Partial purification and properties of S-cysteinyl-hydroxychlorpropham transferase from oat (Avena sativa L.)

S-Cysteinyl and glutathione conjugates of isopropyl-3′-chloro-4′-hydroxycarbanilate (4-hydroxychlorpropham) were synthesized directly in the presence of soluble enzyme systems isolated from etiolated shoots of oat seedlings. The enzyme systems responsible for these reactions were partially purified and charaterized. Enzyme A appeared to be a multicomponent system, equally reactive with either cysteine or glutathione. Enzyme B was twice as active as enzyme A in the formation of S-cysteinyl-hydroxychlorpropham. Affinity chromatography of enzyme A produced an enzyme fraction with properties similar to those of enzyme B. Both enzymes (A and B) were significantly inhibited by increased cysteine concentrations. The reaction of glutathione with enzyme B was limited. However, when low concentrations of a nonreacting effector, cysteine ethyl ether, were added, glutathione conjugation increased significantly. At higher concentrations, cysteine ethyl ester formed a conjugate with 4-hydroxychlorpropham. Isopropyl-5′-chloro-2′-hydroxycarbanilate (2-hydroxy-5-chlorpropham) did not conjugate with either cysteine or glutathione but did react with cysteine ethyl ester. Isopropyl-3′-chlorocarbanilate (chlorpropham) was not a substrate for thioether conjugation. These data suggest that para- and/or ortho-hydroxylated carbanilates and cysteine-related substrates may form thioether conjugates when incubated under appropriate conditions with these complex enzyme systems.     

S-cysteinyl-hydroxychlorpropham: Formation of the S-cysteinyl conjugate of isopropyl-3′-chloro-4′-hydroxycarbanilate in oat (Avena sativa L.)

Isopropyl-3′-chlorocarbanilate (chlorpropham) forms phenolic metabolites, isopropyl-3′-chloro-4′-hydroxycarbanilate (I), and isopropyl-5′-chloro-2′-hydroxycarbanilate (II), in several plant species. In oat, which is a chlorpropham-susceptible plant, I was converted to an S-cysteinyl-conjugate (III). The reaction in vitro was catalyzed by a partially purified, soluble enzyme. The formation of III by the enzyme preparation and by oat shoot sections was compared. Mass spectral data indicated the presence of an aryl-thioether bond, and chloro-, hydroxy-, and isopropylcarbanilate groups in III. The results of this investigation indicate that III was isopropyl-[(2-amino-2-carboxyethyl)thio]-chloro-hydroxycarbanilate (S-cysteinyl-hydroxychlorpropham).     

Firefly luciferase inhibition by isopropyl-3-chlorocarbanilate and isopropyl-3-chloro-hydroxycarbanilate analogues

The firefly luciferase ATP assay was inhibited by the herbicide, isopropyl-3-chlorocarbanilate (I), and by two of its hydroxylated metabolites, isopropyl-5-chloro-2-hydroxycarbanilate (II) and isopropyl-3-chloro-4-hydroxycarbanilate (III). The β-O-glucosides of II and III reversed the inhibition of luciferase. Compounds I and II were linear noncompetitive inhibitors in respect to ATP (K_i ? 20 μM, each) and were linear competive inhibitors in respect to d-luciferin (K_i ? 6 μM, each). Compound III was a linear competitive inhibitor in respect to both ATP and d-luciferin (K_i ? 1 and 6 μM, respectively). The inhibition caused by III appeared to remain competitive for both substrates when AMP was added to the system, but the inhibition exhibited by III with respect to ATP and d-luciferin was more effective (K_i ? 0.5 μM, each). The effects of compounds I, II, and III upon the firefly luciferase ATP assay are discussed, and a relationship between the firefly system and plant susceptibility to compound I is proposed.     

Effect of isopropyl-3-chlorocarbanilate and isopropyl-3-chlorohydroxycarbanilate analogs upon oxidative phosphorylation in plant mitochondria

The effects of the herbicide, isopropyl-3-chlorocarbanilate, and its hydroxylated metabolites, isopropyl-5-chloro-2-hydroxycarbanilate and isopropyl-3-chloro-4-hydroxycarbanilate, upon NADH oxidation, P_i uptake or release, and ATP formation were studied in corn mitochondria. The results indicated that 0.1 mM isopropyl-3-chlorocarbanilate and the 2-hydroxy-metabolite inhibited NADH oxidation by 30% whereas only the 2-hydroxy-metabolite inhibited NADH-linked ATP formation (85–100%). Dinitrophenol and the 2-hydroxy-metabolite exerted similar effects upon respiration, phosphorylation, and ATPase activity. The 4-hydroxy-metabolite (0.1 mM) exerted no effect upon respiration, phosphorylation, or ATPase activity. The β-O-glucoside conjugates of the hydroxymetabolities of isopropyl-3-chlorocarbanilate did not inhibit NADH-linked respiration or phosphorylation at 0.1 mM concentrations. Comparative studies with corn, cucumber, and soybean mitochondria indicated that the parent herbicide and its metabolites affected respiration and phosphorylation activities in a similar manner.     

Enzyme activities and cytochrome P-450 levels of some Japanese quail tissues with respect to their metabolism of several pesticides

In the Japanese quail, cytochrome P-450, A- and B-esterase, amidase, and glutathione S-aryl transferase were assayed in postmitochondrial centrifugal fractions, in microsomes, and supernatant fractions of liver, lungs, kidneys, and testes. Liver microsomes contained the highest A-esterase activity and P-450 levels. B-esterase was more generally distributed and higher in the microsomal tissue fractions. Microsomal amidase activity was highest in quail lung and kidney, and lowest in the liver (per mg protein). Very little difference in glutathione S-aryl transferase activity was noted among the tissues assayed. In vitro metabolism of carbaryl, phosphamidon, and chlorotoluron by the various centrifugal fractions revealed that the production of 1-naphthyl-N-hydroxymethylcarbamate and 1-naphthol, the major metabolites, was greatest in the postmitochondrial fraction of the liver. The major carbaryl metabolite in all other quail tissue fractions was 1-naphthol. Phosphamidon metabolism in postmitochondrial preparations of quail liver was higher than in the supernatant and microsomes. Chlorotoluron metabolism occurred only in the postmitochondrial fractions of quail liver. The major products were the oxidative metabolites, N-(3-chloro-4-methylphenyl)-N′-methylurea and N-(3-chloro-4-hydroxymethylphenyl)-N′-methylurea.     

Metabolism of isopropyl carbanilate (propham) in alfalfa grown in nutrient solution

Alfalfa plants, Moapa variety, were grown in nutrient solution containing isopropylring-[¹⁴C] carbanilate (43.8 μCi/liter propham). After 8 days, 41.2% of the radioactivity initially added to the nutrient culture was recovered; 10.9% of this was from shoots, 3.4% from roots and 26.9% from nutrient medium. Nonextracted residues accounted for 23% of the radioactivity in shoots and 62% of that in roots. The parent herbicide constituted 53 and 38% of the radioactivity extracted from shoots and roots, respectively. The balance of extracted ¹⁴C was polar metabolites which were purified and subjected to enzymatic and acid hydrolysis. Four aglycones were isolated, three of which were purified by thin-layer chromatography and characterized by mass spectrometry. The principal aglycones were: isopropyl-2-hydroxycarbanilate, isopropyl-4-hydroxycarbanilate, and 1-hydroxy-2-propylcarbanilate. The fourth aglycone was not identified.     

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).     

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.     

Soybean shoot metabolism of isopropyl-3-chlorocarbanilate: Ortho and para aryl hydroxylation

This laboratory reported that isopropyl-3-chlorocarbanilate-phenyl-U-¹⁴C (chlorpropham-phenyl-¹⁴C) was absorbed, translocated, and metabolized by soybean plants. Both polar metabolites and insoluble residues were found in roots, whereas only polar metabolites were found in shoot tissues. In both roots and shoots the polar metabolites were shown to be the O-glucoside of isopropyl-2-hydroxy-5-chlorocarbanilate (2-hydroxy-chlorpropham). In shoot tissue there were other polar metabolites that were not identified. The experiments with soybeans have been repeated, but with new isolation and purification procedures. The plants were root treated with both chlorpropham-phenyl-¹⁴C and isopropyl-3-chlorocarbanilate-2-isopropyl-¹⁴C. The roots and shoots were extracted and separated into the polar, nonpolar, and insoluble metabolic components, using the Bligh-Dyer extraction method. The polar metabolites were separated by gel permeation chromatography. Further purification was accomplished on Amberlite XAD-2. The polar metabolites from the shoot and root tissues were hydrolyzed either by β-glucosidase or hesperidinase. The enzyme liberated aglycones were derivatized and separated by gas-liquid chromatography, and the components were characterized by mass spectrometry or NMR. The results of this study showed that the polar metabolites of soybean shoots were 2-hydroxy-chlorpropham and isopropyl-4-hydroxy-3-chlorocarbanilate (4-hydroxy-chlorpropham). These two hydroxy-chlorpropham metabolites were found in soybean shoots at a ratio of approximately 1:1. The only aglycone found in root tissue was 2-hydroxy-chlorpropham. Using the new procedures, no evidence was obtained for the presence of the unidentified polar metabolites that were previously observed in shoot tissues.     

噻二唑基-3-哒嗪酮类化合物的合成及生物活性

将取代的二酰基肼环合后,得到中间体2-芳基-5-氯甲基-1,3,4-噻二唑,然后与2-叔丁基-4-氯-5-羟基-3(2H)-哒嗪酮反应,合成了8个未见文献报道的含噻二唑基哒嗪酮类化合物,其化学结构经1H NMR、高分辩质谱和元素分析确认。生物活性测试结果表明,部分化合物对粘虫P.separate W.有较好的抑制生长活性,其中化合物 3b 的EC50值为21 mg/L。     

Effect of herbicide antidotes on glutathione content and glutathione S-transferase activity of sorghum shoots

The effects of the herbicide antidotes CGA-92194 (α-[(1,3-dioxolan-2-yl-methoxy)-imino]benzeneacetonitrile), flurazole [phenylmethyl 2-chloro-4-(trifluoromethyl)-5-thiazolecarboxylate], dichlormid (2,2-dichloro-N,N-di-2-propenylacetamide), and naphthalic anhydride (1H,3H-naphtho(1,8-cd)-pyran-1,3-dione) on nonprotein thiol content, glutathione content, and glutathione S-transferase (GST) activity in etiolated sorghum (Sorghum bicolor L.) Moench) shoots were examined. CGA-92194 and naphthalic anhydride had no effect on nonprotein thiol or reduced glutathione (GSH) content of sorghum shoots. In contrast, dichlormid and flurazole increased nonprotein thiol content of sorghum shoots by 24 and 48%, respectively. These increases were largely attributable to an increase in GSH. The antidotes increased GST activity less than twofold when using CDNB (1-chloro-2,4-dinitrobenzene) as a substrate. In contrast, when using metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide] as a substrate, the increase in GST activity in response to antidote treatment was much greater: flurazole (30-fold), CGA-92194 (20-fold), naphthalic anhydride (17-fold), dichlormid (5-fold). The degree of protection from metolachlor injury conferred by a particular antidote was strongly correlated (R² = 0.95) with its ability to enhance GST activity, as evaluated with metolachlor as substrate. A comparison of GST activity in untreated and CGA-92194-treated seedlings, over a range of metolachlor concentrations (0.5–500 μM), indicated that the relative enhancement of enzyme activity by CGA-92194 was greater at lower metolachlor concentrations. The rate of nonenzymatic conjugation of metolachlor and GSH in vitro was much less (on a gram fresh weight equivalent basis) than the enzymatic rate. These results are consistent with the hypothesis that the above antidotes protect sorghum by enhancing GST activity which results in accelerated detoxification of metolachlor via GSH conjugation.     

The effect of phenobarbital induction on glutathione conjugation of diazinon in susceptible and resistant house flies

The relationship between glutathione S-transferase activity toward 3,4-dichloronitrobenzene and O-alkyl or O-aryl conjugation of diazinon was investigated in eight strains of house flies. No significant difference was found in the amount of O-aryl conjugation. In contrast, house flies which had higher glutathione S-transferase activity toward 3,4-dichloronitrobenzene also had higher O-alkyl conjugating activity toward diazinon. The glutathione S-transferase(s) in phenobarbital-pretreated flies degraded diazinon faster than those in the nontreated ones. The present results showed that the formation of the O-alkyl conjugate was enhanced by phenobarbital pretreatment, while the formation of the O-aryl conjugate was not affected by induction. Based on these findings, it would appear that one of the multiple forms of glutathione S-transferase is specifically induced and responsible for the increase in O-alkyl conjugation.     

Metabolism of methyl bromide by susceptible and resistant strains of the granary weevil, Sitophilus granarius (L.)

Methyl bromide was metabolized by susceptible and resistant strains of adult granary weevil, Sitophilus granarius (L.), mainly by conjugation with glutathione. S-Methyl glutathione and S-methyl cysteine were produced by both strains and S-methyl glutathione sulfoxide was identified as a metabolite in the resistant strain. In the untreated insects, no significant difference was observed in glutathione S-transferase activity but the resistant contained approximately twice as much glutathione per insect as the susceptible strain. When the insects were treated with methyl bromide, the glutathione content of both strains was lowered; proportionally, however, the decrease was considerably higher in the susceptible than in the resistant strain. These results indicate that conjugation of methyl bromide with glutathione is a major detoxication pathway and tolerance to this fumigant is related, in part at least, to the level of glutathione in the granary weevil.     

Inhibition of carotenogenesis by substituted diphenyl ethers of the m-phenoxybenzamide type

Diphenyl ethers exhibit different modes of action according to their chemical constitution. Diphenyl ethers of the m-phenoxybenzamide type, which were found to be effective on carotenogenesis resulting in an accumulation of colorless carotenoid precursors, mostly phytoene, indicative of inhibition of desaturation, are discussed. As seen with other carotenoid biosynthesis inhibitors, a concurrent loss of chlorophyll was observed as a secondary effect caused by the absence of protective carotenoids. In contrast to peroxidative p-nitrodiphenyl ethers like oxyfluorfen (2-chloro-4-trifluoromethylphenyl-3′-ethoxy-4′-nitrophenyl ether), the m-phenoxybenzamides assayed showed the same phytotoxic mode of action in the dark as observed when using heterotrophic Scenedesmus cultures. As expected, chlorophylls were not affected. The decrease of carotenoids was not due to their degradation but to inhibited carotenogenesis. Examination of carotenoid fractions show that the m-phenoxybenzamides, e.g., 3-(2,5-dimethylphenoxy)-N-ethylbenzamide, used here act similarly to 2-phenylpyridazinones like norflurazon [4-chloro-5-methylamino-2-(2-trifluoromethylphenyl)-pyridazin-3(2H)one]. All these inhibitors strongly decrease the α- and β-carotene content, while xanthophyll content is not lowered as much.     

Studies on residues and the metabolic pathway of methidathion in tomato fruit

The residues and metabolism of methidathion [S-(2, 3-dihydro-5-methoxy-2-oxo-1, 3, 4-thiadiazol-3-ylmethyl) O, O-dimethyl phosphorodithioate] and its secondary metabolites: demethyl-methidathion [S-(2, 3-dihydro-5-methoxy-2-oxo-1, 3, 4-thiadiazol-3-ylmethyl) O-methyl O-hydrogen phosphorodithioate] ( IV ), the sulphide (2,3-dihydro-5-methoxy-3-methylthiomethyl-1,3,4-thiadiazol-2-one) ( I ), tsulphoxide(2,3-dihydro-5-methoxy-3- methylsulphinylmethyl-1,3,4-thiadiazol-2-one) ( II ) and the sulphone (2,3-dihydro-5-methoxy-3-methylsulphonylmethyl-1,3,4-thiadiazol-2-one ( III ) were studied in laboratory-treated tomato fruit. The metabolites and residues of methidathion were determined for the applied doses of 1, 7 and 14 mg of methidathion kg^?1 of fruit. Methidathion was metabolised extensively over a 14-day period. The amount of metabolites formed was a function of both the applied dose as well as the time after application. Major water-soluble metabolites were found to be IV and the cysteine conjugate S-(2,3-dihydro-5-methoxy-2-oxo-1,3,4-thiadiazol-3-ylmethyl)-L-cysteine ( VI ). The chloroform-soluble metabolites were identified as the oxygen analogue of methidathion [S-(2,3-dihydro-5-methoxy-2-oxo-1,3,4-thiadiazol-3-ylmethyl) O, O-dimethyl phosphorothioate] ( V ), the sulphoxide II , and the hydroxy compound 2,3-dihydro-3-hydroxymethyl-5-methoxy-1,3,4-thiadiazol-2-one. The oxygen analogue of methidathion ( V ) was found in small amounts, corresponding to <5% of the added methidathion. Demethyl-methidathion ( IV ) appeared to be a precursor in the formation of the cysteine conjugate VI . The sulphide I seemed to be more reactive in forming the cysteine conjugate than the sulphoxide II or the sulphone III .     

Escherichia coli dihydrodipicolinate synthase and dihydrodipicolinate reductase: kinetic and inhibition studies of two putative herbicide targets

Dihydrodipicolinate synthase (DHDPS) (EC 4.2.1.52) and dihydrodipicolinate reductase (DHDPR) (EC 1.3.1.26) have attracted much recent attention as potential herbicide targets. DHDPS was feedback-inhibited by (S)-lysine; inhibition was reversible and uncompetitive with respect to both (S)-ASA and pyruvate. Homoserine lactone was a reversible non-competitive inhibitor of DHDPS with respect to both (S)-ASA and pyruvate. (R)-Cysteine sulfinic acid and (S)-glutamic acid were reversible uncompetitive inhibitors of DHDPS with respect to (S)-ASA. (S)-Aspartic acid was a reversible mixed-type inhibitor. Dipicolinic acid was a reversible competitive inhibitor of DHDPR with respect to the substrate (4S)-4-hydroxy-2,3,4,5-tetrahydro-(2S)-dipicolinic acid, as was isophthalic acid. Δ³-Tetrahydroisophthalic acid was a moderate inhibitor of both DHDPS and DHDPR. These compounds represent possible leads in the development of novel herbicides. © 1999 Society of Chemical Industry     

Insect glutathione S-transferase: Separation of transferases from fat bodies of American cockroaches active on organophosphorus triesters

Several glutathione S-transferases which catalyze the conjugation of reduced glutathione with organophosphorus triesters were separated from fat bodies of adult female American cockroaches, Periplaneta americana (L.). Two transferases (I, V) were active on diazinon and three transferases (II, III, IV) were active on methyl parathion. The transferase (I) active on the pyrimidinyl moiety of diazinon was distinguishable from the other transferases on the O-methyl portion of methyl parathion, as shown by chromatographic properties, and additionally it was almost inactive or less active on 3,4-dichloronitrobenzene, methyl iodide, p-nitrobenzyl chloride, trans-cinnamaldehyde, and 1,2-epoxy-3-(p-nitrophenoxy)propane. Transferase II had high activities with “aryl” and “aralkyl” compounds, transferase III with “epoxide” and “alkene,” and transferase IV with “alkyl,” “aryl,” and “aralkyl” compounds. This indicated that the transferases had overlapping substrate specificities. The molecular weight was 35,000–37,000 for both of the enzymes active on methyl parathion and diazinon. The pH optima with methyl parathion and diazinon were about 8.5 and 6.5, respectively. At a glutathione concentration of 5 mM, Michaelis constants were 0.28 and 0.13 mM for methyl parathion and diazinon, respectively.     

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.     

Delayed acute toxicity of O,S,S-trimethyl phosphorodithioate and O,O,S-trimethyl phosphorothioate to the rat

The toxicity and LD₅₀ of O,S,S-trimethyl phosphorodithioate were reexamined in the rat. Animals treated orally (single dose) with this compound exhibited early cholinergic signs followed at approximately 5 hr by delayed toxic signs, with an LD₅₀ of 43 mg/kg. Contamination of O,S,S-trimethyl phosphorodithioate by as much as 5% (w/w) O,O,O-trimethyl phosphorothioate provided only limited antagonism to the dithioate's toxicity. In contrast, the addition of 5% O,O,S-trimethyl phosphorodithioate to O,O,S-trimethyl phosphorothioate gave protection against the toxic effects of the latter compound up to 80 mg/kg of toxicant. Pretreatment of rats with as little as 5% O,O,O-trimethyl phosphorothioate, 24 hr prior to treatment with 200 mg/kg O,O,S-trimethyl phosphorothioate, gave complete protection against the toxic effects of this compound. Conversely, administration of 10% (w/w) O,O,O-trimethyl phosphorothioate 4 or 24 hr after treatment with 60 or 80 mg/kg of O,O,S-trimethyl phosphorothioate provided only partial protection at 4 hr and no protection from the effects of the toxicant at 24 hr. The ability of O,O,O-trimethyl phosphorothioate to antagonize the toxicity of this compound depended markedly on the route of administration (oral, intravenous, or intraperitoneal). At 4 hr past treatment with toxicant, only oral administration of the antagonist provided full protection. Intraperitoneal and intravenous administration of antagonist 4 hr after treatment with toxicant were partially effective and completely ineffective, respectively, in halting the toxic effects of this compound.     

Effects of piperonyl butoxide and DEF on the metabolism of methoprene by the imported fire ant,Solenopsis invicta Buren

Studies are presented on the effects of two synergists, piperonyl butoxide and S,S,S-tributyl phosphorotrithioate, on the metabolism of methoprene [isopropyl (2E,4E)-11-methoxy-3,7,11-trimethyldodeca-2,4-dienoate], an insect growth regulator, by the castes of the imported fire ant, Solenopsis invicta Buren. In adults, but not in larvae, pharate pupae, and pupae, piperonyl butoxide, a microsomal enzyme inhibitor, reduced methoprene metabolism by blocking O-demethylation. S,S,S-Tributyl phosphorotrithioate, an esterase and microsomal oxidase inhibitor, was most effective in reducing methoprene metabolism in larvae. In toxicity studies, against pharate pupae, the O-demethylated methoprene metabolite (alcohol-ester) was shown to be more toxic than methoprene. Synergists may be useful in bait formulations used for imported fire ant control to extend the effectiveness of methoprene.