Stereoisomeric separation and toxicity of a new organophosphorus insecticide chloramidophos

Chloramidophos (CP), O,S-dimethyl-[(2,2,2)-trichloro-1-hydroxyethyl]phosphoramidothioate, is a new organophosphorus pesticide (OP) with two chiral centers each on the phosphorus and carbon atoms. Although CP has been widely used in some provinces of China, it has received very limited attention toward its environmental behaviors, in particular, with regard to stereospecificity. In this study, the stereoisomeric separation and toxicity of CP were investigated. All of the four stereoisomers of CP were successfully separated by high-performance liquid chromatography on a Chiralpak AD column. The stereoisomers (pk 1 to pk 4) were distinguishable on their mass and circular dichroism spectra. The inhibition on acetylcholinesterases (AChE, in vitro) and the acute aquatic toxicity to Daphnia magna (in vivo) tested with optically pure stereoisomers of CP showed its stereoselectivity. The inhibitory potency toward AChE decreased in the order of pk 4 > pk 3 > pk 2 > pk 1. In comparison, the acute toxicity to D. magna was in the order of pk 3 > pk 2 > pk 1 > pk 4. The stereoselectivity was found to be isomer-dependent, with 1.1-18.1-fold differences (in vitro) and 1.2-13-fold differences (in vivo) among the stereoisomers. These results suggest that the overall toxicity of chiral OPs should be assessed using their individual enantiomers.     

Developmental toxicity of flupyrazofos, a new organophosphorus insecticide, in rats.

Flupyrazofos is a new type of pyrazole organophosporus insecticide, which has a high activity against the diamond-back moth (Plutella xylostella). The potential of this agent to induce developmental toxicity was investigated in the Sprague-Dawley rat. One hundred mated females (sperm in vaginal LAVAGE=day 0) were distributed among three treated groups and a control group. Flupyrazofos was administered by gavage to pregnant rats from days 7-17 of gestation at dose levels of 0, 5, 12 and 30 mg/kg/day. All dams were subjected to the caesarean section on day 20 of gestation and their fetuses were examined for external, visceral and skeletal abnormalities. At 30 mg/kg, maternal effects including mortality (4.3%), clinical signs of toxicity, decreased food intake, suppressed body weight, and increased weight of adrenal glands, kidney and heart were observed in dams. Litter values for corpora lutea, implantations, sex ratio and litter size were within the normal range. However, a reduction in the fetal weight and an increase in the incidence of fetal skeletal retardations were observed. At 12 mg/kg, toxic effects including mortality (4.2%), nasal discharge and some fetal skeletal retardation were observed. There were no signs of either maternal toxicity or embryotoxicity at 5 mg/kg. The results show that flupyrazofos induces fetal growth retardation only at maternally toxic doses in rats and the no-observed-adverse-effect levels (NOAELs) of this agent are considered to be 5 mg/kg for both dams and fetuses.     

Passive avoidance retention in mice tolerant to the organophosphorus insecticide disulfoton

Male mice were administered the organophosphorus insecticide disulfoton (O,O-diethyl S-[2-(ethylthio)ethyl] phosphorodithioate) daily for 14 days at a dosage rate of 10 mg/kg/day. Signs of poisoning disappeared after 5 to 8 days of treatment indicating that the animals had developed tolerance to disulfoton toxicity. Acetylcholinesterase activity was inhibited 75 to 93% in hippocampus, cerebral cortex, striatum, and cerebellum of tolerant animals. Binding of the muscarinic cholinergic antagonist [³H]quinuclidinyl benzilate was significantly decreased in all these brain areas. The decrease was due to a decrease in the number of receptors (B max) without any change in the affinity constant K_d. Sodium-dependent high-affinity choline uptake was unchanged in the hippocampus of tolerant animals. When tested in a single trial passive avoidance task, mice tolerant to disulfoton did not show any difference from controls in 24-hr retention. These results indicate that the decreased number of muscarinic receptors in brain of disulfoton-tolerant animals does not affect their response latencies in a passive avoidance task.     

Dose-dependent toxicity of diphenhydramine overdose

BACKGROUND: Diphenhydramine (DPHM) overdose is a frequent cause of acute poisoning. Although its clinical features are well known, information about the dose-dependent toxicity of DPHM is still scarce. The objective of this study was to investigate the dose-dependent toxicity of DPHM in patients with acute DPHM poisoning. METHODS: We have analyzed retrospectively all well-documented cases with DPHM monointoxications reported by physicians to the Swiss Toxicological Information Centre (STIC) between January 1984 and April 1996. In addition, a prospective study focusing on ingested DPHM doses and severity of symptoms was performed between May 1996 and December 1998. RESULTS: The retrospective and prospective studies included 232 and 50 patients with DPHM monointoxications, respectively. In both studies, mild symptoms (somnolence, anticholinergic signs, tachycardia, nausea/vomiting) occurred in 55-64%, moderate symptoms (isolated and spontaneously resolving agitation, confusion, hallucinations and ECG disturbances) in 22-27% and severe symptoms (delirium/psychosis, seizures, coma) in 14-18% of patients. Moderate symptoms occurred above ingested doses of 0.3 g DPHM. For severe symptoms the critical dose limit was 1.0 g DPHM. Although the frequency of delirium/psychosis remained constant or even decreased, coma and seizures were significantly (p<0.05) more frequent in the >1.5-g compared with the 1.0- to 1.5-g-dose group. CONCLUSIONS: These data demonstrate a clear dose-dependent acute toxicity of DPHM. They indicate that only patients with DPHM ingestions above 1.0 g are at risk for the development of severe symptoms and, therefore, should be hospitalized. Thus, the results contribute to the data basis required for a cost effective management of patients with DPHM overdose.     

群体性有机磷杀虫剂中毒救治探讨

目的探讨群体性有机磷杀虫剂（OPI）中毒救治的临床特点。方法回顾性分析一次突发群体性OPI中毒的临床资料。结果 55例OPI中毒患者为集体生活、统一进食,在相同时间段出现头昏、恶心、呕吐、上腹部疼痛,其中,32例出现皮肤潮湿多汗,1例出现心慌、心悸和燥动不安,1例出现幻觉、躁狂、谵妄、尿潴留,大多数患者、家属、老师出现不同程度的紧张恐慌情绪;40例胆碱酯酶（ChE）活力降低,15例ChE活力正常;55例患者经排毒、解毒、对症、支持治疗均痊愈。结论群体性OPI中毒救治过程中要区别对待不同患者,注意心因性反应影响病情判断,合理使用解毒药物,防治阿托品中毒发生。     

Extremely sensitive biomarker of acute organophosphorus insecticide exposure

Egasyn-beta-glucuronidase complex is located at the luminal site of liver microsomal endoplasmic reticulum. When organophosphorus insecticides (OP) are incorporated into the liver microsomes, they become tightly bound to egasyn, a carboxylesterase isozyme, and subsequently, beta-glucuronidase (BG) is dissociated and released into blood. Consequently, the increase in plasma BG activity becomes a good biomarker of OP exposure. Thus, the single administration of EPN (O-ethyl O-p-nitrophenylphenylphosphonothioate), acephate and chlorpyrifos increased plasma BG activity in approximately 100-fold the control level in rats. The increase in plasma BG activity after OP exposure is a much more sensitive biomarker of acute OP exposure than acetylcholinesterase (AChE) inhibition.     

Evaluation of oxidative stress and genotoxicity in organophosphorus insecticide formulators

The aim of this study was to evaluate genotoxicity and oxidative stress in workers who formulate organophosphorus (OP) pesticides. In this survey, blood leukocytes and erythrocytes of a group of 21 pesticide formulating workers and an equal number of control subjects were examined for genotoxicity and oxidative stress parameters. The mean comet tail length and mean comet length were used to measure DNA damage. Lipid peroxidation level, catalase, superoxide dismutase (SOD) and glutathione peroxidase activities in erythrocytes were analysed as biomarkers of oxidative stress. In addition, the acetylcholinesterase activity was measured as a biomarker of toxicity. The average duration of employment of workers in the factory was 97 months. Results indicated that chronic exposure (multiple-dose, greater than or equal to 6 months duration) to OP pesticides was associated with increased activities of catalase, SOD and glutathione peroxidase in erythrocytes. The level of lipid peroxidation and acetylcholinesterase activity did not show any significant differences between the two groups. The results also indicated that chronic exposure to OP pesticides was associated with increased DNA damage. It is concluded that human chronic exposure to OP pesticides may result in stimulated antioxidant enzymes and increased DNA damage in the absence of depressed acetylcholinesterase levels. Routine genotoxicity monitoring concomitant to acetylcholinesterase activity in workers occupationally exposed to OP insecticides is suggested.     

Effect of preinduction of metallothionein on paraquat toxicity in mice

The effect of pretreatment with metallothionein (MT)-inducing metals (Zn, Cu, Bi, Co, Cd or Hg) on paraquat (PQ) toxicity was investigated in mice. PQ lethality was remarkably reduced by pretreatment with the above MT-inducing metals. The protective effect of pretreatment with these metals on PQ lethality was significantly correlated with MT levels in the lung, a target tissue of PQ toxicity, in mice administered MT-inducing metals, but not with MT in the liver or kidney. The increase in pulmonary lipid peroxidation in mice treated with PQ was significantly inhibited by Zn pretreatment. Zn was the most effective of the MT-inducing metals used in this experiment in protecting mice against PQ lethality. Of those monitored, the only pulmonary free radical scavenging factor increased by Zn pretreatment was MT. Other free radical scavenging factors (activities of Superoxide dismutase, glutathione peroxidase and catalase, and concentration of non-protein thiols level) were not influenced by Zn treatment. These results indicate that the induction of pulmonary MT protects against the lethality and lung toxicity of PQ. Pulmonary MT may scavenge free radicals produced by PQ, thereby protecting against lethal pulmonary toxicity.     

Effect of ascorbate on the toxicity of morphine in mice

The effects of ascorbic acid on the toxicity of morphine in mice were investigated. An intraperitoneal dose of sodium ascorbate (1 G/kg) injected 10 min prior to morphine (500 mg/kg, i.p.) was found to provide significant protection against mortality due to respiratory depression, while having no effect on the lethality of the pentobarbital. Pretreatment with ascorbate had no effect on the distribution of morphine in brain tissue, nor did it alter the pH of the plasma. Administration of ascorbate in vivo also produced no inactivation of binding to opioid receptors. It is postulated that ascorbate antagonizes the lethality of morphine by selectively affecting neuronal activity.     

Effect of L-cystine on toxicity of paraquat in mice.

The protective effect of L-cystine on the toxicity of paraquat (PQ) in mice was studied. Lipid peroxidation in the lung significantly increased after oral administration of PQ (200 mg/kg) and the increase in lipid peroxidation was prevented by L-cystine treatment (300 mg/kg). PQ administration produced an increase in superoxide dismutase (SOD) activity and a decrease in glutathione peroxidase (GSH-Px) activity in the lung at 24 h after PQ. L-Cystine treatment significantly prevented the changes in SOD and GSH-Px activity in the lung after PQ. L-Cystine treatment prevented the decrease in non-protein sulfhydryl (NP-SH) content in the lung after PQ administration. The tissue distribution and excretion of PQ after PQ administration were not changed by L-cystine treatment. Plasma aspartate aminotransferase activity did not change after PQ administration. These results suggest that L-cystine protects against the toxicity of PQ by maintaining reduced glutathione levels in the cells.     

Environmental pyrethroid and organophosphorus insecticide exposures and sperm concentration

BACKGROUND: There is growing concern that poisoning and other adverse health effects are increasing because organophosphorous (OP) insecticides are now being used in combination with pyrethroid (PYR) insecticides to enhance the toxic effects of PYR insecticides on target insects, especially those that have developed PYR resistance. OBJECTIVES: We conducted a pilot biomonitoring study to determine whether men in our reproductive cohort study were being exposed to pesticides environmentally by virtue of frequenting an agricultural setting. METHODS: We screened 18 randomly selected urine samples collected from male participants of reproductive age for 24 parent compounds and metabolites of pesticides and examined the results in relation to sperm concentration. RESULTS: Results showed high prevalence of exposure to OP and PYR pesticides and our preliminary analyses provided some suggestion that the higher exposure group had lower sperm concentration. CONCLUSIONS: The potential of OP/PYR mixtures to have enhanced human toxicity needs more research attention.     

Spinosad insecticide: subchronic and chronic toxicity and lack of carcinogenicity in CD-1 mice.

The potential toxicologic and oncogenic effects of spinosad, a natural fermentation product with insecticidal properties, were investigated. The 13-week toxicity study consisted of groups of 10 CD-1 mice/sex provided diets containing 0, 0.005, 0.015, 0.045, or 0.12% spinosad (Study 1). The 0.12% group was terminated on Test Day 44 due to mortality and overt clinical signs of toxicity. An 18-month chronic oncogenicity study consisted of groups of 50 CD-1 mice/sex provided diets containing 0, 0.0025, 0.008, or 0.036% spinosad (Study 2). Two interim groups of 10 mice/sex/group were terminated after 3 and 12 months. Females given 0.036% were terminated on Day 455 due to markedly lower body weights and feed consumption, as well as excessive mortality. Because of the early termination of the female high-dose group, additional groups of 10 male and female mice (12-month interim necrospy) and 50 male and female mice (18-month necropsy) were provided diets containing 0, 0.0008, or 0.024% spinosad (Study 3) to fully assess potential chronic toxicity and oncogenicity. Standard toxicologic parameters were evaluated consistent with existing regulatory guidelines. The primary effect in the 13-week and 18-month studies was intracellular vacuolation of histiocytic and epithelial cells in numerous tissues and organs at doses of > or = 0.015%. The histological vacuolation corresponded to ultrastructural lysosomal lamellar inclusion bodies. This alteration was consistent with phospholipidosis, a condition that results from accumulation of polar lipids in lysosomes. Lesions with no apparent direct relation to vacuolation were hyperplasia of the glandular mucosa of the stomach, skeletal muscle myopathy, bone marrow necrosis, and anemia with associated splenic hematopoiesis. The incidence of tumors in mice given spinosad was not increased relative to controls at any dose level. The no observed effect level for the 13-week study was 0.005% (6 mg/kg/day) spinosad, and for the chronic toxicity/oncogenicity study was 0.008% (11 mg/kg/day) spinosad for male and female CD-1 mice.     

Effect of diethyldithiocarbamate on the toxicity of amphetamine in aggregated mice

Summary These studies were performed to investigate the influence of diethyldithiocarbamate on the toxicity of amphetamine in aggregated mice. In the same mice changes in body temperature and spontaneous motility were determined. In the other experiments brain catecholamine concentrations were assayed. Diethyldithiocarbamate potentiated the toxicity of amphetamine in some experimental conditions and slightly antagonized it in others. These effects are discussed in relation to behavioral and biochemical changes observed in mice.     

紫杉醇长效微球对小鼠生物相容性和血液毒性研究

目的研究紫杉醇长效微球对小鼠的生物相容性和血液毒性。方法 18只小鼠分别于背部皮下注射20 mg紫杉醇长效微球。于第0、5、10、15、20、25 d分别处死3只小鼠,取皮肤组织观察炎症反应。另取18只小鼠,分为长效微球组、紫杉醇注射液（Taxol）组和对照组（以生理盐水作为对照）3组,每组6只,进行血细胞计数的测定。结果紫杉醇长效微球皮下注射后开始出现轻微的炎症反应,未观察到明显血管与纤维的增生,第15日后已经恢复正常;皮下注射微球后,引起了白细胞计数的轻微下降并随后维持恒定,而紫杉醇注射液在给药后前2周,小鼠的白细胞计数出现显著下降,显示紫杉醇长效微球对小鼠的血细胞损伤性较小。结论紫杉醇长效缓释注射微球具有良好的生物相容性,皮下注射后对血细胞无明显杀伤作用,安全性较高。     

Effect of dietary butylated hydroxyanisole on methylazoxymethanol acetate-induced toxicity in mice

Administration of butylated hydroxyanisole (BHA), a widely used food additive, has been found to inhibit the carcinogenic and toxic effects of various chemicals in animal models. To study the relationship of dietary BHA to the acute toxicity of methylazoxymethanol (MAM) acetate, a colonspecific carcinogenic compound, groups of female CF₁ mice were fed NIN-07 diet containing 0, 300, 1000, 3000 or 6000 ppm BHA or a semipurified diet containing 0 or 6000 ppm BHA for 4 wk, and were injected ip with MAM acetate (20 mg/kg body weight) at the end of the first 2 wk and again 4 days later. At levels of 300–6000 ppm. BHA was found to protect against death caused by MAM acetate. The mortality rates in MAM-treated mice were 80 and 92% in those fed the diets with no BHA and 0 and 1% in those fed 6000 ppm BHA, and were inversely related to the amount of BHA in the diet. The protection was associated with increased levels of hepatic cytochrome P-450 and b₅ and with a reduction in necrotic changes in the liver.     

A case of unusual suicidal poisoning by the organophosphorus insecticide dimethoate

A 20-year-old male who attempted suicide by injecting subcutaneously 10 ml of Sistemin 40 (40% dimethoate) was admitted 16 h later. General weakness, muscular fibrillations and a marked inhibition of red blood cell and serum cholinesterases were the prominent signs of intoxication. The antidotal treatment of intermittent boluses of atropine, oxime HI-6 and diazepam was combined with symptomatic therapy. Cholinesterase activity decreased within the next 3 d. In contrast to the marked general improvement of the patient, the return of cholinesterase activities was very slow. The patient was discharged 24 d after the poisoning with no notable consequences which could be ascribed to the intoxication.     

Enantioselectivity in zebrafish embryo toxicity of the insecticide acetofenate

Enantioselectivity in separation and toxicology of chiral xenobiotics have become one of the frontier topics interfacing chemistry and toxicology. In this study, enantiomers of insecticide acetofenate (AF) were separated on selected chiral columns by HPLC, and enantioselectivity in developmental toxicity was evaluated using the zebrafish embryo-larval assays. The AF enantiomers were baseline separated on Chiralcel OD, Chiralpak AD, and Sumichiral OA-2500I columns under optimized conditions. Pure enantiomers were obtained on Chiralcel OD. Optical rotatory dispersion (ORD) and circular dichroism (CD) detectors were used to determine the elution order and CD spectra of the enantiomers. The absolute configuration of enantiomers was identified as S-(+)-AF and R-(-)-AF by the octant rule from force-field calculations and CD spectra. The individual enantiomers were used in 4-day zebrafish embryo-larval bioassays, and a series of developmental end points were measured and compared. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to investigate the expressions of estrogen receptor alpha (ERalpha) in zebrafish embryo exposed to varying enantiomers. While the enantiomers showed no difference in acute toxicity, significant enantioselectivity was observed in developmental toxicities such as yolk sac edema and pericardial edema. The data of qRT-PCR showed that there was about 3.2-fold induction in the mRNA levels of ERalpha between fish exposed to (+)-AF and (-)-AF. The results suggest that enantioselectivity may occur at the developmental level even in the absence of selective acute toxicity and should be considered when evaluating ecotoxicological effects of chiral contaminants.     

Resistance to organophosphorus agent toxicity in transgenic mice expressing the G117H mutant of human butyrylcholinesterase

Organophosphorus toxicants (OP) include chemical nerve agents and pesticides. The goal of this work was to find out whether an animal could be made resistant to OP toxicity by genetic engineering. The human butyrylcholinesterase (BChE) mutant G117H was chosen for study because it has the unusual ability to hydrolyze OP as well as acetylcholine, and it is resistant to inhibition by OP. Human G117H BChE, under the control of the ROSA26 promoter, was expressed in all tissues of transgenic mice. A stable transgenic mouse line expressed 0.5 microg/ml of human G117H BChE in plasma as well as 2 microg/ml of wild-type mouse BChE. Intestine, kidneys, stomach, lungs, heart, spleen, liver, brain, and muscle expressed 0.6-0.15 microg/g of G117H BChE. Transgenic mice were normal in behavior and fertility. The LD50 dose of echothiophate for wild-type mice was 0.1 mg/kg sc. This dose caused severe cholinergic signs of toxicity and lethality in wild-type mice, but caused no deaths and only mild toxicity in transgenic animals. The mechanism of protection was investigated by measuring acetylcholinesterase (AChE) and BChE activity. It was found that AChE and endogenous BChE were inhibited to the same extent in echothiophate-treated wild type and transgenic mice. This led to the hypothesis that protection against echothiophate toxicity was not explained by hydrolysis of echothiophate. In conclusion, the transgenic G117H BChE mouse demonstrates the factors required to achieve protection from OP toxicity in a vertebrate animal.