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Bis-propargylcalix[4]arene-stabilized silver nanoparticles (BPCA-Ag NPs), as a chemosensor for detecting an isocarbophos (ICP) pesticide in an aqueous medium, are reported in this work. The nanoparticles were characterized by UV-visible spectroscopy, dynamic light scattering, zeta potential and high-resolution transmission electron microscopy techniques. It was observed that the BPCA-Ag NPs had a high selectivity for isocarbophos with a detection limit of 1.0 × 10 -6  M. According to the result of this research, the BPCA-Ag NPs were found to be useful for the colorimetric detection of isocarbophos in an aqueous medium. It provides a new method for in situ detection of isocarbophos using host-guest interaction., (© 2022. The Author(s), under exclusive licence to The Japan Society for Analytical Chemistry.)

We have previously reported that the long-term exposure of Isocarbophos, a kind of organophosphorus compounds, induces vascular dementia (VD) in rats. Studies have also shown that organophosphorus compounds have adverse effects on offsprings. Vitamin B6 is a coenzyme mainly involved in the regulation of metabolism and has been demonstrated to ameliorate VD. Sphingosine-1-phosphate (S1P), a biologically active lipid, plays a vital role in the cardiovascular system. However, whether S1P is involved in the therapeutic effects of Vitamin B6 on posterior cerebral artery injury has yet to be further answered. In the present study, we aimed to explore the potential influence of Vitamin B6 on Isocarbophos-induced posterior cerebral artery injury in offspring rats and the role of the S1P receptor in this process. We found that Vitamin B6 significantly improves the vasoconstriction function of the posterior cerebral artery in rats induced by Isocarbophos by the blood gas analysis and endothelium-dependent relaxation function assay. We further demonstrated that Vitamin B6 alleviates the Isocarbophos-induced elevation of ICAM-1, VCAM-1, IL-1, and IL-6 by using the enzyme-linked immunosorbent assay kits. By performing immunofluorescence and the western blot assay, we revealed that Vitamin B6 prevents the down-regulation of S1P in posterior cerebral artery injury. It is worth noting that Fingolimod, the S1P inhibitor, significantly inhibits the Vitamin B6-induced up-regulation of S1P in posterior cerebral artery injury. Collectively, our data indicate that Vitamin B6 may be a novel drug for the treatment of posterior cerebral artery injury and that S1P may be a drug target for its treatment., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Locust plagues are still worldwide problems. Selecting active enantiomers from current chiral insecticides is necessary for controlling locusts and mitigating the pesticide pollution in agricultural lands. Herein, two enantiomers of isocarbophos (ICP) were separated and the enantioselectivity in insecticidal activity against the pest Locusta migratoria manilensis (L. migratoria) and mechanisms were investigated. The significant difference of LD 50 between (+)-ICP (0.609 mg/kg bw) and (-)-ICP (79.412 mg/kg bw) demonstrated that (+)-ICP was a more effective enantiomer. The enantioselectivity in insecticidal activity of ICP enantiomers could be attributed to the selective affinity to acetylcholinesterase (AChE). Results of in vivo and in vitro assays suggested that AChE was more sensitive to (+)-ICP. In addition, molecular docking showed that the -CDOKER energies of (+)-ICP and (-)-ICP were 25.6652 and 24.4169, respectively, which suggested a stronger affinity between (+)-ICP and AChE. Significant selectivity also occurred in detoxifying enzymes activities (carboxylesterases (CarEs) and glutathione S-transferases (GSTs)) and related gene expressions. Suppression of detoxifying enzymes activities with (+)-ICP treatment suggested that (-)-ICP may induce the detoxifying enzyme-mediated ICP resistance. A more comprehensive understanding of the enantioselectivity of ICP is necessary for improving regulation and risk assessment of ICP., (Copyright © 2021. Published by Elsevier Inc.)

A novel colorimetric sensing platform based on the peroxidase activity of hemin regulated by oligonucleotide and pesticide was reported for the ultrasensitive and selective detection of isocarbophos. Oligonucleotides can accumulate on the surface of hemin in acid condition and temporarily inhibit its catalytic activity, which results in the loss of one electron of TMB molecule and produce the blue products. With the addition of isocarbophos, the pesticide molecules can interact with oligonucleotides to form some complexes, which relieve the inhibition of ssDNA to hemin and further enhance its catalytic activity. Thus, the TMB molecules are further oxidized to lose another electron and produce the yellow product in a few minutes, which has the characteristic absorption peak at 450 nm. The color change of the sensing system is related to the amount of isocarbophos, so this method can quickly discriminate whether the target pesticide exceeds the maximal residue limit just by naked eyes. To improve the performance of sensing platform, some important parameters like buffer condition and ssDNA have been investigated, and the peroxidase activity of hemin was further studied to verify the catalytic mechanism. The proposed sensing platform has a detection limit as low as 0.6 μg/L and displays good selectivity against other competitive pesticides. Moreover, the developed sensing platform also exhibits favorable accuracy and stability, indicating that it has potential applications in the detection of pesticide residues in agricultural products. Graphical abstract A novel colorimetric sensing platform based on oligonucleotides and pesticide regulation; the peroxidase catalytic activity of hemin was firstly reported for the ultrasensitive and selective detection of isocarbophos pesticide.

Isofenphos-methyl (IFP) is a very active and persistent chiral insecticide. However, IFP has lower activity against acetylcholinesterases (AChEs). Previously, it was confirmed that phosphorothioate organophosphorus pesticides with N-alkyl (POPN) require activation by oxidative desulfuration and N-dealkylation. In this work, we demonstrated that IFP could be metabolized in human liver microsomes to isofenphos-methyl oxon (IFPO, 52.7%), isocarbophos (ICP, 14.2%) and isocarbophos oxon (ICPO, 11.2%). It was found that (R)-IFP was preferentially degraded compared to the (S)-enantiomer, and the enantiomeric fraction (EF) value reached 0.61 at 60 min. However, (S)-enantiomers of the three metabolites, were degraded preferentially, and the EF values ranged from 0.34 to 0.45. Cytochrome P450 (CYP) isoforms CYP3A4, CYP2E1, and CYP1A2 and carboxylesterase enzyme have an essential role in the enantioselective metabolism of IFP; but, the enzymes that participate in the degradation of IFP metabolites are different. The AChE inhibition bioassay indicated that ICPO is the only effective inhibitor of AChE. The covalent molecular docking has proposed that the metabolites of IFP and its analogs after N-dealkylation and oxidative desulfuration will possess the highest inhibitory activity against AChE. This study is the first to demonstrate that ICPO can be regarded as a potential biomarker for the biomonitoring of IFP and ICP exposure in humans., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Although isocarbophos is a widely used insecticide, its toxicity to aquatic organisms has not been well characterized. In this study, zebrafish were exposed to isocarbophos at concentrations of 50 µg L -1 and 200 µg L -1 to assess its bioaccumulation, metabolic disruption, and oxidative stress. Metabolomics analysis based on 1 H NMR spectroscopy showed that 50 µg L -1 and 200 µg L -1 isocarbophos exposure induced increases in leucine, isoleucine, valine, and alanine compared to levels in the control. Lactate, creatine, and taurine were reduced in the 50 µg L -1 isocarbophos exposure group, and only lactate decreased in response to 200 µg L -1 isocarbophos. After zebrafish were exposed to 50 and 200 µg L -1 isocarbophos for 28 days, the activities of antioxidant enzymes (SOD, CAT, and GPx) and GSH contents decreased significantly in the liver. This result indicates that there was significant oxidative stress in the liver. Furthermore, changes in metabolite profiles significantly covaried with changes in several oxidative stress endpoints based on partial least squares regression. These results will contribute to the environmental risk assessment of isocarbophos and clarify the mechanism underlying its toxicity in zebrafish., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Vascular dementia, being the most severe form of vascular cognitive impairment (VCI), is caused by cerebrovascular disease. Whether organophosphorus causes VCI remains unknown. Isocarbophos (0.5 mg/kg per 2 days) was intragastrically administrated to rats for 16 weeks. The structure and function of cerebral arteries were assayed. The learning and memory were evaluated by serial tests of step-down, step-through and morris water maze. Long-term administration of isocarbophos reduced the hippocampal acetylcholinesterase (AChE) activity and acetylcholine (ACh) content but did not alter the plasma AChE activity, and significantly damaged the functions of learning and memory. Moreover, isocarbophos remarkably induced endothelial dysfunction in the middle cerebral artery and the expressions of ICAM-1 and VCAM-1 in the posterior cerebral artery. Morphological analysis by light microscopy and electron microscopy indicated disruptions of the hippocampus and vascular wall in the cerebral arteries from isocarbophos-treated rats. Treatment of isocarbophos injured primary neuronal and astroglial cells isolated from rats. Correlation analysis demonstrated that there was a high correlation between vascular function of cerebral artery and hippocampal AChE activity or ACh content in rats. In conclusion, chronic administration of isocarbophos induces impairments of memory and learning, which is possibly related to cerebral vascular dysfunction., (© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

The benthic fauna is of great importance to assess the environmental fate of contaminations in aquatic ecosystem. In this study, tubificids were exposed to both laboratory-contaminated aqueous phases and spiked sediment to study the bioaccumulation of isocarbophos (ICP). Two types of spiked sediments were used in the spiked sediment experiment. During the exposure period, an enantioselective bioaccumulation was found in spiked water treatment, with concentrations of the (-)-ICP higher than that of the (+)-ICP, but no enantioselectivity was detected in the spiked sediment treatments. However, different bioaccumulation patterns were observed in the two spiked sediment treatments. Results showed that for spiked forest field sediment (FF sediment) incubation, bioaccumulation was governed by the concentrations in soil. Whereas ICP was bioaccumulated dominantly from overlying water in spiked Chagan Lake sediment (CG sediment) test. The dissipation rates were proved different in the two sediments and ICP dissipated much faster in CG sediment than that in FF sediment. Significant difference in ICP's half-life was also observed between worm-present and worm-free treatments in FF sediment. The detections of concentrations in overlying water indicated that much more ICP diffused to aquatic phase with the present of tubificids., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

An axenic bacterial strain, Lysinibacillus sp. KB1, was isolated from malathion-contaminated soil. It tolerated malathion up to 0.15 % and, under aerobic conditions, utilized it as sole carbon source. 20 % malathion and 47 % malaoxon were degraded out of the initially provided malathion. Two metabolites, mal-monocarboxylic acid and mal-dicarboxylic acid, were detected within 7 days at 30 °C. Esterase activity of the strain was 240 ± 2.5 U/ml after 7 days of growth. Sterilized soil mixed with malathion showed rapid degradation of malathion when inoculated with strain KB1 as compared to the uninoculated soil.

This work presents the development of bioassays and biosensors for the detection of insecticides widely used in the treatment of olive trees. The systems are based on the covalent immobilisation of acetylcholinesterase on magnetic microbeads using either colorimetry or amperometry as detection technique. The magnetic beads were immobilised on screen-printed electrodes or microtitration plates and tested using standard solutions and real samples. The developed devices showed good analytical performances with limits of detection much lower than the maximum residue limit tolerated by international regulations, as well as a good reproducibility and stability.

Acetylcholinesterase (AChE), encoded by the Ace gene, is the primary target of organophosphates (OPs) and carbamates (CBs) in insects. Ace mutations have been identified in OP and CB resistant strains of Musca domestica. In this study, the Ace gene was partially amplified and sequenced at amino acid positions 260, 342, and 407 to determine the frequencies of these mutations in housefly samples collected from farms and garbage disposal sites of 16 provinces in the Aegean and Mediterranean regions of Turkey. In addition, the percent remaining AChE activities in these samples were assayed by using three OPs (malaoxon, paraoxon, and dichlorvos) and one CB (carbaryl) compound as inhibitors. In all the analyzed samples, 13 different combinations at the three amino acid positions were identified and the L/V260-A/G342-F/Y407 combination was found in the highest frequency. No susceptible individual was detected. The highest mean percent remaining AChE activities were detected in the individuals having the L260-A/G342-F/Y407 genotype when malaoxon and paraoxon were used as inhibitors and in the individuals with the L260-A342-F/Y407 combination when dichlorvos and carbaryl were used as inhibitors. The obtained data were heterogeneous and there was no exact correlation between the molecular genetic background and the resistance phenotypes of the flies. The findings of this study at the molecular and biochemical levels indicate the presence of significant control problems in the field., (© 2011 The Society for Vector Ecology.)

The simultaneous use of the repellent DEET, pyridostigmine, and organophosphorus pesticides has been assumed as a potential cause for the Gulf War Illness and combinations have been tested in different animal models. However, human in vitro data on interactions of DEET with other compounds are scarce and provoked the present in vitro study scrutinizing the interactions of DEET, pyridostigmine and pesticides with human acetylcholinesterase (hAChE) and butyrylcholinesterase (hBChE). DEET showed to be a weak and reversible inhibitor of hAChE and hBChE. The IC(50) of DEET was calculated to be 21.7mM DEET for hAChE and 3.2mM DEET for hBChE. The determination of the inhibition kinetics of pyridostigmine, malaoxon and chlorpyrifos oxon with hAChE in the presence of 5mM DEET resulted in a moderate reduction of the inhibition rate constant k(i). The decarbamoylation velocity of pyridostigmine-inhibited hAChE was not affected by DEET. In conclusion, the in vitro investigation of interactions between human cholinesterases, DEET, pyridostigmine, malaoxon and chlorpyrifos oxon showed a weak inhibition of hAChE and hBChE by DEET. The inhibitory potency of the tested cholinesterase inhibitors was not enhanced by DEET and it did not affect the regeneration velocity of pyridostigmine-inhibited AChE. Hence, this in vitro study does not give any evidence of a synergistic effect of the tested compounds on human cholinesterases., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Soybean fields provide habitats for many species of amphibians. However, the persistence and health of amphibian populations may be at risk from the increasing use of pesticides and other agricultural chemicals. We examined the activities of acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and carboxylesterases (CbEs) in 11 syntopic species of larval anurans. In vitro effects of malaoxon causing 50% BChE inhibition (IC(50)) were also studied. In addition, we calculated a relative risk index (RI) based on the geographic distributions of the anurans, the phenology of soybean cultivation, and basal enzymatic values related to potential pesticide detoxification. Among the 11 species, AChE activity varied from 17.5 ± 1.6 to 68.2 ± 4.7 nmol min(-1) mg(-1) protein (PT). BChE activity also varied significantly, ranging from 3.3 ± 0.4 to 7.5 ± 0.4 nmol min(-1) mg(-1)PT. Both measures of CbE activities varied widely (CbE α-NA: 2.1 ± 0.5-12.4 ± 1.1 nmol min(-1) mg(-1) PT; CbE-4NPV: 21.8 ± 1.8-102.6 ± 7.9 nmol min(-1) mg(-1) PT). We also corroborate that lower BChE activity levels for the tadpoles were associated at minor IC(50) values. The results of this study demonstrate significant variation in enzymatic levels among several tadpole species and intermediate to high RI values for 7 species. Based on these results, it appears that a conversion of native ecosystems to soybean crops may lead to increased ecological risk for anuran amphibians., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Isocarbophos is a widely used organophosphorus insecticide that has caused environmental pollution in many areas. However, degradation of isocarbophos by pure cultures has not been extensively studied, and the degradation pathway has not been determined. In this paper, a highly effective isocarbophos-degrading strain, scl-2, was isolated from isocarbophos-polluted soil. Strain scl-2 was preliminarily identified as Arthrobacter sp. based on its morphological, physiological, and biochemical properties, as well as 16S rDNA analysis. Strain scl-2 could utilize isocarbophos as its sole source of carbon and phosphorus for growth. One hundred mg/l isocarbophos could be degraded to a nondetectable level in 18 h by scl-2 in cell culture, and isofenphos-methyl, profenofos, and phosmet could also be degraded. During the degradation of isocarbophos, the metabolites isopropyl salicylate, salicylate, and gentisate were detected and identified based on MS/MS analysis and their retention times in HPLC. Transformation of gentisate to pyruvate and fumarate via maleylpyruvate and fumarylpyruvate was detected by assaying for the activities of gentisate 1,2- dioxygenase (GDO) and maleylpyruvate isomerase. Therefore, we have identified the degradation pathway of isocarbophos in Arthrobacter sp. scl-2 for the first time. This study highlights an important potential use of the strain scl-2 for the cleanup of environmental contamination by isocarbophos and presents a mechanism of isocarbophos metabolism.

Organophosphorus pesticides (OPs) are ubiquitous in the environment and are highly toxic to amphibians. They deactivate cholinesterase, resulting in neurological dysfunction. Most chemicals in this group require oxidative desulfuration to achieve their greatest cholinesterase-inhibiting potencies. Oxon derivatives are formed within liver cells but also by bacterial decay of parental pesticides. This study examines the toxicity of chlorpyrifos, malathion and diazinon and their oxons on the foothill yellow-legged frog (Rana boylii). R. boylii is exposed to agricultural pesticides in the California Central Valley. Median lethal concentrations of the parental forms during a 96 h exposure were 3.00 mg/L (24h) for chlorpyrifos, 2.14 mg/L for malathion and 7.49 mg/L for diazinon. Corresponding oxons were 10 to 100 times more toxic than their parental forms. We conclude that environmental concentrations of these pesticides can be harmful to R. boylii populations.

Four organophosphorus compounds: azinphos-methyl, chlorpyrifos, malathion and malaoxon in aqueous solution were degraded by using a 125 W xenon parabolic lamp. Gas chromatography-mass spectrometry (GC-MS) was used to monitor the disappearance of starting compounds and formation of degradation products as a function of time. AChE-thermal lens spectrometric bioassay was employed to assess the toxicity of photoproducts. The photodegradation kinetics can be described by a first-order degradation curve C=C0e(-kt), resulting in the following half lives: 2.5min for azinphos-methyl, 11.6 min for malathion, 13.3 min for chlorpyrifos and 45.5 min for malaoxon, under given experimental conditions. During the photoprocess several intermediates were identified by GC-MS suggesting the pathway of OP degradation. The oxidation of chlorpyrifos results in the formation of chlorpyrifos-oxon as the main identified photoproduct. In case of malathion and azinphos-methyl the corresponding oxon analogues were not detected. The formation of diethyl (dimethoxy-phosphoryl) succinate in traces was observed during photodegradation of malaoxon and malathion. Several other photoproducts including trimethyl phosphate esters, which are known to be AChE inhibitors and 1,2,3-benzotriazin-4(3H)-one as a member of triazine compounds were identified in photodegraded samples of malathion, malaoxon, and azinphos-methyl. Based on this, two main degradation pathways can be proposed, both result of the (P-S-C) bond cleavage taking place at the side of leaving group. The enhanced inhibition of AChE observed with the TLS bioassay during the initial 30 min of photodegradation in case of all four OPs, confirmed the formation of toxic intermediates. With the continuation of irradiation, the AChE inhibition decreased, indicating that the formed toxic compounds were further degraded to AChE non-inhibiting products. The presented results demonstrate the importance of toxicity monitoring during the degradation of OPs in processes of waste water remediation, before releasing it into the environment.

Engineered variants of Drosophila melanogaster acetylcholinesterase (AChE) were used as biological receptors of AChE-multisensors for the simultaneous detection and discrimination of binary mixtures of cholinesterase-inhibiting insecticides. The system was based on a combination of amperometric multielectrode biosensors with chemometric data analysis of sensor outputs using artificial neural networks (ANN). The multisensors were fully manufactured by screen-printing, including enzyme immobilisation. Two types of multisensors were produced that consisted of four AChE variants each. The AChE mutants were selected in order to obtain high resolution, enhanced sensitivity and minimal assay time. This task was successfully achieved using multisensor I equipped with wild-type Drosophila AChE and mutants Y408F, F368L, and F368H. Each of the AChE variants was selected on the basis of displaying an individual sensitivity pattern towards the target analytes. For multisensor II, the inclusion of F368W, which had an extremely diminished paraoxon sensitivity, increased the sensor's capacity even further. Multisensors I and II were both used for inhibition analysis of binary paraoxon and carbofuran mixtures in a concentration range 0-5 microg/l, followed by data analysis using feed-forward ANN. The two analytes were determined with prediction errors of 0.4 microg/l for paraoxon and 0.5 microg/l for carbofuran. A complete biosensor assay and subsequent ANN evaluation was completed within 40 min. In addition, multisensor II was also investigated for analyte discrimination in real water samples. Finally, the properties of the multisensors were confirmed by simultaneous detection of binary organophosphate mixtures. Malaoxon and paraoxon in composite solutions of 0-5 microg/l were discriminated with predication errors of 0.9 and 1.6 microg/l, respectively.

Extensive utilization of pesticides against insects provides us with a good model for studying the adaptation of a eukaryotic genome to a strong selective pressure. One mechanism of resistance is the alteration of acetylcholinesterase (EC 3.1.1.7), the molecular target for organophosphates and carbamates. Here, we report the sequence analysis of the Ace gene in several resistant field strains of Drosophila melanogaster. This analysis resulted in the identification of five point mutations associated with reduced sensitivities to insecticides. In some cases, several of these mutations were found to be combined in the same protein, leading to different resistance patterns. Our results suggest that recombination between resistant alleles preexisting in natural populations is a mechanism by which insects rapidly adapt to new selective pressures.

The state of California conducted aerial applications of malathion (MA) bait over urban areas in the southern California air basin in order to eradicate the Mediterranean Fruit Fly (Ceratitis capitata). Concern about the potential human health effects of this activity prompted a risk assessment conducted by the California Department of Health Services. Estimates of potential human exposures to MA and its primary breakdown product, malaoxon, (MO) are based on assumptions of daily human activities which influence the rate of contact with MA and MO. Several exposure scenarios, representing a range of activity levels from sedentary to very active, are used as surrogates for a variety of human activities. For each exposure scenario, acute dose rates are calculated using both the mean and the mean plus 2 standard deviations (SD) (98% upper confidence limit (UCL)) of measured environmental values. Chronic dose rates are calculated using long-term averages incorporating degradation and multiple applications at 14 day intervals, and the estimated 98% UCL for these averages. Based on this model, estimated adult dermal doses (1-246 micrograms/kg-d) are up to about 2000-fold higher than the estimated inhalation doses (0.01-0.1 microgram/kg-d) but are comparable to the doses from ingestion of contaminated unwashed backyard vegetables (30-80 micrograms/kg-d). For the individual who does not consume backyard vegetables, therefore, almost the whole dose of MA or MO would be due to contacting contaminated surfaces with skin.

Between August 1989 and July 1990, California conducted a Mediterranean fruit fly eradication project in southern California which included repeated aerial applications of malathion bait to urban areas where approximately 1.6 million people resided. Concern about the safety of these applications prompted the California Department of Health Services to prepare a risk assessment. The current work presents the estimates of environmental levels of malathion and malaoxon, derived from mass deposition rates during application and monitoring of air. We estimated short and long-term malathion and malaoxon levels on outdoor surfaces, plants, and soils (0.1 and 1.0 cm mixing depth), using a simple first-order exponential decay model and literature half-life values ranging from three to nine days. Direct monitoring data were used to characterize short-term air levels. Average and upper-bound malathion levels immediately following an application were 0.091 microgram/m3 and 0.207 microgram/m3 in air; 22 and 52 mg/m2 on outdoor surfaces; 3.8 and 9.6 micrograms/g in plants; and 1.5 and 3.5 micrograms/g in soil (1 cm mixing depth). Malaoxon levels were 0.039 microgram/m3 and 0.110 microgram/m3 in air; 0.15 and 0.46 mg/m2, 0.03 and 0.09 microgram/g, and 0.01 and 0.03 microgram/g, respectively. Estimates of average and upper-bound levels over the duration of the eradication program were roughly one-third the immediate levels in all media. No field measurements were available for co-products other than malaoxon, for environmental persistence, or for concentrations on surfaces, soil, or plants after repeated applications. Because of limited monitoring data, and the likelihood that similar pest-eradication programs will be conducted, additional studies are warranted to more accurately characterize the environmental fate of malathion and human exposures after aerial application to residential communities.

Acetylcholinesterase from the heads of insecticide-resistant and -susceptible houseflies (Musca domestica L.) was studied in vitro. The enzymes could not be distinguished electrophoretically, and their behaviour on polyacrylamide-disc-gel electrophoresis was influenced by the presence of Triton X-100 in both the homogenate and the gels. In the absence of detergent, the acetylcholinesterase was heterogeneous, but behaved as a single enzyme when it was present. By analogy with studies of acetylcholinesterase from other sources, these observations were attributed to aggregation of the enzyme when not bound by membranes. The enzyme from resistant flies was more slowly inhibited than the susceptible enzyme, bimolecular rate constants (ki) differing by approx. 4-20-fold for a range of organophosphorus compounds. The kinetics of inhibition of acetylcholinesterase were consistent with the results of electrophoresis, i.e. they corresponded to those of a single enzyme in the presence of Triton X-100, but a mixture of enzymes in its absence. The susceptibility of the more sensitive components in these mixtures was determined.

Malathion and its oxygen analogue, malaoxon, are carcinogenic in Osborne-Mendel and Fischer-344 rats. Benign and malignant neoplasms at all sites were increased in Osborne-Mendel and Fischer-344 male and female rats ingesting malathion. Both Osborne-Mendel and Fischer-344 male rats were more susceptible than the female rats. The organ sites were not the same for Fischer-344 and Osborne-Mendel rats. Osborne-Mendel rats developed neoplasms of the endocrine organs, brain, and liver. Fischer-344 rats had neoplasms of the adrenal medulla, organs with squamous cells, lung, and hematopoietic system. Fischer-344 male rats given malathion were more susceptible to chronic renal disease, parathyroid hyperplasia, metastatic calcification, and atrophy of the testes than were Osborne-Mendel male rats. They also had ulcers of the forestomach. Chronic renal disease and atrophy of the testes affected the health of the animals and interfered with the development of neoplasms. Malathion increased the incidence of neoplasms of the liver in B6C3F1 male mice. Male mice also had atrophy of the testes. Benign and malignant neoplasms at all sites, and in the endocrine organs, were increased in Fischer-344 male and female rats ingesting malaoxon. Neoplasms were present in the adrenal medulla, organs with squamous cells, liver, and hematopoietic system. Both male and female Fischer-344 rats receiving malaoxon had chronic renal disease and the male rats had parathyroid hyperplasia and metastatic calcification.

In the early 1970s the National Cancer Institute (NCI) studied malathion and the oxygen analog, malaoxon, for possible carcinogenicity in rats and mice. The results from these long-term studies were reported in three NCI Technical Reports with the conclusions that neither chemical was shown to be carcinogenic in rodents. In response to the renewed public health interest and concern about the increasing use of malathion in agriculture and especially its use to eliminate Mediterranean fruit fly infestations in California and Florida during the 1980s, the National Toxicology Program (NTP) in consultation and agreement with the NCI reevaluated the histopathology of the NCI studies of malathion in Osborne-Mendel and Fischer 344 rats and of malaoxon in Fischer 344 rats. The NTP histopathology reexamination confirmed the original NCI interpretative conclusions that malathion was not carcinogenic. For the malaoxon study, the only difference between the original and subsequent interpretations was for C-cell neoplasms of the thyroid gland, in that the NTP concluded there was equivocal evidence of carcinogenicity for male and female F344 rats.