Carbon disulphide toxicology: The present picture

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Vol. I4 pp. 57-65

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Pergamon

Press 1976. Printed

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See tion ARTICLES

CARBON

DISULPHIDE

OF GENERAL TOXICOLOGY:

Current questions regarding the results of industrial exposure to carbon disulphide (CSJ were presented at the Third International Symposium on Toxicology of Carbon Disulphide, held in Cairo and Alexandria in May 1974 (Lieben, Archs erwir. Hlth 1974, 29, 173). The main interest centred on the cardiovascular effects of CS,, and its effects on blood, liver, kidneys and gonads, and on the value of its metabolites as possible early indicators of significant exposure. Threshold limit values (TLVs), which differ widely between East and West, came in for scrutiny, with several authorities supporting a case for reducing the TLV from 20 to 10 ppm CS, despite the difficulty apparently experienced by some companies in maintaining even the higher value. The determination of two CS2 metabolites, 5-mercaptothiazolidone and thiourea, was suggested as a possible improvement on the merely qualitative urinary iodine-azide test for detecting significant exposure. No agreement was reached during the symposium on whether women should be permitted to work at all under conditions involving exposure to CS2. East Germany and the USSR are not at present prepared to restrict the exposure of women to CS,, but several other countries have prohibited such exposure on the strength of demonstrations of teratogenesis induced in experimental animals (Lieben, lot. cir.). A number of new advances in CS2 toxicology were discussed in the rather tardily reported 1972 Yant Memorial Lecture (Teisinger, Aln. irld. Hyg. Ass. J. 1974, 35, 55). Rats exposed to 1.2-2.4 mg CS,/litre for 6 hours daily for l&50 weeks developed a disturbance in pyridoxine metabolism which could be counteracted by giving pyridoxine supplements, but the neuropathy that developed improved only slowly, and it seems possible that CS2 may also upset nicotinic acid metabolism. CS, has been reported to inhibit serotonin activity in the brain by reducing monoamine-oxidase activity. A diminution in the tissue levels of copper and zinc available for enzyme systems occurs in CS2 exposure and may be involved in the toxic action, although whether CS2 itself or one of its metabolites (mercaptothiazolidone or thiourea) is primarily responsible for CS2 toxicity is so far unknown. A relationship between exposure to CS2 and coronary heart disease probably exists, but studies of the lipid disturbances involved have given contradictory results, and there is no more than a strong suspicion that exposure to CS, aggravates the atherosclerotic process (Teisinger, lot. cit.). Other reports claim that CS2 has a toxic effect on the optic nerve and produces changes in the retina suggestive of diabetic retinopathy, and that it decreases secretion of

INTEREST THE PRESENT PICTURE

testosterone and inhibits spermatogenesis (Teisinger. lot. cir.). Some of these aspects of CS2 toxicity may now be examined in more detail. Nervous

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Briiderl & Benini (Schwei:. wed. Wschr. 1974. 104, 15) report five instances in which moderate chronic industrial intoxication by CS, resulted in polyneuropathy as well as the vaguer symptoms of insomnia, irritability, gastric disorders, circulatory disorders, loss of weight, loss of libido and loss of potency. The patients came from a group of six men aged 41-63 years, who had been exposed in a viscose plant to CS, concentrations ranging up to 65 mg/m3 for several years. Sleep disturbances, fatigue, nausea and headache were encountered after periods of exposure ranging from 1 to 26 years, and these effects were complicated by the polyneuritic symptoms of paraesthesia, muscle weakness and cramping pains in the extremities. There were signs of improvement after termination of exposure to CS2, but these features persisted to some extent in three patients. Among the sequelae to CS2 intoxication were intermittent claudication, loss of detectable pulse in the foot. atrophy of the skin of the lower thigh, giddy spells and premature senility. The tisssue half-life of CS2 in rats in relation to its effect on braincatecholamine concentrations was studied by Magos et al. (Iut. Arch. Arbeitsrned. 1974, 32, 289) who exposed rats to 2 mg CS,/litre either for I hour or for two Chour periods on consecutive days. The whole-body burden of CS2 after the l-hour exposure was 160 nmol/g and it fell exponentially from this level with a half-life of 35 minutes. In blood and liver, CS2 concentrations were similar to the whole-body concentration, but their rates of decrease showed initially a rapid phase, in which the half-life was 10 minutes, followed by a slower phase with a half-life approaching 35 minutes. Immediately after the two Chour periods of exposure, brain concentrations of noradrenaline and dopamine were below control values; after 20 hours the level of noradrenaline was still depressed, while that of dopamine had returned to the control level. Sensitivity to amphetamine-induced stereotypy was still enhanced after 20 hours. Thus, in spite of its relatively rapid elimination from the body, CS2 can have a cumulative depressant effect on catecholamines. These results support the hypothesis that disorders of catecholamine metabolism may be a factor in the development of ischaemic heart disease in persons occupationally exposed to CS2 (Cited in F.C.7: 1974, 12, 260). In rats exposed to 2.5 mg CSJlitre for 15 hours, glutamate 57

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Articlesof generalinterest

amount of cytochromeP-450 presentin the liver at the time of dosing,and wasalsorelated,but in a more complexway, to the severity of the toxic changesinduced in the liver. Incubation of CS2 with rat-liver microsomesproduceda marked lossof cytochrome P-450only whenNADPH waspresent.and the addition of EDTA, which preventsmalonaldehydeformaAdrenals tion, did not influencethis reaction.Thus, the oxidaThe adrenalglandsof rats exposedto 2 mg CS2/ tive metabolismof CS2 appearsto be an essential litre for 4 hours on two successive days showeda step in its production of liver toxicity. In phenobarbitone-treated rats, a rapid increasein doublingof their dopaminecontent (Jarvis& Magos, Naunyn-Schnriedebergs Arch. exp. Path. Pharmak. the watercontent of the liver. maximalat 12-16hours 1973,278,207)and the increasein dopamineconcent- and returning to control values by 48 hours, was ration was significantly enhancedby pretreatment apparentafter exposureto 4 mg CS,/litre for 4 hours with phenobarbitoneor by starvation before expo- (Butler et al. J. Path. 1974, 113, 53). At the same sure.The highestadrenaldopaminecontent observed time, the concentrationof Na’ and K’ in the liver was in C&-treated animalsthat were both starved increasedwhen calculated on the dry weight. One and pretreatedwith phenobarbitone,but no increase effect of CS2 may be to causedissolution of the smooth endoplasmicreticulum in the liver, and an occurredin the absenceof CS2 exposure. increasedwater uptake by osmosis;meanwhilean inLiver tact Na+ pump maintains normal liver concenFreundt et al. (Int. Arch. Arbeitsmed. 1974,32, 297) trations of Na’ and K’, and during the recovery reported that pretreatmentof rats with three succes- phase the damagedmembrane,included in memsive daily dosesof 80 mg phenobarbitone/kg,their brane-boundautophagicvacuoles,does not impede subsequent exposurefor 8 hours to 20 or 200 ppm the return of cytoplasmichydration to normal. The CS, and finally the injection of 100 mg hexobarbi- sameauthors(idem, ibid 1974,113, 79) have reported tone/kg failed to causeany significant fat accumu- that the treatment describedabove resulted in an lation in liver cellsor to alter serumlevelsof gluta- early and consistentloss of glucose-6-phosphatase mic-oxalacetic and glutamic-pyruvic transaminase and aniline hydroxylasefrom the endoplasmicreticu(SGOT and SGPT).Pretreatmentwith hexobarbitone lum of the centrilobular areasof the liver. Loss of followed by exposureto 200 ppm CS, for 8 hours acid phosphatasealso occurred during the phaseof on two successive daysalsofailed to producehepatic hydropic degeneration,while during recovery large fatty change.In contrast, pretreatmentwith pheno- acidphosphatase-positive bodieswere seenwithin the barbitonefollowed 24 hourslater by oral intubation hydropic areas. There was no detectable loss of of 1 ml C$/kg in olive oil (1 ml/kg) resultedin exten- adenosine triphosphatasefrom the plasmamembrane. sivefatty depositsin the Kupffer cellsand in the cen- Thesefindings indicate that CS2 damagesonly the trilobular, intermediateand acino-peripheralareasof smoothendoplasmicreticulum. the liver, and in a rise in esterifledfatty acids and Butler et al. (ibid 1974,112, 147)failed to find any SGOT. Without phenobarbitonepretreatment,orally ultrastructural changesin the hepatic parenchymal administeredCS, producedlessextensivefatty depo- cells of rats exposedto 4 mg CS,/litre for 4 hours sition, and olive oil alone had no effect. Thus it without phenobarbitonepretreatment. Phenobarbiappearsthat occupationalexposureto the TLV of tone alone induceda proliferation of the smoothen20 ppm CS2 with occasionalpeaks of 200 ppm is doplasmicreticulum; this wasmost prominentin the probably unlikely to produce degenerative liver centrilobularcellsbut alsoappearedin the periportal damagein personswith a healthy liver who are being cells, Exposure of rats to CS, following starvation treated therapeuticallywith a barbiturate drug. and phenobarbitonetreatmentresultedin dilatation of In rat-liver microsomal preparations incubated the cisternaeof the rough endoplasmicreticulum of with CS, labelledwith 14Cor 3sS,carbonyl sulphide the centrilobularhepatocytes,followedby an increase (COS)wasproducedin the presenceof NADPH but in sizeof the vesiclesfor 12-16hours, the mitochonnot in its absence(Dalvi et al. Life Sci. 1974,14,1785). dria and other organellesremainingnormal. During The production of COS was inhibited in an atmos- the recovery phasethe vesiclescollapsed,the cytophereof CO. It is thereforesuggested that (by analogy plasm reverted to normal and large autophagic with the metabolismof parathion to paraoxon) CS2 vacuolesappeared.The C&-induced lesionsin the may be metabolizedto COS with the releaseof reac- liver appeared,therefore,to dependon induction of tive sulphurand its covalent binding to the microso- the smooth endoplasmicreticulum by phenobarbima1membrane.This secondreaction may be respon- tone. siblefor the liver necrosisproducedby CSI in phenobarbitone-pretreated rats(Dalvi et al. lot. cit.). de Mat- Circulatory effects on the eye teis & Seawright(Chemico-Biol.Interactions 1973, 7, Teisinger(lot. cit.) has referred to the toxic effect 375)reported that whereasthe pretreatmentof rats of CS2 on the optic nerve, and to retinal changes with phenobarbitoneand starvation enhancedthe resemblingthoseof diabetesmellitus.A study of the liver toxicity of CS,, the administrationof a small microcirculation of the ocular fundus of workers prior dose of CS2 diminishedthe liver toxicity pro- exposedto CS2 has been describedby Raitta et al. ducedby the standarddose.A study of the excretion (Albrecht v. Graefes Arch. klin. exp. Ophthal. 1974,191, of 14C02after an ip doseof 14CSzshowedthat the 151).Ophthalmologicalexaminationwas carried out amountof 14C02exhaledwaslinearly relatedto the on 100menwho hadbeenexposedto CS2in a viscosedecarboxylaseactivity in the brain was diminished (Tarkowski,Int. Arch. Arbeitsrned. 1974,33, 79).This effectmay explain the reducedI’-aminobutyratelevels observedin the brain after CS, exposure(Cited in F.C.?Y1973,11, 918). Pyridoxal phosphatefailed to protect rats againstthis enzymeinhibition.

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Articles of general interest

rayon plant for l-27 years (mean 15 years) and exposureto 20 ppm CS2 for 6 hours on 5 successive of 97 controls who worked in a paper mill. The only daysproduceda delay in amidopyrineexcretionsixnisignificantdifferencesobservedwere a delay in peri- lar to that seenafter a single6-hour exposureto 40 papillary filling, either circumferentialor segmental ppm CS,, so that somecumulative effect was indior both, and an increasein the meancalibre of the cated. The authors ‘suggestthat inhibition of drug retinal arteriolesin 68 of the exposedmencompared metabolismof this type might piove valuable as a with 38 of the controls. Thesechangescould not be sensitivetest for exposureof workers to CS2. related to ageing,smokinghabits, serum-cholesterol The predictionof an individual worker’ssusceptibilevels or glucosetolerance,and intraocular tension lity to CS2was the subjectof a study by Djuric’ et was within normal limits. Thus, no retinopathy di- al. (Arch envir. Hlth 1973,26, 287),involving the use rectly attributable to CS2 exposure was detected. of disulfiram.This drug and CS2are apparentJymetaHowever,it is possiblethat the wideningof the retinal bolized by the sameenzyme system,or by related arterioles reflected the release or the increased systems, and thereforemay be competitive.The metaappearanceof a circulatingmediatoraffectingthe vas- bolite from disulfiram is diethyldithiocarbamate culature, and this may be related to other suspected (DDC). A singledoseof 500mg disul!iramwasgiven but not confirmedcardiovasculareffectsof CS2. to 33viscoseworkersknown to be susceptibleto CS2 sincethey had developedpolyneuritis as a result of Detection of excessive exposure exposurein the past, to 21 workers who had been The necessityfor morespecificand quantitative in- exposedto high concentrationsof CS2for 7-14 years dicesof early exposureto CS, was stressedduring without developing signsof intoxication and to 18 the Third International Symposium(Lieben,lot. cit.). workerswho had beenexposedto sub-TLV concentAt presentthe qualitative method of urine analysis rations for 11-15 years without developingsignsof by the iodine-azidereaction,which dependsupon the intoxication. A urine specimenwas taken from each presenceof the CS, metabolite, thiourea, is used. subject 5 hours after the disulfiram administration Mack et al. (Biochem. Pharmac. 1974, 23, 607) have and assayedphotometricallyfor DDC. IncreasedCS2 reported that in 19 healthy subjectsexposedfor 6 exposurewas associatedwith a decreasein urinary hours to 10-80 ppm CS2, there was first a marked excretion of DDC, the lowestexcretion rate of DDC depressionin the excretion of a doseof 7 mg amido- beingin thoseworkerswho had oncedevelopedsigns pyrine/kg and subsequentlyan increase.At 10 ppm, of chronic CS2intoxication. The authorssuggestthat CS, markedly reducedthe urinary excretion of the a potential for greatermetabolismof compoundsconamidopyrine metabolite, 4-aminoantipyrine,but the taining sulphur may go with a higher resistanceto early deficit wascompensated almostcompletelydur- CS2intoxication and vice versa. It is thereforepossing the later excretion phase.Inhibition of the drug ible that a worker’sdegreeof susceptibilityto chronic excretion was reversible,and increasedwith increas- CS2exposurecould be predictedfrom the knowledge ing degreeof exposureto CS,; it was no longer de- of how efficiently hecan excretea singledoseof disultectable18hoursafter exposureto 20ppm CS2.Daily firam [P. Cooper-BIBRA]

PATTERNS OF CHLOROFORM METABOLISM We recently reported the results of teratology studieson chloroform administeredorally and by inhalation (Cirerl i,t F.C.7: 1975. 13, 402). Although theseand numerousother fairly sophisticatedtoxicologicaland biochemicalstudieshavebeencarried out on chloroform. its metabolism appears to have receivedonly limited attention in the past, most of the investigationsreportedhaving beenin connexion with carbon tetrachloride metabolism, in which chloroform is apparently formed as an intermediate (Hathway, Arzneimittel-Forsch. 1974,24, 173; Uehleke et al. Xenobiorica 1973,3, 1). Its metabolicfate has recently receivedmore systematicconsideration, however,having beeninvestigatedin the mouse,rat and squirrelmonkey (Brown et al. ibid 1974,4, 151). The animalswere given 14C-labelledchloroform, with a specificactivity of about 5 &i/kg, by gavage in a doseof 60 mg/‘kgand werekept in glassmetabolism cages.The exhaled air was passedthrough a seriesof traps containing, first, anhydrous magnesiumperchlorateand silicagel to removewater, then toluene to remove toluene-solublemetabolitesand chloroform, and finally a mixture of cellosolveand ethanolamineto trap carbon dioxide (CO,), and the

contents of each trap was assayed for retained radioactivity. Blood sampleswere taken from mice and squirrelmonkeys,tolueneextractsof bile samples from frozen monkey sectionswere analysedby gasliquid chromatography,and the faeces,urine and carcasses of treatedanimalswere examinedfor radioactivity. Whole-bodyautoradiographywascarriedout on the rat and monkey. Distinct speciesdifferenceswere found in the patternsof chloroformexcretion,but three mousestrains all gave similar results. After administration of a singleoral dose,a peak level of chloroform (about 3 pg/ml) appearedin the blood of the mice within 1 hour. Within 24 hours of dosing,some80% of the radioactivity appearedin the ethanolaminetraps as 14C02while about 139; was presentin the urine as bicarbonateand/or carbonate.Approximately 6%was found in the toluenetraps and about 2.5% remained in the carcass48 hours after treatment. Excretion in the rat followed a similar pattern, although about 66’4 of the radioactivity appearedas 14C02and 20% asexhaledchloroform or its toluene-solublemetabolites in the first 48 hours. A small amount (about 8%) appearedin the urine and faecesover the 96