The metabolism of DDT in vivo by the Japanese quail (Coturnix coturnix japonica)

Male and female Japanese quail (Coturnix coturnix japonica) were given intraperitoneal injections of [¹⁴C]DDT in ethanol at a rate of 13.4 mg/kg body wt. Fifty-six days later the tissues and droppings were analysed for total ¹⁴C and metabolites. The rate of loss of ¹⁴C in droppings was very similar in males and females. The maximal rate was reached on the third day, and 65–66% of the injected dose was voided by the fifty-sixth day. Ninety-three to ninety-four percent of the ¹⁴C in droppings and 83–90% of the ¹⁴C in tissues were extracted by solvents. Combined extracts from males and females were used for determination of DDT and its metabolites. Expressing all results as percentages of injected dose, the following were isolated from droppings: DDA (24%), DDT (3%), DDD (5.1%), DDE (11%), and uncharacterised polar metabolites (17%). Twenty-five percent of the dose was retained in the tissues and this was largely accounted for as DDT (10.4%) and DDE (10.5%). Of the total metabolites found 31% was DDE (almost equally divided between tissues and droppings) and 35% was DDA (almost entirely in droppings). Since DDD was not found in significant quantities in tissues, the substantial quantities in droppings were probably produced from DDT by the action of microorganisms.     

Comparative metabolism of 1,1-Bis-(p-chlorophenyl)-2-nitropropane (Prolan) in mouse,insects, and in a model ecosystem

¹⁴C-labeled Prolan or 1,1-bis-(p-chlorophenyl)-2-nitropropane was found to be some-what more biodegradable than DDT. This insecticide, although highly resistant to microsomal metabolism, was degraded by elimination to 1,1-bis-(p-chlorophenyl)-1-propene, and by reduction to 1,1-bis-(p-chlorophenyl)-2-aminopropane. The major degradative pathway, however, was by oxidation to 1,1-bis-(p-chlorophenyl)-2-propanone, to 1,1-bis-(p-chlorophenyl)-pyruvic acid, to bis-(p-chlorophenyl)-acetic acid, and ultimately to p,p′-dichlorobenzophenone. Therefore the ultimate degradative products of Prolan are identical to those produced from DDT.     

Physiological basis for antagonism of clethodim by imazapic on goosegrass (Eleusine indica (L.) Gaertn.)

Greenhouse and laboratory experiments were conducted to determine the effect of imazapic on the herbicidal activity of clethodim on goosegrass. Imazapic did not affect absorption of [¹⁴C]clethodim by goosegrass. Averaged across the two treatments of clethodim alone and clethodim plus imazapic, absorption was 36 and 89% of applied [¹⁴C]clethodim at 0.5 and 96 h, respectively. The majority of [¹⁴C]clethodim (79% of applied) was absorbed by 24 h. Translocation of ¹⁴C was not affected by imazapic, and 3.6% of applied ¹⁴C had translocated into the portion of the shoot below the treated leaf at 96 h after treatment. Metabolism of clethodim was not affected by the presence of imazapic. Three major metabolites of clethodim were detected in treated tissue at all harvest intervals. The majority (58%) of [¹⁴C]clethodim was converted to a relative polar metabolite form 96 h after treatment, whether clethodim was applied alone or in the presence of imazapic. One day after treatment, the photosynthetic rate in plants treated with imazapic decreased below the rate in the non-treated check, and was less for 8 days, the duration of the study. These data suggest that the antagonism of clethodim by imazapic may be caused by imazapic reducing the photosynthetic rate of goosegrass and therefore the sensitivity of ACCase to clethodim.     

Binding of [14C]DDT and [14C]dicyclohexylcarbodi-imide to a lipoprotein of the membrane sector of mitochondrial ATpase

Intact mitochondria, isolated from red coxal muscle of the American cockroach (Periplaneta americana L.), were incubated in the presence of 1,1,1-trichloro-2,2-bis(4-chloro[¹⁴C]phenyl)ethane ([¹⁴C]DDT) to isolate a suspected binding site for DDT in the membrane sector of the mitochondrial ATPase. The requirements for the binding of DDT were compared with those for the binding of dicyclohexyl[¹⁴C]carbodi-imide([¹⁴C]DCCD), a potent inhibitory probe of mitochondrial ATPase activity. [¹⁴C]DDT appeared to bind to a proteolipid of the membrane sector, which also binds [¹⁴C]DCCD. Exchange experiments, with [¹⁴C]DCCD, [¹⁴C]DDT and unlabelled DDT at different concentrations, indicated that DDT and DCCD may be acting on a similar protein. This protein may act as the energy transducing protonophore required for the synthesis and hydrolysis of ATP in coupled mitochondria. Inhibition of mitochondrial ATPase activity may be a consequence of DDT and DCCD binding to this proteolipid protonophore, resulting in the disruption of energy transduction in muscle and nerve.     

Metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide] inhibition of gibberellin precursor biosynthesis

[2-¹⁴C]Mevalonic acid incorporation into gibberellic acid precursors was measured in cell-free extracts from sorghum [Sorghum bicolor (L.) Moench var. G-522 DR] coleoptiles. ¹⁴C incorporation into ent-kaur-16-ene was inhibited ca. 90% by 10^?7 to 10^?4M metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide]. [¹⁴C]Geranylgeraniol (GG) content increased. [¹⁴C]Farnesol content was not altered and [¹⁴C]geraniol content decreased. Total ¹⁴C incorporation was decreased by metolachlor. In the safener [α-(cyanomethoximino)benacetonitrile]-treated sorghum seed coleoptile cell-free system, total ¹⁴C incorporation increased, [¹⁴C]kaurene and relative kaurence content increased 4× up to 10⁵M metolachlor, and [¹⁴C]farnesol, and [¹⁴C]GG contents increased while relative farnesol and relative GG contents were not influenced by metolachlor. Thus, the inhibition of kaurene synthesis by metolachlor was reversed by the safener. Since the biosynthetic processes are mevalonic acid → geraniol → farnesol → GG → copalylol → kaurene, these data corroborate a proposed gibberellic acid biosynthesis inhibition between GG and kaurene as well as a partial blockage between mevalonic acid and geraniol. Thus, a portion of metolachlor-induced growth inhibitions of sorghum could be explicable on the basis of gibberellic acid biosynthesis inhibitions.     

Effect of DDT on photosynthesis of Selanastrum capricormutum

The effect of DDT (2,2-bis-(p-chlorophenyl)-1,1,1-trichloroethane) on carbon assimilation of a green alga, Selanastrum capricormutum was studied. DDT at concentrations between 3.6 and 36 ppb was inhibitory to the photosynthetic CO₂ fixation (ethanol-soluble and/or ethanol-insoluble) and the longer the exposure to DDT, the greater the inhibition. Kinetic studies of photosynthetic CO₂ fixation indicated that DDT stimulated the incorporation of carbon-14 into glycolic acid, a major compound of photorespiration and caused the concomittant suppression of flow of carbon-14 into aspartic acid, a major component of the C₄-dicarboxylic acid pathway. The shift from an efficient pathway into a nonefficient pathway by DDT was interpreted to be through interruption of cyclic photophosphorylation.     

[14C]Dicofol association to cellular components of Azospirillum lipoferum

The bacterium Azospirillum lipoferum is able to survive in high concen-trations of the organochlorine acaricide dicofol [1,1-bis-(4-chlorophenyl)-2,2,2-trichloroethanol]. It accumulates this chemical in the cell envelope where it is protected against hydrolysis. We investigated the nature of cell envelope molecules with which [¹⁴C]dicofol is associated; no indication of [¹⁴C]dicofol–saccharide bonds was found. We concluded that about 80% of the total [¹⁴C]dicofol found in the cells was associated with lipids and the remaining 20% with proteins. Electrophoresis did not indicate any correlation of a specific protein band with [¹⁴C]dicofol radioactivity peaks. After Folch partition, [¹⁴C]dicofol distribution in TLC analysis showed 60% of [¹⁴C]dicofol–lipid bonds related to neutral lipids, 20% to phospholipids and the remaining 20% of the bonds associated with other lipids. Experimental results suggested that [¹⁴C]dicofol associates mainly with membrane domains near proteins and that this association influences membrane fluidity as well as enzymatic activity. © 1998 SCI     

DDT and pyrethroids: Receptor binding and mode of action in the house fly

The mode of action of DDT and pyrethroids was investigated in the house fly, Musca domestica L, using drug:receptor binding techniques. Both in vivo and in vitro binding studies demonstrated the existence of membrane receptors which bind specifically to [¹⁴C]DDT and [¹⁴C]cis-permethrin. The receptors show properties to be expected of a critical target site of these insecticides. These include negative temperature correlation with binding, relatively nonsensitivity to DDE, and sensitivity to Ca²⁺. The receptor sites are readily saturated at 45–90 nM [¹⁴C]DDT and have an apparent disassociation constant (K_d) of 12.2 nM. The maximum number of binding sites was estimated to be 17 pmol DDT/mg membrane protein (0.34 pmol/house fly head). Competition studies showed DDT, cis-permethrin, and cypermethrin bind to the same receptor but not at precisely the same site. The addition of Ca²⁺ to the incubation buffer significantly inhibited the binding of both [¹⁴C]DDT and [¹⁴C]cis-permethrin, suggesting the receptor binding is Ca²⁺ sensitive and may have a role in ion conductance.     

S-Ethyl dipropylthiocarbamate, N,N-Diallyl-2-chloroacetamide,and 2-chloroallyl diethyldithiocarbamate influence on sulfhydryl-dependent sucrose accumulation

l-[U-¹⁴C]sucrose accumulation by phloem sieve tube members (PSTM) of wheat (Triticum aestivum L. ‘Holley’) and sorghum (Sorghum bicolor L. ‘G522 DR’) was inhibited by the nonpermeant sulfhydryl inhibitor p-chloromercuribenzenesulfonic acid (PCMBS), and this inhibition was reversed by the permeant sulfhydryl protectants dithiothreitol (DTT) and dithioerythritol (DTE). S-Ethyl dipropylthiocarbamate (EPTC) (≤0.1 mM) did not inhibit [¹⁴C]sucrose accumulation by wheat or sorghum PSTM. N-N-Diallyl-2-chloroacetamide (CDAA) (1 mM) inhibited [¹⁴C]sucrose accumulation by sorghum but not by wheat PSTM. The inhibition of [¹⁴C]sucrose accumulation in sorghum PSTM by the membrane permeant CDAA was reversed by DTT. Sorghum growth was inhibited by <1 μM CDAA. Membrane permeant 2-chloroallyl diethyldithiocarbamate (CDEC) (0.1 mM) inhibited [¹⁴C]sucrose accumulation by PSTM of sorghum but not wheat. The inhibition of sucrose accumulation in sorghum PSTM by 0.1 mM CDEC was reversed by DDT.     

Metabolic resistance mechanisms of the housefly (Musca domestica) resistant to pyraclofos

In vivo and in vitro metabolism of pyraclofos labeled with ¹⁴C on benzene ring was studied in the pyraclofos-resistant and -susceptible female houseflies. In vivo metabolism studies, the metabolic rate of pyraclofos was the same in both strains. Pyraclofos primarily undergoes metabolic detoxification by cleavage of P-S-alkyl bond, and cleavage of the P-O-aryl bond followed by CHP [1-(4-chlorophenyl)-4-hydroxypyrazole]]-glucose conjugation. Cleavage of P-O-aryl bond and CHP-glucose conjugation is more predominant in the resistant strain whereas the cleavage of P-S-propyl bond resulting in EHP-CHP [O-1-(4-chlorophenyl)pyrazol-4-yl ethyl hydrogen phosphate] is more preferred in the susceptible strain. CHP production by P-O-aryl bond cleavage was controlled by P450 monooxygenase and esterase. UDP-glucosyltransferase appeared to play an important role in the pyraclofos metabolism of the resistant strain. Production of CHP-glucose conjugate was largely reduced by piperonyl butoxide and S,S,S-tributylphosphorotrithioate in both strains. EHP-CHP production seemed to be controlled by P450 monooxygenase and stimulated by UDP-glucose.     

Acifluorfen metabolism in soybean: Diphenylether bond cleavage and the formation of homoglutathione, cysteine, and glucose conjugates

Metabolism of the substituted diphenylether herbicide, acifluorfen [sodium 5-(2-chloro-4-trifluoromethylphenoxy)-2-nitrobenzoate], was studied in excised leaf tissues of soybean [Glycine max (L.) Merr. ‘Evans’]. Studies with [chlorophenyl-¹⁴C]- and [nitrophenyl-¹⁴C]acifluorfen showed that the diphenylether bond was rapidly cleaved. From 85 to 95% of the absorbed [¹⁴C]acifluorfen was metabolized in less than 24 hr. Major polar metabolites were isolated and purified by solvent partitioning, adsorption, thin layer, and high-performance liquid chromatography. The major [chlorophenyl-¹⁴C]-labeled metabolite was identified as a malonyl-β- -glucoside (I) of 2-chloro-4-trifluoromethylphenol. Major [nitrophenyl-¹⁴C]-labeled metabolites were identified as a homoglutathione conjugate [S-(3-carboxy-4-nitrophenyl) γ-glutamyl-cysteinyl-β-alanine] (II), and a cysteine conjugate [S-(3-carboxy-4-nitrophenyl)cysteine] (III).     

β-d-Glucosyl and O-malonyl-β-d-glucosyl conjugates of pentachlorophenol in soybean and wheat: Identification and enzymatic synthesis

The polar metabolite fraction formed by soybean cell suspension cultures from [¹⁴C]pentachlorophenol (PCP) was fractionated by repeated high-performance liquid chromatography. The β-d-glucopyranosyl conjugate of PCP was identified by cochromatography, by chemical and enzymatic degradation, and by mass spectroscopic comparison with the authentic compound. The O-(malonyl)-β-d-glucopyranosyl conjugate of PCP was identified by chromatographic and electrophoretic comparison with the enzymatically synthesized compound, by specific chemical and enzymatic degradation, and by mass spectroscopy. Soluble protein extracts from both soybean and wheat cells catalyzed (a) the UDPG-dependent conversion of PCP to the β-d-glucopyranosyl conjugate, and (b) the malonyl-SCoA-dependent further conversion to the O-(malonyl)-β-d-glucopyranosyl conjugate. These enzymes were partially purified and had apparent molecular weights between 40,000 and 48,000. Free PCP was again released from the conjugates upon incubation with plant malonyl esterase and/or β-glucosidase preparations.     

Comparative effects of three isoxazole-urea herbicidal derivatives on the metabolism of enzymatically isolated soybean leaf cells*

The effects of the herbicide isouron and of its plant degradation products designated as metabolite l {N-[5-(1,1-dimethylethyl)-3-isoxazolyl]-N-methylurea} and metabolite 2 {N-[5-(1,1-dimethylethyl)-3-isoxazolyl]-urea} on the metabolism of enzymatically isolated leaf cells of soybean [Glycine max (L.) Merr., cv. Essex] were compared under laboratory conditions. Photosynthesis, protein synthesis, ribonucleic acid synthesis, and lipid synthesis were assayed by the incorporation of NaH¹⁴CO₃, [¹⁴C]-leucine, [¹⁴C]-uracil, and [¹⁴C]-acetate, respectively, into the isolated cells. Time-course and concentration studies included incubation periods of 30, 60, and 120 min and concentrations of 0.1, 1, 10 and 100 μM of the three herbicides. The urea derivative of isouron (metabolite 2) was the least active of the three compounds. The activity of the mono-methylated derivative of isouron (metabolite 1) was comparable to that of isouron and the sensitivity of the four processes to both chemicals decreased in the order: photosynthesis > ribonucleic acid synthesis > lipid synthesis > protein synthesis. The concentration of isouron that caused a 50% inhibition of photosynthesis of the isolated soybean leaf cells was calculated at 0.51 μM. The effects of isouron and metabolite 1 on photosynthesis, lipid and RNA synthesis appeared to be independent of incubation lime as maximal inhibition occurred within 30 min. Inhibition of protein synthesis by both chemicals was time-dependent, increasing in magnitude with concomitant increases in incubation time.     

Behaviour of ddt in three soils exposed to solar radiations under different conditions

Volatilization, mineralization, degradation and binding of soil-applied [¹⁴C]DDT were studied in three different soils from a tropical region of southern India subjected to solar irradiation and flooding for a period of 42 days. The soil types–red cotton soil, nursery soil and canal bank soil–differed in their organic carbon content, pH and texture. Under unflooded conditions, volatile losses were highest in the sandy canal bank soil. Flooding significantly enhanced volatilization, and this effect was maximal in the nursery soil, which had the highest organic carbon. The soils fully exposed to solar radiations in quartz tubes registered 1.5-1.8 times greater volatility. The volatilized organics contained appreciable quantities of DDE under both flooded and unflooded conditions. In addition, greater quantities of DDD volatilized from the flooded systems. The rate of formation of DDE was faster when soils were irradiated in quartz tubes. Mineralization remained minimal throughout the period of exposure and flooding the soil appeared to reduce further the [¹⁴C]carbon dioxide evolution. Canal bank soil exhibited the least mineralization and degradation. The data indicate that volatilization was significantly influenced by solar radiation and flooding to a much greater degree than by the differences in soil properties. Binding of DDT to soil was significantly increased by flooding the soil, thus leaving up to 33% of the initial DDT as bound residues in the nursery soil.     

The metabolism of fenvalerate in plants: The conjugation of the acid moiety

The metabolism of the pyrethroid insecticide fenvalerate [(RS)-α-cyano-3-phenoxybenzyl (RS)-2-(4-chlorophenyl)-3-methylbutyrate] ( I ), and of its most insecticidal (αS,2S) isomer ( II ), has been examined in cabbage plants grown and treated under laboratory conditions with [¹⁴C]chlorophenyl- and [ring-¹⁴C]benzyllabelled preparations of the two compounds. Both insecticides disappeared from the treated leaves with similar half-lives of approximately 12–14 days; they underwent ester cleavage to a significant extent, together with some hydroxylation at the 2- or 4-position of the phenoxy ring, and hydrolysis of the nitrile group to amide and carboxyl groups. Most of the carboxylic acids and phenols thus produced occurred as glycoside conjugates. In separate experiments, the uptake and metabolism of 2-(4-chlorophenyl)-3-methylbutyric acid ( X ), the acidic half of the molecule, were examined in the laboratory, using abscised leaves of kidney bean, cabbage, cotton, cucumber and tomato plants. The acid X was found to be readily converted, mainly into glucose and 6-O-malonylglucose esters in kidney bean, cabbage and cucumber plants, into glucosylxylose, sophorose and gentiobiose esters in cotton, and into two types of triglucose esters with differing isomerism in tomato. One of the acetyl derivatives of the trisaccharide conjugates was identical with the synthetic deca-acetyl derivative of the [1 → 6]-triglucose ester.     

Formulation and adjuvant effects on the absorption and translocation of 14C-clethodim in wheat (Triticum aestivum L.)

A new formulation of clethodim {(E,E)-(±)-2-(1-[([3-chloro-2-propenyl]oxy)imino]propyl)-5-(2-[ethylthio]propyl)-3-hydroxy-2-cyclohexen-1-one} is labeled for the control of grasses and volunteer grass crops, including glyphosate-resistant corn. The effects of the formulation (new: 0.12 kg L⁻¹ and current: 0.24 kg L⁻¹) and adjuvants (ammonium sulfate [AMS], crop oil concentrate [COC] or both) on the absorption and translocation of the ¹⁴C-clethodim was determined at 1, 4, 12, 24, 48, and 72 h after treatment (HAT) in wheat under greenhouse conditions. The absorption of the ¹⁴C-clethodim with the 0.12 kg L⁻¹ formulation was higher than that with the 0.24 kg L⁻¹ formulation, especially at 24 HAT and beyond, regardless of the presence or absence of an adjuvant. The addition of an adjuvant increased the absorption of the ¹⁴C-clethodim with the 0.12 kg L⁻¹ formulation at all harvest times, except at 72 HAT. However, there were no differences in the ¹⁴C-clethodim absorption among the adjuvants added to the 0.12 kg L⁻¹ formulation, except at 48 and 72 HAT. Most of the ¹⁴C-clethodim remained in the treated leaf independent of the formulation or adjuvant. The formulation did not have an impact on the distribution of the absorbed ¹⁴C-clethodim; however, the presence of an adjuvant increased the movement of the ¹⁴C-clethodim out of the treated leaf. Most of the absorbed ¹⁴C-clethodim remained in the treated leaf and a negligible amount translocated to the root. These results demonstrated the improved absorption of clethodim with a formulation containing half of the active ingredient (0.12 kg L⁻¹) and the inclusion of both AMS and COC.     

Comparative metabolism of atrazine and EPTC in proso millet (Panicum miliaceum L.) and corn

The purpose of this study was to examine the differential activities of proso millet (Panicum miliaceum L.) and corn (Zea mays L.) with respect to atrazine [2-chloro-4-(ethylamino)-6-(isopropylamino)-S-triazine] and EPTC (S-ethyldipropyl thiocarbamate) metabolism. GSH-S-transferase was isolated from proso millet shoots and roots. When assayed spectrophotometrically using CDNB (1-chloro 2,4-dinitrobenzene) as a substrate, the shoot enzyme had only 10% of the activity of corn shoot enzyme while the root enzyme had 33% the activity of corn root enzyme. However, when proso millet shoot GSH-S-transferase was assayed in vitro using ¹⁴C-ring-labeled atrazine, it degraded the atrazine to water-soluble products at the same rate as the corn shoot enzyme. Incubation of excised proso millet and corn roots with [¹⁴C]EPTC indicated that uptake of EPTC was similar in both plants. However, proso millet metabolized the EPTC to water-soluble products at only half the rate of corn. Glutathione levels of proso millet roots were 35.9 μg GSH/g fresh wt, compared with 65.4 μg GSH/g fresh wt for corn. However, a 2.5-day pretreatment with R-25788 (N,N-diallyl-2-2-dichloroacetamide) elevated proso millet GSH levels to 62.7 μg GSH/g fresh wt. R-25788 did not elevate the activity of proso millet GSH-S-transferase, in contrast to its effects on corn. We conclude that differences in response to atrazine and EPTC in proso millet and corn are a result of their differential metabolism.     

DDE inhibition of marine algal cell division and photosynthesis per cell

Exuviella baltica, a marine dinoflagellate, was exposed to DDE, the major metabolite of DDT, at a concentration of 25 μg/liter of medium. At intervals of 1 hr and 1, 2, 3, and 4 days, cells were withdrawn from the culture, washed, and reseeded in DDE-free medium, and their growth (cell division) and photosynthesis were monitored for 14 days. No increase in cell numbers occurred until cells were removed from DDE, and lag phases, proportional to the duration of DDE exposure and lasting up to 5 days, preceded exponential growth. Cell densities comparable to controls were eventually reached in all treated cultures. A similar pattern of ¹⁴C uptake per milliliter of culture and per cell was observed. A 1-hr exposure to DDE resulted in a maximum reduction of 45% in carbon fixed per cell, while longer exposures caused reductions as great as 91%, relative to controls.     

Residues of DDT in cottonseed after spraying with DDT and torbidan

DDT at 1.12 and 2.24 kg/ha a.i. and Torbidan at 5 and 10 litre formulation/ha (1 and 2 kg DDT/ha) were sprayed five times on cotton over a period of 15 weeks. Seeds from the first pick of the crop were found to contain as residues pp′-DDT and pp′-DDE [1,1-dichloro-2,2-di- (4-chlorophenyl)ethylene]. The highest residue level (0.783 parts/10⁶) was found in seeds from Torbidan 10 litre/ha treatment.     

In vitro metabolism of pentachloronitrobenzene to pentachloromethylthiobenzene by onion: Characterization of glutathione S-transferase,cysteine C-S lyase,and S-adenosylmethionine methyl transferase activities

Pentachloromethylthiobenzene (PCTA) was synthesized in vitro from pentachloronitrobenzene (PCNB) at pH 7.9 by an enzyme system from onion root that required dithiothreitol, glutathione, and S-adenosylmethionine. The soluble enzyme system was isolated from onion root by ammonium sulfate fractionation and differential centrifugation. The system contained glutathione S-transferase activity with PCNB, C-S lyase activity with S-(pentachlorophenyl)cysteine, S-adenosylmethionine methyl transferase activity with pentachlorothiophenol (PCTP), and presumably several peptidase activities. All activities were stable when the crude enzyme system was stored at ?25°C. Evidence for the following sequence of reactions in PCTA synthesis was presented: PCNB→¹S-(pentachlorophenyl)glutathione→²S-(pentachlorophenyl)-γ-glutamylcysteine→³S-(pentachlorophenyl)cysteine→⁴ PCTP→⁵ PCTA. The first reaction was studied with [¹⁴C]PCNB. Reactions 2–4 were studied with S-([¹⁴C]pentachlorophenyl)glutathione, S-([¹⁴C]pentachlorophenyl)cysteine, and peptide inhibitors. Reaction 5 was studied with [¹⁴C]PCTP, S-[¹⁴C]adenosylmethionine, and inhibitors. The possible use of the enzyme system in the characterization of other glutathione conjugates was discussed.