2 : 0.1 00%o f ch: 100%berrzene ;idministered. ''C-be.nzene, a c:oniporien of a r u b t w solvent containing 0.365b e i 'ne, vd 1s snpli :d togical k t t o the forearm of thc m n k e fs. sing 1.t.e paler teral cc:rrt:rtioii factor o f 2.44, ttic: ac tu3. :nei:ra' ion wa 0.085 .t':.O:Cl6% of the benzent: portion ::.36%1o) f the ipplied dme, or 0.00029%benzene in re1.a- i in lo 1he s c ~ l v ~a~pnptiie 2. T h e 0.1 72%and O.GSO% Dent:[ * x i o niigures suggesc t h i t wnen similar amounts of benrrne .m presenred in diluted and concentrated forms, p r a ?cirtiorrally less benzene rnay be absorbed in the diluted Tcrm. In the multiple exposure experiment, 100 91 of full;!:-ength (100%)benzene (unlabelled) was applied to the * f i x t imonkey's forearm on 10 occasions at 15-min intera i s , followed by a 50 1 close of '*C-benzene. The penetrai o n of fiat eleventh dose was 0.848%-approxirnately 4.5 bmes the 0.172% p e n e m t i o n reported in the single exposure study. The single and multiple exposure results cannot be com3ared in a linear retationship, because in the multiple exposire experiment the first 10 doses were 100pI each, rather h a n 50 PI. T h e larger doses were to approximate the effects If chronic exposure to full-strength benzene. It was thought h a t t h e use of 100pl doses of full-strength benzene might lave a more deleterious effect on the skin than 10 similar ioses in 50 pi amounts. T h e results of the stripped skin experiment using full- itrength benzene suggest that the multiple exposure experment may approximate benzene's chronic effects. After the stratum corneum layer of the forearm skin had been itripped using cellophane tape, the penetrxion was 0.909%, Hhich is in a close range to the 0.848%actual penerration 'eported in the multiple exposure study. In the multiple exposure experiment using rubber sollent labelled with 14C-benzene, 10 separate 100pl doses were applied at 10.min intervals. The penetration was 3.431 % of the benzene portion (0.35%)of the applied dose, lr 0.00108%benzene in relation to the solvent applied. rhis is approximately 4 times the single dose actual pene- ration (using a 50 PI dose rather than 100 PI doses) of 3.0805% of the benzene portion (0.36%)of the applied h e , or 0.00029%in relation to the solvent applied. To approximate chronic exposure to benzene contained n rubber solvent, 190 crl doses, rather than 50 MIdoses *ere used in the m u i t i p l e ~ x p o s u r e xperiment wirn ,I sol'en t. The 0.431 % actual penetration of benzene compu red In this multiple-exposure experiment is slightly more than half the 0.848% penetration in the multiple-exposure experlrnent using full-strength benzene. A similar relationship between penetration of full-strength benzene and benzene in rubber solvent was found in the single-exposure experiments reported previously. In the solvent experiment, how- cvz'!, t l x bmzerie pener:s:icn OF ~ i 1i 0 d 9 ; t s VI'.\ 9/11 le the fall-strength b m r r n e ixperirnent neau~cd~ 1 ~ 1 ) :le penetration of the e.eventh dose. The actrial penetration of I4C. kierizerie ir rubiicr scrlverlt :Iii'l:ugh the rhesus palm wai 0.651 %, This Ienet s t i o n vm g w t e r than the ;?enetra::ion thrciJgh the rlii sus I oreirrn of 0.0805%,and indicates t h a t a t least in th: rilciws rntwk:y, the palm is probablf a poorer barrier f(lr be,izrnC d u n the forearm. I t is n o t p o s i b i e t o compare r t esus p ~ l r n a r al)scirption to absorption through the hurnaii palm sincc iio literature i s available o n i.his w b j e c t I t n a y b,: q w j i:iJmedwhether the relationship between rhesus fmmi m d palrnsr absorption 01' benzene will be r e w a n t to m n . i3;tsed on the limited d a u with compounds c.ther r h m b m zene in man, the absorption through the hurian p3lm n.1j found to be approximately equal to absorption throush the foreirrn.'* Adequate quantitative identification of systemicAl\' absorbed benzene requires that the chemical and i t s n i m b olites be isolated from biological fluids or excreted pruducts. This task is complicated by the relatively simplc! nature of this chemical and the biochemical ubiquity of its compounds and metaboii tes. Much of absorbed benzene is metabolized and the metabolite pool is derived from various sources in addition to the exogenous chemical.' Because of normal biological variability, it is difficult to quantitate the exogenous benzene source of such r n e u b olites. Even with many of the highly specific and sensitive methods of modern analytical technology, this remains a problem. Radiochemical methods provide a means of sclectively tracing the biochemical fate of radioisotopically labelled benzene because the labelled moieues are retained in metabolites and can be detected with great sensitivity 2nd accuracy. Evaluation of the older literature. The most pertinent older data is cited by Hanke et a].' The present cornmsnts are confined to questions of analytical methodology 3nd to quantitative studies relevant :o human absorption. This includes three publications appearing from 1946 to 1961. T w o r e p ~ r t s " ~involving human subjects conciudcd t h ~ t there was n o percutaneous absorption of benzene. The m a lyucal methodology then available limits the reliability of these studies. These methods were based upon a change in the sulfur ratio of total urinary sulfate, due to changes in the organic vs inorganic sulfate, t h a t would result from sulfa= conjugation of phenol formed metabolically from a n y absorbed benzene. Since the total sulfur ratio normally varied between 75% and 9S%, and the amount of sulfate normally excreted in urine is relatively high (;lpprLnimately 2,000 mg/L), this is n o t a reasonably sensiiive method for measuring benzene absorption. The rncthod would probably not detect increases in urinary phenol excretion of thc order of 10 mg,;L Hsnke e t ai.' employed xalg:ical methods suffliicnily sensitive to detect benzene absorption, although not sufticiently prccise to accurately quantitate that absorption. One method involved the direct assay of excreted phcnol obtained by chemical hydrolysis of urinary phenol sulf.itcs. The results indicate-that urinary phenol was elevatcd in individuals dermally exposed to benzene (maximum V.L!UI: of 1 .S8 to 25.3 m g / L 2s compared to maximum of 1 G.0 immgelLnidnvnaoluneesxpoocsceudrrienddaivmidounaglsi)n.diLvaidrguealvsa. rNiaotiromnasli2n4ehxprer--. urinary excretion of totd phenol varied from less than 10 mg to more than SO mg' The authors acknowledged the problem of quantitation and discussed various methods of calculation and also arrived at the unjustified conclusion that "the results would have less error if physiologic excre- tion was calculatedas the average of all four methods of calculation." The authors did not present statistical evaluation of their fiata, nor did they present sufficient data to allow application of statistical evaluations. These data apply to continuous occluded exposure of a 35 to 45 cm2 area of human forearm by 60 m&m2 benzene for 1.25 to 2 hr. For these conditions the authors calculated an average absorption rate of 100%benzene of 0.24 mglcm2. hr. This calculation takes into account that not all of the benzene absorbed systemically appears in ihe urine as phenol. Their data, however, do not allow an estimation of the reliability of this figure. I t is based on an estimated average "surplus" of approximately 5 mg of excreted phenol for exposed individualsabove an estimated Jverage level of approximmly 5 mg for nonexposed individ- uals. Since values for nonexposed individuals were reported to vary between 3.0 and 16.0 with an average of 7.2 mg, it would seem that the calculated value of 0.24 mglcm2-hr i s subject to considerable error and may be substantially over- estimated. The same study employed an alternative method in which absorption was determined as the difference between the amounts of benzene recoverable by evaporationfrom the skin surface at the time o f application and after 1.25 hr of continuous occluded exposure (17.6 mg applied to 8 cm2). While this method i s characterized as more "direct" chem- ically, it i s actually less direct biologically in that it does not necessarily measure benzene reaching the systemic circulation. The unrecoverable benzene presumably consisted of that systemically absorbed plus that retained temporarily in the stratum corneum. Since a substantial portion of the unrecoverable benzene may not have been systemically absorbed during the 1.25-hr exposure period, the calculated "absorption rate" of 0.4 mg/cm2. hr is likely to be substantially overestimated. i n spite of the deficiencies cited and t h e limited precis- ion, the similarity of the results obtained by the two inde- pendent methods used by Hanke e t ai.' is at least indicative of an absorption rate upper limit of less than 0.24 to 0.4 mg /cm2.hr for the severe conditions of exposure employed, These conditions involved continuous exposure by direct contact with 100% benzene and surface occlusion for 1.25 to 2 hr. Respecting the effect of occlusion, thc occlusive method employed by Hanke e t ai. m3y have h3d the effect of increasingthe reportedabsorption rare more than 10. fold.9 The presented percutaneous penetration data can be corn pared to levels o f benzene absorbed by workers from airborne exposure. However, before these estimates are accep. ted for general use, we believe that confirmatory CUtaneOUS human penetration experiments should be performed. Although the present experimental data are the most c o n plete available, we believe that this should be interpreted in a tentative way until human data becomes available. *.**.*.*+* Submitted for publication July25,1980; revised; accepted for publication April 15, 1981. Requests for reprints should be sent to: Howard Maibach, M.D., University of California Medial School, S i n Francisco, CA 94143. REFERENCES 1. OSHA, Final Environmental Impact Statement on a Standard for Occupational Exposure to Benzene (January,1978). 2. Drinkingwater and health: Recommendation of the National Aademy of Sciences. 1977 (July 11).Fed Reg (5ummory) 35764.35777. 3. Feldmann, R.J., and Maihch, H.I. 1970. Absorption of some organic compoundsthrough the skin in man. j Invest DennuroI 54: 399-404. 4. Wester, R.C, and Mlibach, H.I. 1975. Rhesus monkey as an animal model for percutaneouspenetration. In Animui Models in Dermatology, H.I. Maibach, ed., pp. 133-37. Edinburgh: Churchill Livingstone. 5. Ccasaro, AN. 1946. Is benzeneabsorption possible?Med Lovoro 37: 151-56 (Id). 6. Conu, G.L., and Malragliati, A. 1955. Study of the percutaneous benzene absorption. Med Lovom 46: 194-98 (Ital). 7. Metabolism. Altmann, P.L, and Dittmer, DJ., ds.Fed Am Soc L r p Bioi, Bethesda, Maryland, pp. 515,519,525; Neish, AC in Biochemistry of Phenolic Compounds, J. 8. Harborne, ed., Academic Pres, N.Y., 1964, p. 295. 8. Hanke, 1.; Duuiewiw, T.; and Piotrowski, J.1961. Absorption of benzene through the skin in man. Med Prog 12: 413-26. 9. Feldmann, R.J., and Maibach, H.I. 1965. Penetration of "C hydrocortisone through normal skin. Arch Dermotol 91: 661-66. I 1 I ! i I i260 Archives o f Environmental Hell[*