Dose-related inhibition of brain and plasma cholinesterase in neonatal and adult rats following sublethal organophosphate exposures.

Developing mammals are markedly more sensitive to acute toxicity from exposure to a variety of organophosphorus (OP) pesticides. The present study examined dose-related inhibition of both brain and plasma cholinesterase activity in neonatal and adult rats exposed to sublethal doses of one of three common OP pesticides, methyl parathion, parathion and chlorpyrifos. Effective dose 50 (i.e., ED50 or dose which would inhibit 50% of the cholinesterase activity) values were determined and then correlated with an indicator of acute toxicity, the maximal tolerated dose (MTD). It was found that ED50 estimates for both brain and plasma cholinesterase correlated highly (r = 0.932-0.992) with previously derived MTD values. In no case was there a significant difference between in vivo brain and plasma cholinesterase inhibition across doses in neonatal rats was high (r = 0.962-0.975) but lower in adults (r = 0.700-0.943). The results suggest that in vivo inhibitory potency of the three OPs towards either brain or plasma ChE activity is highly correlated with sensitivity to acute toxicity in both neonatal and adult rats. Additionally, under defined experimental conditions, plasma ChE inhibition may be a useful quantitative index for the degree of brain cholinesterase inhibition following OP exposures.     

Effects of Acute Exposure to Methyl Parathion on Carbohydrate Metabolism of Indian Catfish (Heteropneustes fossilis)

Abstract: Exposure of the Indian catfish (Heteropneustes fossilis) to a high sublethal concentration of 5.6 p.p.m. (0.8 of the 96 hr LC50) of methyl parathion for 3,6,12,48, and 96 hrs affected carbohydrate metabolism. Muscle glycogen levels decreased significantly at 3,6,12 and 96 hrs; liver glycogen content declined at 6 hrs but there was a resynthesis of hepatic glycogen stores at 12 hrs. Blood glucose levels in fish were elevated at 3 and 6 hrs. Mean values for blood pyruvate were elevated significantly at 6,12, and 96 hrs. Blood lactate level was elevated at 3 hrs but hypolactaemia resulted at 48 and 96 hrs in pesticide-treated fish. The observed effects of methyl parathion on carbohydrate metabolism in fish are discussed in relation to acute stress syndrome. Measurement of carbohydrate metabolites in fish for 3 hrs or longer could prove useful as a rapid method for evaluating toxicity of pesticides and other toxicants.     

Comparative toxicity, anticholinesterase action and metabolism of methyl parathion and parathion in sunfish and mice

The in vitro metabolism of methyl parathion (O,O-dimethyl O-p-nitrophenyl phosphorothioate) and parathion (O,O-diethyl O,p-nitrophenyl phosphorothioate) and the sensitivities of the target cholinesterases to inhibition by their oxygen analogs were studied in sunfish (Lepomis gibbosus) and mice to determine the basis for the low toxicity of methyl parathion in sunfish (LD50 > 2500 mg/kg). The LD50 values of parathion and methyl parathion in mice were 13.5 and 11 mg/kg, respectively, and the times to death were much shorter for both compounds in mice than in fish. Low sensitivity of fish cholinesterases to paraoxon as compared to mice accounted for the 10-fold lower toxicity of parathion in fish (LD50, 110 mg/kg). By contrast, sunfish had similar cholinesterase sensitivities to methyl paraoxon and paraoxon. Differences in rates of oxidative formation of the oxygen analog or oxidative cleavage to p-nitrophenol and the corresponding dialkyl thiophosphate could not account for the selective resistance of sunfish to methyl parathion toxicity. Fish and mouse liver homogenates catalyzed a glutathione (GSH)-dependent metabolism of methyl parathion and methyl paraoxon but not of parathion or paraoxon. Additionally, hydrolysis of methyl paraoxon by fish liver homogenates exceeded that for parathion by 5-fold, while methyl paraoxon hydrolysis in mice was $\text{1}\text{2}$ of that of paraoxon. Apparently, a longer time to death in fish provided the opportunity for GSH-dependent and hydrolytic detoxification, which favored methyl parathion and methyl paraoxon relative to parathion and paraoxon. Although in mice the GSH-dependent enzymes also favored detoxification of methyl parathion and methyl paraoxon, this is apparently of less importance because of their high cholinesterase sensitivity and because cleavage and hydrolysis favored parathion and paraoxon.     

Involvement of oxidative stress in methyl parathion and parathion‐induced toxicity and genotoxicity to human liver carcinoma (HepG2) cells

Methyl parathion (C₈H₁₀NO₅PS) and parathion (C₁₀H₁₄NO₅PS) are both organophosphate insecticides (OPI) widely used for household and agricultural applications. They are known for their ability to irreversibly inhibit acetylcholinesterase which often leads to a profound effect on the nervous system of exposed organisms. Many recently published studies have indicated that human exposure to OPI may be associated with neurologic, hematopoietic, cardiovascular, and reproductive adverse effects. Studies have also linked OPI exposure to a number of degenerative diseases including Parkinson's, Alzheimer's, and amyotrophic lateral sclerosis. Also, oxidative stress (OS) has been reported as a possible mechanism of OPI toxicity in humans. Hence, the aim of the present investigation was to use human liver carcinoma (HepG₂) cells as a test model to evaluate the role of OS in methyl parathion‐ and parathion‐induced toxicity. To achieve this goal, we performed the MTT [3‐(4, 5‐dimethylthiazol‐2‐yl)‐2, 5‐diphenyltetrazolium bromide] assay for cell viability, lipid peroxidation assay for malondialdehyde (MDA) production, and Comet assay for DNA damage, respectively. Results from MTT assay indicated that methyl parathion and parathion gradually reduce the viability of HepG₂ cells in a dose‐dependent manner, showing 48 h‐LD₅₀ values of 26.20 mM and 23.58 mM, respectively. Lipid peroxidation assay resulted in a significant increase (P < 0.05) of MDA level in methyl parathion‐ and parathion‐treated HepG₂ cells compared with controls, suggesting that OS plays a key role in OPI‐induced toxicity. Comet assay indicated a significant increase in genotoxicity at higher concentrations of OPI exposure. Overall, we found that methyl‐parathion is slightly less toxic than parathion to HepG₂ cells. The cytotoxic effect of these OPI was found to be associated, at least in part, with oxidative cell/tissue damage. © 2011 Wiley Periodicals, Inc. Environ Toxicol, 2013.     

Rapid toxicity assessment using esterase biomarkers in Brachionus calyciflorus (rotifera)

We have developed biomarkers of sublethal toxicity in the freshwater rotifer Brachionus calyciflorus based on the reduction of enzyme activity. Esterase and phospholipase A2 activity was quantified in single rotifers using image analysis and a fluorescence detection system. Esterase activity was localized in the gut and phospholipase A2 activity in the corona of females. Quantitation of enzyme activity demonstrated that toxicant stress reduced activity in a dose-dependent manner. Concentration-response relationships are described for 10 compounds representing a variety of toxicant classes and NOECs are reported. Esterase and phospholipase A2 activities were generally less sensitive end points than reproduction NOECs, but usually were more sensitive than LC₅₀s. Since in vivo enzyme activity can be assessed in 1 h, these biomarkers will be useful where rapid results are important. The cost of performing in vivo enzyme inhibition tests is substantially less than traditional whole animal tests because these require three times more person-hours to execute. Obtaining test animals by hatching cysts, their sensitivity to toxicants, and the rapid results make the rotifer esterase and phospholipase A2 tests good candidates for inclusion in a test battery for rapid toxicity assessment. © 1994 by John Wiley & Sons, Inc..     

Using Proales similis (Rotifera) for toxicity assessment in marine waters

There is a need to develop more animal species for assessing toxicity in marine environments. Cyst‐based toxicity tests using invertebrates are especially fast, technically simple, cost‐effective, and sensitive to a variety of toxicants. Over the past 30 years, a variety of toxicity endpoints have been measured using the marine rotifer Brachionus plicatilis hatched from cysts, including mortality, reproduction, ingestion, swimming, enzyme activity, and gene expression. A consensus has developed that the most ecologically relevant toxicity measurements should be made using more than one species. Furthermore, it has been noted that the rotifer species toxicant sensitivity distribution is much broader than which endpoint is measured. This implies that toxicity should be measured with the simplest, fastest, least expensive test available on as many species as feasible. If a battery of test species is to be used to estimate toxicity, diapause egg‐based toxicity tests that do not require culturing of test animals will be key. In this paper, we describe how diapause eggs of a new marine rotifer, Proales similis, can be produced, stored and hatched under controlled conditions to produce animals for toxicity tests. Methods are described for quantifying the toxicity of copper, mercury and cadmium based on mortality, ingestion, reproduction, and diapause egg hatching endpoints. We found that reproduction and ingestion endpoints were generally more sensitive to the metals than mortality or diapause egg hatching. When the copper sensitivity of P. similis was compared to Brachionus manjavacas and B. plicatilis using an ingestion test, similar EC50s were observed. In contrast, the B. rotundiformis ingestion EC50 for copper was about 4× more sensitive. Although diapause egg hatching was not the most sensitive endpoint, it is the most ecologically relevant for assessing sediment toxicity. Our discovery of diapausing eggs in the P. similis life cycle has created a conundrum. We have not observed males or sex in P. similis populations, which is a direct contradiction to the orthodox view of the monogonont rotifer life cycle. Work is needed to clarify how diapause egg production is accomplished by P. similis and whether sexual reproduction is involved.     

Acute toxicity tests using rotifers. III. Effects of temperature,strain, and exposure time on the sensitivity of Brachionus plicatilis

As part of the development of a standardized acute toxicity test, the effect of cyst age, strain, temperature, and exposure time on the toxicity of 21 chemicals to the estuarine rotifer Brachionus plicatilis was investigated. Toxicity was chemical specific, with LC₅₀s ranging from 0.061 mg · L^?1 for mecury to 598 mg · L^?1 for 2,4-dichlorophenoxyacetic acid. Intralaboratory coefficients of variation averaged 11%, at least three times lower than for other aquatic invertebrate acute tests. The age of rotifer cysts stored up to 27 months had no effect on the sensitivity of test animals, but significant differences in sensitivity were detected among rotifer strains. Test temperatures of 25, 30, and 35°C generally yielded lower LC₅₀s than at 20°C. LC₅₀s decreased by 80–90% for cadmium and pentachlorophenol when toxicant exposure time was increased from 24 to 72 h. Life table analysis of rotifer survival in the controls revealed that 72 h is the longest acute test possible without feeding. A comparison of the sensitivity of the rotifer test to that of sea urchin (Arabacia punctulata) early embryo, sea urchin sperm cell, Microtox®, and Mysidopsis bahia tests revealed comparability for several compounds. However, no species is consistently the most sensitive to all compounds.     

Effect of piperonyl butoxide on the metabolism of dimethyl and diethyl phosphorothionate insecticides

The effect of piperonyl butoxide on the acute toxicity of phosphorothionate insecticides was studied in male mice. One hour after piperonyl butoxide (400 mg/kg), the toxicity of the dimethyl phosphorothionates, methyl parathion and Guthion, was antagonized, whereas the toxicity of their respective diethyl homologs, parathion and Ethyl Guthion, was potentiated. Piperonyl butoxide did not appreciably alter the toxicity of the oxygen analogs of these compounds. Pretreatment with SKF 525-A (50 mg/kg) modified the toxicity of the phosphorothionates in a manner qualitatively similar to piperonyl butoxide pretreatment. Plasma concentrations of all four insecticides were increased three- to sevenfold in piperonyl butoxide-pretreated mice. This increase may result in a greater total oxon formation; however, reactivation in vitro of esterases inhibited in vivo was 5 to 10 times more rapid following methyl parathion or Guthion challenge than after their diethyl homologs. Although a greater total oxon formation-cholinesterase inhibition is possible for both dimethyl and diethyl phosphorothionates following piperonyl butoxide pretreatment, rapid reactivation of inhibited nerve tissue cholinesterases after dimethyl phosphorothionate challenge appears to compensate for further inhibition occurring at a decreased rate. The net result would be a reduction in dimethyl phosphorothionate toxicity. In contrast, slow reactivation of inhibited nerve tissue cholinesterases following diethyl phosphorothionate challenge appears unable to compensate for increased oxon formation-cholinesterase inhibition. The net result is a potentiation of the toxicity of the diethyl-substituted compounds.     

Toxicity of triphenyltin chloride to the rotifer Brachionus koreanus across different levels of biological organization

Although triphenyltin (TPT) compounds are ubiquitous pollutants in urbanised coastal environments in Asian regions, their toxicities to marine organisms are still poorly known. This study was designed to investigate the toxicity of triphenyltin chloride (TPTCl) on the rotifer Brachionus koreanus across different levels of biological organisation. Firstly, we concurrently performed a 24 h static‐acute toxicity test and a 6‐day semi‐static multigenerational life‐cycle test using the rotifer. Our results demonstrated that the 24‐h median lethal concentration of TPTCl for the rotifer was 29.6 μg/L and the 6‐day median effect concentration, based on the population growth inhibition, was 3.31 μg/L. Secondly, we examined the expression of 12 heat shock protein (hsp) genes, four glutathione S‐transferase (GST) genes, one retinoid X receptor (RXR) gene and 13 cytochrome P450 (CYP) genes in the rotifers after exposure to 20 µg/L TPTCl for 24 h. Among these studied genes, hsp90α2, GST‐O and CYP3045C1 were the most significantly up‐regulated genes with a relative expression level up to 32.9, 4.4 and 62.6 folds, respectively. The expression of these three genes in the rotifers showed an increasing trend in the first few hours of TPTCl exposure, peaked at 3 h (hsp90α2 and GST‐O) and 12 h (CYP3045C1) respectively, and then gradually returned to a lower level at 24 h. Such up‐regulations of hsp and GST genes probably offer cellular protection against the TPT‐mediated oxidative stress while the accelerated induction of CYP genes possibly facilitates the detoxification of this toxicant in the rotifer. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 13–23, 2016.     

Potential use of the oligochaete Limnodrilus profundicola V., as a bioindicator of contaminant exposure

Cholinesterase (ChE) and ethoxyresorufin‐O‐deethylase (EROD) were of special interest to this study as these biochemical tools have been widely used for the determination of exposure to pollutants. In this study, the freshwater oligochaete Limnodrilus profundicola was tested for its potential as a bioindicator of freshwater pollution. For this purpose, the ChE and EROD activities of L. profundicola and the level of polycyclic aromatic hydrocarbons (PAH) of water samples collected from different sites along the Curuksu stream on the Menderes River (the ancient Meander) running through south‐western Turkey were studied. First, these activities were characterized using, as model substrates, acetylthiocholine (ATC), propionylthiocholine (PTC), and butyrylthiocholine (BTC). Then, the in vivo effects of insecticides and pollutants on these activities were investigated. L. profundicola were exposed to various doses of methyl‐parathion, methomyl, and deltamethrin. Although significant inhibition of ChE was detected with each of the insecticides, the highest level of inhibition was observed with methyl‐parathion. In addition to the inhibition of ChE, the activity of EROD was induced by exposure to oil‐contaminated sediments. Thus, although L. profundicola has a reputation for being very resistant to pollution (although it is not insensitive to it), we demonstrated that it may potentially be used as a bioindicator species for contaminant exposure when ChE and EROD are used as biomarkers. © 2010 Wiley Periodicals, Inc. Environ Toxicol 26: 37–44, 2011.     

Assessment of the lethal and sublethal effects of 20 environmental chemicals in zebrafish embryos and larvae by using OECD TG 212

Fish embryo toxicity tests are used to assess the lethal and sublethal effects of environmental chemicals in aquatic organisms. Previously, we used a short‐term toxicity test published by the Organization for Economic Co‐operation and Development (test no. 212: Fish, Short‐term Toxicity Test on Embryo and Sac‐Fry Stages [OECD TG 212]) to assess the lethal and sublethal effects of aniline and several chlorinated anilines in zebrafish embryos and larvae. To expand upon this previous study, we used OECD TG 212 in zebrafish embryos and larvae to assess the lethal and sublethal effects of 20 additional environmental chemicals that included active pharmaceutical ingredients, pesticides, metals, aromatic compounds or chlorinated anilines. Zebrafish embryos (Danio rerio ) were exposed to the test chemicals until 8 days post‐fertilization. A delayed lethal effect was induced by 16 of the 20 test chemicals, and a positive correlation was found between heart rate turbulence and mortality. We also found that exposure to the test chemicals at concentrations lower than the lethal concentration induced the sublethal effects of edema, body curvature and absence of swim‐bladder inflation. In conclusion, the environmental chemicals assessed in the present study induced both lethal and sublethal effects in zebrafish embryos and larvae, as assessed by using OECD TG 212. Copyright © 2017 John Wiley & Sons, Ltd.     

Mass spectrometric detection of CYP450 adducts following oxidative desulfuration of methyl parathion

Cytochrome P450 (CYP)‐mediated desulfuration of methyl parathion results in mechanism‐based inhibition of the enzyme. Although previous data suggest that reactive sulfur is released and binds to the apoprotein, the identities of neither the adduct(s) nor the affected amino acid(s) have been clearly determined. In this study, nanospray tandem mass spectroscopy was used to analyze peptide digests of CYP resolved by SDS–PAGE from liver microsomes of male rats following incubation in the absence or presence of methyl parathion. Oxidative desulfuration was confirmed by measurement of methyl paraoxon, and inhibition of specific CYP isozymes was determined by measurement of testosterone hydroxylation. Total CYP content was quantified spectrophotometrically. Incubation of microsomes with methyl parathion decreased CYP content by 58%. This effect was not associated with a comparable increase in absorbance at 420 nm, suggesting the displacement of heme from the apoprotein. Rates of testosterone 2β‐ and 6β‐hydroxylation, respectively, were reduced to 8 and 2%, implicating CYP3A and CYP2C11 in the oxidative desulfuration of methyl parathion. Mass spectrometric analysis identified 96 amu adducts to cysteines 64 and 378 of CYP3A1. In addition, a peptide containing cysteine 433 that coordinates with heme was possibly modified as it was detected in control, but not methyl parathion samples. A comparison of rat CYP3A1 with human CYP3A4 suggests that cysteines 64 and 378 reside along the substrate channel, remote from the active site. Alteration of these residues might modulate substrate entry to the binding pocket of the enzyme. Copyright © 2012 John Wiley & Sons, Ltd.     

Toxicity of parathion on embryo and yolk‐sac larvae of gilthead seabream (Sparus aurata l.): Effects on survival,cholinesterase, and carboxylesterase activity

This study was conducted to examine the acute toxicity of the organophosphorus pesticide (OP) parathion on embryos and yolk‐sac larvae of gilthead seabream (Sparus aurata), and to investigate the effects of this compound on cholinesterase and carboxylesterase activity of seabream larvae in the phase of endogenous feeding. The 72‐h LC50 for yolk‐sac larvae (0.523 mg L^?1) was about two‐fold lower than the 48‐h LC50 for embryos (1.005 mg L^?1). Parathion significantly inhibited the activity of ChE and CaE activity in yolk sac larvae but there were not significant differences in the sensitivity of both esterases to parathion as inferred by their 72‐h IC50 values. Larvae exposed to parathion for 72 h showed a 70% inhibition of the whole body acetylcholinesterase at approximately the LC50. © 2009 Wiley Periodicals, Inc. Environ Toxicol 25: 601–607, 2010.     

Methyl parathion activation by a partially purified rat brain fraction

Organophosphorus pesticides are one of the most commonly used insecticide classes. They act through a potent inhibition of acetylcholinesterase (AChE). Many of them must undergo transformation into the corresponding oxon analogs to inhibit AChE. This study showed that a brain tissue subfraction transformed methyl parathion (O,O-dimethyl O-p-nitrophenyl phosphorothioate) in vitro. Methyl parathion activation was assayed by solvent extraction of the products followed by HPLC and GC-MS analyses and, indirectly, by the inhibition of AChE present in the incubation mixture. The lack of impairment of AChE after 2 h of incubation of the brain subfraction with methyl parathion and, alternatively, with NADPH, CO, SKF 525-A, piperonyl butoxide or nitrogen indicated that this brain subfraction transformed methyl parathion without the involvement of a mixed-function oxidative pathway.The results from HPLC analysis did not show a peak corresponding to methyl paraoxon (O,O-dimethyl O-p-nitrophenylphosphate), but showed the production of an unidentified peak which eluted nearby standard methyl parathion (retention times of 10.65 and 8.86 min, respectively). GC-MS analysis suggested that the unidentified product could be a methyl parathion isomer.     

Consideration of dosimetry in evaluation of ToxCast™ data

The US Environmental Protection Agency (US EPA) Toxcast? program has the stated goal of predicting hazard, characterizing toxicity pathways and prioritizing the toxicity testing of environmental chemicals through the use of in vitro high‐throughput screening (HTS) assays. This analysis integrates data from biomonitoring and from in vivo toxicity and pharmacokinetic studies to examine the physiological relevance of the tested and responding in vitro concentrations for five case study chemicals: triclosan, 2,4‐dichlorophenoxyacetic acid, perfluorooctanoic acid, monobutyl phthalate and mono‐2(ethylhexyl)phthalate. This analysis also examines the ToxCast? phase 1 data set for approximately 50 chemicals belonging to four ‘common mechanism groups’ which have been the subject of cumulative risk assessments by the US EPA for both the pattern of key responses and the relative potencies of included chemicals compared with the in vivo relative potencies. Responding concentrations in vitro were generally in the range of serum or plasma concentrations associated with no‐observed to lowest‐observed effect levels for the case study chemicals, while available biomonitoring data demonstrating actual exposures were generally lower. ToxCast? assay endpoints related to acetylcholinesterase (AChE) inhibition had low sensitivity for detecting organophosphate pesticides but good sensitivity for detecting N‐methyl carbamates. However, in vitro relative potencies did not correlate with in vivo potency. Both qualitative and quantitative predictive power is probably affected by the lack of comprehensive metabolic activity in most current in vitro systems explored in the ToxCast? program, and this remains a fundamental challenge for high‐throughput toxicity screening efforts. Copyright © 2011 John Wiley & Sons, Ltd.     

Tissue lipid levels of the clam,Villorita cyprenoides var. cochinensis,following exposure to endosulfan,malathion, and methyl parathion

Sublethal stress of toxicants on organisms is generally measured through metabolic changes. The effects of three pesticides (endosulfan, malathion, and methyl parathion) on tissue lipid levels in the estuarine bivalve, Villorita cyprenoids var. cochinensis, have been measured. The reduction in lipid content was expressed as a function of the pesticide concentrations and the duration of the exposure periods (24, 48, 72, and 96 h). The results point to the existence of a direct relationship between the applied stress and the fall in the lipid content. The order of toxicity was endosulfan > malathion > methyl parathion. The depletion in lipid content observed may reflect an adaptive response of the organism to pesticide stress. Thus lipids in bivalves have been shown to be an important reserve material that may be utilized under stressed conditions as an energy source. © by John Wiley & Sons, Inc.     

The influence of age on the toxicity and metabolism of methyl parathion and parathion in male and female rats

To determine the mechanisms responsible for the variations in toxicity of methyl parathion and parathion, the in vitro metabolism of these insecticides and cholinesterase sensitivity to their respective oxygen analogs methyl paraoxon and paraoxon were studied in male and female rats of several ages. For rats of five ages studied (1, 12–13, 23–24, 35–40, and 56–63 days), there was a gradual decrease in susceptibility to poisoning by both insecticides with increasing age up to 35–40 days for both sexes. Age differences in susceptibility were not related to differences in sensitivity of cholinesterase to inhibition by paraoxon or methyl paraoxon in vitro. Oxidative formation of the oxygen analogs, oxidative aryl cleavage, and glutathione-dependent dealkylation and dearylation of methyl parathion and parathion were assayed in liver homogenates of male and female rats of the five ages. Rates of enzymatic detoxification of their corresponding oxygen analogs by A-esterase, glutathione-S-aryl-, and -S-alkyl-transferase and inactivation by binding were also investigated. Correlation coefficients for rates of metabolism versus LD50 values for the different ages were calculated. In general, changes in LD50 values with age for methyl parathion and parathion correlated better with changes in rates of reactions which represented detoxification pathways for methyl paraoxon and paraoxon than for reactions which represented direct metabolism of the parent insecticides. Both male and female rats became much less sensitive to the acute lethal effects of methyl paraoxon and paraoxon with increasing age. This is consistent with a hypothesis that changes in LD50 values of methyl parathion and parathion with age are due to changes in rates of metabolism of the oxygen analogs.     

Studies of the amplification of carbaryl toxicity by various oximes.

The administration of 2-pyridine aldoxime methyl chloride (2-PAM Cl) is a standard part of the regimen for treatment of human overexposure to many organophosphorus pesticides and nerve agents. However, some literature references indicate that poisoning by carbaryl (1-naphthyl N-methyl carbamate), an insecticide in everyday use, is aggravated by the administration of 2-PAM Cl. This effect has been reported in the mouse, rat, dog and man. We have found that the inhibition of both eel acetylcholinesterase (eel AChE, EC 3.1.1.7) and human serum cholinesterase (human BuChE, EC 3.1.1.8) by carbaryl was enhanced by several oximes. Based on 95% confidence limits the rank order of potentiation with eel AChE was TMB-4 = Toxogonin > HS-6 = HI-6 > 2-PAM Cl. By the same criterion, the rank order of potentiation with human BuChE was TMB-4 > Toxogonin > HS-6 = 2-PAM Cl. Carbaryl-challenged mice also reflected a potentiation since TMB-4 exacerbated the toxicity more than 2-PAM Cl. Our hypothesis is that certain oximes act as allosteric effectors of cholinesterases in carbaryl poisoning, resulting in enhanced inhibition rates and potentiation of carbaryl toxicity.     

Subchronic neurotoxicity of chlorpyrifos,carbaryl, and their combination in rats

Anticholinesterase pesticides have been widely used in agricultural and domestic settings and can be detected in the environment after long‐term use. Although the acute toxic effects of chlorpyrifos and carbaryl have been well described, little is known about the chronic toxicity of the pesticides mixture. To investigate their chronic neurotoxicity, Wistar rats were exposed to chlorpyrifos, carbaryl, and their mixture (MIX) for 90 consecutive days. The activities of serum cholinesterase (ChE) as well as acetylcholinesterase (AChE) and neuropathy target esterase (NTE) in nerve tissues were determined. Furthermore, the histopathological examination was carried out. The results showed that ChE activity significantly decreased in all treated rats except the rats treated with low dose carbaryl. Treatment with middle‐ and high‐dose chlorpyrifos and MIX in rats significantly inhibited AChE activity in the central nervous tissues, whereas treatment with carbaryl alone did not. In sciatic nerve, AChE activity was significantly inhibited by high‐dose carbaryl and MIX, but not by chlorpyrifos alone. No significant NTE inhibition was observed in all treatment groups. Histopathological examination revealed that both chlorpyrifos and MIX treatment induced hippocampal damage. However, no obvious hippocampal damage was found in carbaryl‐treated rats. Carbaryl and MIX, but not chlorpyrifos alone, induced pathological damage of sciatic nerve. Taken together, all of the results indicated that chlorpyrifos and carbaryl have different toxicological target tissues in nervous system and showed corresponding effects in the nervous tissues, which may reflect the different sensitivity of central and peripheral nervous tissues to different pesticides individually and in combination. © 2013 Wiley Periodicals, Inc. Environ Toxicol 29: 1193–1200, 2014.     

Esterase inhibition and reactivation in relation to piperonyl butoxide-phosphorothionate interactions

The effect of piperonyl butoxide on the acute toxicity of phosphorothionate insecticides was studied in male mice. One hour after piperonyl butoxide (400 mg/kg), the toxicity of the dimethyl phosphorothionates, methyl parathion and Guthion, was antagonized, whereas the toxicity of their respective diethyl homologs, parathion and Ethyl Guthion, was potentiated. Piperonyl butoxide did not appreciably alter the toxicity of the oxygen analogs of these compounds. Pretreatment with SKF 525-A (50 mg/kg) modified the toxicity of the phosphorothionates in a manner qualitatively similar to piperonyl butoxide pretreatment. Plasma concentrations of all four insecticides were increased three- to sevenfold in piperonyl butoxide-pretreated mice. This increase may result in a greater total oxon formation; however, reactivation in vitro of esterases inhibited in vivo was 5 to 10 times more rapid following methyl parathion or Guthion challenge than after their diethyl homologs. Although a greater total oxon formation-cholinesterase inhibition is possible for both dimethyl and diethyl phosphorothionates following piperonyl butoxide pretreatment, rapid reactivation of inhibited nerve tissue cholinesterases after dimethyl phosphorothionate challenge appears to compensate for further inhibition occurring at a decreased rate. The net result would be a reduction in dimethyl phosphorothionate toxicity. In contrast, slow reactivation of inhibited nerve tissue cholinesterases following diethyl phosphorothionate challenge appears unable to compensate for increased oxon formation-cholinesterase inhibition. The net result is a potentiation of the toxicity of the diethyl-substituted compounds.