Piperonyl butoxide, a synergist of pesticides can elicit male-mediated reproductive toxicity.

A semi-synthetic methylenedioxyphenyl compound piperonyl butoxide (PBO) has been used as a ubiquitous synergist to increase the insecticidal effect of pesticides for agricultural and household use. Despite previously demonstrated effects of PBO, the detailed mechanism of PBO in spermatozoa and reproductive toxic effects on male germ cells have not been fully elucidated. Therefore, this study evaluated the effects of PBO on various sperm functions during capacitation and clarified the mechanisms of reproductive toxic effects on male fertility at different concentrations of PBO (0.1, 1, 10, and 100 µM). Sperm motility and kinematics were assessed using computer-assisted sperm analysis and the status of capacitation was evaluated using combined H33258/chlortetracycline (CTC) staining. Intracellular adenosine triphosphate (ATP) and cell viability levels were also measured. In addition, protein kinase A (PKA) activity and protein tyrosine phosphorylation were evaluated. In addition, in vitro fertilization was performed to determine the effects of PBO on cleavage and blastocyst formation rates. We found that PBO significantly decreased sperm motility, kinematics, and acrosome-reacted and capacitated spermatozoa. In addition, PBO suppressed the intracellular ATP levels and directly affected cell viability. Moreover, PBO detrimentally decreased the activation of PKA and altered the levels of tyrosine-phosphorylated proteins. Consequently, cleavage and blastocyst formation rates were significantly reduced in a dose-dependent manner. In line with our observations, the synergist of pesticides PBO may directly and/or indirectly cause disorder in male fertility. Hence, we suggest that careful attention is made to consider reproductive toxicity when using PBO as a synergist.

Piperonyl butoxide: Mode of action analysis for mouse liver tumour formation and human relevance.

In a 79 week bioassay the pesticide synergist piperonyl butoxide (PBO) was shown to significantly increase the incidence of hepatocellular adenoma (but not hepatocellular carcinoma) in male CD-1 mice at dietary levels of 100 and 300 mg/kg/day PBO and in female mice at a dietary level of 300 mg/kg/day. As PBO is not a genotoxic agent, a series of investigative studies were undertaken to elucidate the mode of action (MOA) for PBO-induced mouse liver tumour formation. Male CD-1 mice were fed diets to provide intakes of 0 (control), 30, 100 and 300 mg/kg/day PBO and for purposes of comparison 500 ppm sodium phenobarbital (NaPB), a known constitutive androstane receptor (CAR) activator, for 7 and 14 days. Treatment with 100 and 300 mg/kg/day PBO and 500 ppm NaPB increased relative liver weight which was associated with hepatocyte hypertrophy, with hepatocyte replicative DNA synthesis (RDS) being increased after 7 days treatment. The treatment of CD-1 mice with 30-300 mg/kg/day PBO for 14 days resulted in significant dose-dependent increases in hepatic microsomal cytochrome P450 (CYP) content and 7-pentoxyresorufin O-depentylase (PROD) activity and in hepatic Cyp2b10 mRNA levels. In contrast, PBO produced a biphasic effect on markers of activation of the peroxisome proliferator-activated receptor alpha (PPARα), with small increases in microsomal lauric acid 12-hydroxylase activity and hepatic Cyp4a10 mRNA levels being observed in mice given 100 mg/kg/day with PBO, with either no increase or a significant inhibition being observed in mice given 300 mg/kg/day PBO. The hepatic effects of PBO in male CD-1 mice were generally similar to those produced by NaPB and were reversible after the cessation of treatment for 28 days. Studies were also performed in male C57BL/6J (wild type) mice and in hepatic CAR and pregnane X receptor (PXR) knockout mice (CAR KO/PXR KO mice), where in the CAR KO/PXR KO mice PBO had little effect on markers of CAR activation, but produced some increases in markers of PPARα activation. The treatment of male CD-1 mouse hepatocytes for 4 days with 5-50 µM PBO, 10-1000 µM NaPB and 25 ng/mL epidermal growth factor (EGF) resulted in significant increases in hepatocyte RDS. While treatment of hepatocytes from one male and one female human donor with 5-500 µM PBO and 10-1000 µM NaPB for 4 days had no effect on hepatocyte RDS, treatment with EGF resulted in significant increases in RDS in both human hepatocyte preparations. In summary, PBO is predominantly a hepatic CAR activator at carcinogenic dose levels in CD-1 mice, with activation of hepatic CAR resulting in a suppression of the effect of PBO on hepatic PPARα. A robust MOA for PBO-induced mouse liver tumour formation has been established, this MOA being similar to that previously identified for NaPB and some other rodent liver CAR activators. Based on the lack of effect of PBO on RDS in human hepatocytes, it is considered that the MOA for PBO-induced mouse liver tumour formation is qualitatively not plausible for humans.

Integrated analysis of in vitro data and the adverse outcome pathway framework for prioritization and regulatory applications: An exploratory case study using publicly available data on piperonyl butoxide and liver models.

The integration of existing knowledge to support the risk assessment of chemicals is an ongoing challenge for scientists, risk assessors and risk managers. In addition, European Union regulations limiting the use of new animal testing in cosmetics makes already existing information even more valuable. Applying a previous SEURAT-1 program framework to derive predictions of in vivo toxicity responses for a compound, we selected piperonyl butoxide (PBO) as a case study for identification of knowledge and methodology gaps in understanding a compound's effects on the human liver. This is investigated through integration of data from human in vitro transcriptomics studies, biological pathway analysis, chemical and disease associations, and adverse outcome pathway (AOP) information. The outcomes of the analysis are used to generate AOPs of liver-related endpoints, identifying areas of concern for risk assessors and regulators. We demonstrate that integration of data through already existing and publicly available tools can produce outcomes comparable to those that may be found through more conventional time- and resource-intensive methods. It is also expected that, with more refinement, this approach could in the future provide evidence to support chemical risk assessment, while also identifying data gaps for which additional testing may be needed.

Toxicity and sublethal effects of fluralaner on Spodoptera litura Fabricius (Lepidoptera: Noctuidae).

The increasing occurrence of resistance to chemical insecticides in insect pest populations is a serious threat to the integrity of current pest management strategies, and exploring new alternative chemistries is one important way to overcome this obstacle. Fluralaner, as a novel isoxazoline insecticide, has broad spectrum activity against a variety of insect pests, but little data is available about its effect on Lepidopterans. The effects of fluralaner on Spodoptera litura Fabricius, a widespread and polyphagous pest, were evaluated in the present study. Our results showed younger larvae were more susceptible to fluralaner treatment, but feeding and topical applications were similarly effective in 3rd instar larvae. Synergism assays indicated that piperonyl butoxide (PBO) could increase the toxicity of fluralaner to S. litura to a certain degree and P450 may be involved in the detoxification of fluralaner in vivo. Sublethal developmental effects included reduced larval body weight, decreased pupation and emergence, and notched wings in adults, accompanied by changes in the transcript levels of chitinase 5 (CHT5) and juvenile hormone acid methyltransferase (Jhamt), genes vital for insect development. Above results manifested that fluralaner is highly toxic to S. litura larvae via either topical or oral application and provide an indication of how this insecticide is metabolized in vivo. Further, our results provided a foundation for further development of fluralaner as a new tool in insect pest management.

Toxicological actions of plant-derived and anthropogenic methylenedioxyphenyl-substituted chemicals in mammals and insects.

The methylenedioxyphenyl (MDP) substituent is a structural feature present in many plant chemicals that deter foraging by predatory insects and herbivores. With increasing use of herbal extracts in alternative medicine, human exposure to MDP-derived plant chemicals may also be significant. Early studies found that most MDP agents themselves possess relatively low intrinsic toxicity, but strongly influence the actions of other xenobiotics in mammals and insects by modulating cytochrome P-450 (CYP)-dependent biotransformation. Thus, after exposure to MDP chemicals an initial phase of CYP inhibition is followed by a sustained phase of CYP induction. In insects CYP inhibition by MDP agents underlies their use as pesticide synergists, but analogous inhibition of mammalian CYP impairs the clearance of drugs and foreign compounds. Conversely, induction of mammalian CYP by MDP agents increases xenobiotic oxidation capacity. Exposure of insects to MDP-containing synergists in the environment, in the absence of coadministered pesticides, may also enhance xenobiotic detoxication. Finally, although most MDP agents are well tolerated, several, typified by safrole, aristolochic acid, and MDP-kavalactones, are associated with significant toxicities, including the risk of hepatotoxicity or tumorigenesis. Thus, the presence of MDP-substituted chemicals in the environment may produce a range of direct and indirect toxicities in target and nontarget species.

Statistical evaluation of mortality in long-term carcinogenicity bioassays using a Williams-type procedure.

Several doses and a control group can be compared under order restriction using the Williams procedure for normally distributed endpoints assuming variance homogeneity. Comparison of the survival functions represents a secondary endpoint in long-term in vivo bioassays of carcinogenicity. Therefore, a Williams-type procedure for the comparison of survival functions is proposed for the assumption of the Cox proportional hazards model or the general frailty Cox model to allow a joint analysis over sex and strains. Interpretation according to both statistical significance and biological relevance is possible with simultaneous confidence intervals for hazard ratios. Related survival data can be analyzed using the R packages survival, coxme, and multcomp. Together with the R packages MCPAN and nparcomp, Dunnett- or Williams-type procedures are now available for the statistical analysis of the following endpoint types in toxicology: (i) normally distributed, (ii) non-normally distributed, (iii) score (ordered categorical) data, (iv) crude proportions, (v) survival functions, and (vi) time-to-tumor data with and without cause-of-death information.

Ultraviolet radiation increases sensitivity to pesticides: synergistic effects on population growth rate of Daphnia magna at low concentrations.

In the present study we aimed to investigate whether UV-B radiation can exacerbate effects of pesticides fenoxycarb, pirimicarb, and tebufenpyrad on the survival, reproduction, and population growth rate of the standard test species Daphnia magna. We applied sublethal pesticides' concentrations and UV doses and observed no effects on survival. However, we observed synergistic effects of UV and pesticides on both cumulative reproduction and population growth rate (21 days) for fenoxycarb (100 µg/L) and pirimicarb (10 µg/L), but a less-than-additive effect for tebufenpyrad (5-10 µg/L). In the series exposed to UV and fenoxycarb or pirimicarb, the population growth rate dropped down to 0.1, while in the control series it was around 0.3. The results indicate that concentrations of some toxicants that are nontoxic in standard tests can cause harmful population-level effects when combined with UV.

Evaluation of in vivo liver genotoxic potential of Wy-14,643 and piperonyl butoxide in rats subjected to two-week repeated oral administration.

Wy-14,643 (WY), a peroxisome proliferator-activated receptor-alpha agonist, and piperonyl butoxide (PBO), a pesticide synergist, induce oxidative stress and promote hepatocarcinogenesis in the liver of rodents. These chemicals belong to a class of non-genotoxic carcinogens, but DNA damage secondary to the oxidative stress resulting from reactive oxygen species generation is suspected in rodents given these chemicals. To examine whether WY or PBO have DNA-damaging potential in livers of rats subjected to repeated oral administration for 14 days, the in vivo liver comet assay was performed in partially hepatectomized rats, and the expression of some DNA-repair genes was examined. Then, to examine whether they have genotoxic potential, the in vivo liver initiation assay was performed in rats. In the comet assay, positive results were obtained at 3 h after the last treatment of WY, and some DNA-repair genes such as Apex1, Mlh1, Xrcc5, and Gadd45 were up-regulated in the liver. In the liver initiation assay, negative results were obtained for both WY and PBO. The results of the present study suggest that WY, but not PBO, causes some DNA damage in livers of rats, but such DNA damage was repaired by the increased activity of some DNA repair genes and may not lead to a DNA mutation.

Mechanistic study on hepatocarcinogenesis of piperonyl butoxide in mice.

To clarify the mechanism of piperonyl butoxide (PBO)-induced hepatocarcinogenesis in mice, male mice were subjected to a two-thirds partial hepatectomy, N-diethylnitrosamine (DEN) initiation, and a diet containing 0.6% PBO for eight weeks. The incidence of gamma-glutamyl transpeptidase (GGT)-positive foci and PCNA-positive cells was significantly increased in the DEN + PBO group compared with the DEN-alone group. Real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis showed up-regulation of genes related to metabolism, such as cytochrome P450 1A1 and 2B10, and metabolic stress, such as Por, Nqo1, Nrf2, abcc3, and abcc4. Early responsive genes downstream of mitogen-activated protein kinase (MAPK), such as c-fos, c-jun, c-myc, and activating transcription factor 3 (ATF3), were also up-regulated in this group. Positive immunohistochemical staining for ATF3 was diffusely observed in nonproliferating hepatocytes of the DEN + PBO group, but altered foci were negative or weakly positive for ATF3. The nuclei of hepatocytes within ATF3-negative foci were positive for cyclin D. Thus PBO can induce oxidative stress, activate the MAPK pathway, and increase ATF3 transcript levels in hepatocytes outside the altered foci during the early stage of PBO-induced hepatocarcinogenesis in mice.

Threshold dose of piperonyl butoxide that induces reactive oxygen species-mediated hepatocarcinogenesis in rats.

To determine the threshold dose of piperonyl butoxide (PBO) that induces hepatocellular tumor-promoting effects, reactive oxygen species (ROS) generation, and drug-metabolizing enzymes that protect against ROS generation, partial hepatectomized rats were fed diets containing 0, 0.015, 0.03, 0.06, 0.125, 0.25, or 0.5% PBO after an i.p. injection of N-diethylnitrosamine (DEN) to initiate hepatocarcinogenesis. Histopathologically, Glutathione S-transferase placental form (GST-P)-positive foci were significantly increased in a dose-dependent manner in rats given 0.25% PBO or higher. The formation of microsomal ROS in the liver was significantly increased in 0.25 and 0.5% PBO. Real-time RT-PCR showed that the expression of the CYP1A1, UDPGTr-2, and Mrp3 genes was significantly upregulated in rats given 0.03% PBO or higher. These results suggest that 0.25% is the threshold dose of PBO that induces ROS-mediated hepatocarcinogenesis in rats, although the CYP1A1 gene that is related to ROS generation and the UDPGTr-2 and Mrp3 genes that are involved in protection against ROS were induced in the livers of rats even at a PBO dose of 0.03%.

Molecular pathological analysis for determining the possible mechanism of piperonyl butoxide-induced hepatocarcinogenesis in mice.

Piperonyl butoxide (PBO), alpha-[2-(2-butoxyethoxy)ethoxy]-4,5-methylene-dioxy-2-propyltoluene, is widely used as a synergist for pyrethrins. In order to clarify the possible mechanism of non-genotoxic hepatocarcinogenesis induced by PBO, molecular pathological analyses consisting of low-density microarray analysis and real-time reverse transcriptase (RT)-PCR were performed in male ICR mice fed a basal powdered diet containing 6000 or 0 ppm PBO for 1, 4, or 8 weeks. The animals were sacrificed at weeks 1, 4, and 8, and the livers were histopathologically examined and analyzed for gene expression using the microarray at weeks 1 and 4 followed by real-time RT-PCR at each time point. Reactive oxygen species (ROS) products were also measured using liver microsomes. At each time point, the hepatocytes of PBO-treated mice showed centrilobular hypertrophy and increased lipofuscin deposition in Schmorl staining. The ROS products were significantly increased in the liver microsomes of PBO-treated mice. In the microarray analysis, the expression of oxidative and metabolic stress-related genes--cytochrome P450 (Cyp) 1A1, Cyp2A5 (week 1 only), Cyp2B9, Cyp2B10, and NADPH-cytochrome P450 oxidoreductase (Por) was over-expressed in mice given PBO at weeks 1 and 4. Fluctuations of these genes were confirmed by real-time RT-PCR in PBO-treated mice at each time point. In additional real-time RT-PCR, the expression of Cyclin D1 gene, key regulator of cell-cycle progression, and Xrcc5 gene, DNA damage repair-related gene, was significantly increased at each time point and at week 8, respectively. These results suggest the possibility that PBO has the potential to generate ROS via the metabolic pathway and to induce oxidative stress, including oxidative DNA damage, resulting in the induction of hepatocellular tumors in mice.

Toxicity of aflatoxin B1 to Helicoverpa zea and bioactivation by cytochrome P450 monooxygenases.

Infestation of corn (Zea mays) by corn earworm (Helicoverpa zea) predisposes the plant to infection by Aspergillus fungi and concomitant contamination with the carcinogenic mycotoxin aflatoxin B1 (AFB1). Although effects of ingesting AFB1 are well documented in livestock and humans, the effects on insects that naturally encounter this mycotoxin are not as well defined. Toxicity of AFB1 to different stages of H. zea (first, third, and fifth instars) was evaluated with artificial diets containing varying concentrations. Although not acutely toxic at low concentrations (1-20 ng/g), AFB1 had significant chronic effects, including protracted development, increased mortality, decreased pupation rate, and reduced pupal weight. Sensitivity varied with developmental stage; whereas intermediate concentrations (200 ng/g) caused complete mortality in first instars, this same concentration had no detectable adverse effects on larvae encountering AFB1 in fifth instar. Fifth instars consuming AFB1 at higher concentrations (1 microg/g), however, displayed morphological deformities at pupation. That cytochrome P450 monooxygenases (P450s) are involved in the bioactivation of aflatoxin in this species is evidenced by the effects of piperonyl butoxide (PBO), a known P450 inhibitor, on toxicity; whereas no fourth instars pupated in the presence of 1 mug/g AFB1 in the diet, the presence of 0.1% PBO increased the pupation rate to 71.7%. Pupation rates of both fourth and fifth instars on diets containing 1 mug/g AFB1 also increased significantly in the presence of PBO. Effects of phenobarbital, a P450 inducer, on AFB1 toxicity were less dramatic than those of PBO. Collectively, these findings indicate that, as in many other vertebrates and invertebrates, toxicity of AFB1 to H. zea results from P450-mediated metabolic bioactivation.

Structure-activity relationship study of alkynyl ether insecticide synergists and the development of MB-599 (verbutin).

Structure-activity relationships of aryl alkynyl synergists of the general formula of Ar-Q-R, where Q represents a bridging structure, were studied using a standardised testing system and Relative Potency values. Ethers, esters, oxime ethers, amides and amines were prepared and evaluated. The length of the R-alkynyl chain, the role of the bridge and the substitution of the aromatic ring were examined systematically. The most potent compounds possessed an aromatic ring connected via a bridge of three atoms to an alkynyl chain, forming together a linear side-chain of six atoms. Several highly potent compounds were synthesised of which one (MB-599; proposed common name verbutin) was selected for development as a selective insecticide synergist in crop protection. Its high potential at practical insecticide:synergist ratios makes possible the reduction of the total amount of insect-control chemicals applied, and its use as an additive to produce new formulations of existing insecticides makes it highly advantageous in resistance management, giving a new tool to sustain the effectiveness of a wide range of insecticides. A product containing a (1+1) mixture of verbutin and beta-cypermethrin was launched in Hungary in 2002.

Oncogenicity studies of piperonyl butoxide in rats and mice.

1. The oncogenicity of Piperonyl butoxide (PBO) has been studied in the mouse and rat. CD-1 mice were administered PBO in the diet at target doses of 0, 30, 100 and 300 mg/kg/day for 79 weeks and Sprague-Dawley rats 0, 30, 100 and 500 mg/kg/day for 104/105 weeks. 2. At termination of the study in the mouse there was evidence of increased liver weights and an increased incidence of eosinophilic adenomas at 100 and 300 mg/kg/day in males and 300 mg/kg/day in females. 3. In rats there was increased liver weights at 100 and 500 mg/kg/day associated with hepatocyte hypertrophy in both male and female rats. There was no increased incidence of neoplasia at any site. Hypertrophy and hyperplasia of thyroid follicles was observed at 500 mg/kg/day in both sexes. 4. The observations reflect the expected changes related to the induction of the mixed function oxygenase group of enzymes. In the mouse the increased incidence of eosinophilic adenomas is not considered relevant for human risk evaluation.

Effect of piperonyl butoxide on cell replication and xenobiotic metabolism in the livers of CD-1 mice and F344 rats.

Male CD- 1 mice were fed diets containing 0 (control), 10, 30, 100, and 300 mg/kg/day piperonyl butoxide (PBO) and 0.05% sodium phenobarbital (NaPB) and male F344 rats were fed diets containing 0 (control), 100, 550, 1050, and 1850 mg/kg/day PBO and 0.5% NaPB for periods of 7 and 42 days. In both species PBO and NaPB increased relative liver weight and whereas PBO produced a midzonal (mouse) or periportal/midzonal (rat) hypertrophy, NaPB produced a centrilobular hypertrophy. In the rat, individual cell necrosis was also observed at 42 days after high doses of PBO. Replicative DNA synthesis, assessed as the hepatocyte labeling index following implantation of 7-day osmotic pumps containing 5-bromo-2'-deoxyuridine during Study Days 0-7 and 35-42, was increased in mice given 300 mg/kg/day PBO and NaPB for 7 days and in rats given 550 and 1050 mg/kg/day PBO and NaPB for 7 days and 1050 mg/kg/day PBO for 42 days. While PBO had no effect on body weights in mice, the body weights of rats given 550, 1050, and 1850 mg/kg/day PBO for 42 days were reduced to 92, 89, and 70% of control, respectively. PBO induced microsomal cytochrome P450 content and mixed function oxidase activities in the mouse and rat, although the effects were less marked than those produced by NaPB. In summary, this data demonstrates that PBO can produce liver enlargement in the mouse and the rat which is associated with induction of xenobiotic metabolism, hypertrophy, and hyperplasia. The hepatic effects of PBO in the mouse were similar to but less marked than those produced by NaPB. In the rat high doses of PBO were hepatotoxic and resulted in a marked reduction in body weight. Thus while the reported formation of eosinophilic nodules in mouse liver by PBO may occur by a mechanism(s) similar to that of NaPB and other nongenotoxic enzyme inducers, the reported tumor formation in rats at greater than the maximum tolerated dose is most likely associated with marked enzyme induction in conjunction with a regenerative hyperplasia resulting from PBO-induced hepatotoxicity.

Chronic toxicity studies of piperonyl butoxide in CD-1 mice: induction of hepatocellular carcinoma.

Male and female CD-1 mice (51-104 mice/group) were administered piperonyl butoxide (alpha-[2-(2-butoxyethoxy)ethoxy-4,5-methylenedioxy-2-propyltol uene) in the diet at levels of 0 (control), 0.6 and 1.2% for 52 weeks (1 year). Hepatocellular carcinomas were induced in treated groups in a dose-dependent manner. The incidences of hepatocellular carcinoma were 11.3 and 52.0% in male mice given 0.6 and 1.2% piperonyl butoxide, and 41.2% in female mice given 1.2%. Piperonyl butoxide is thus a hepatocarcinogen to mice as it is known to be to rats.

Morphometric and immunohistochemical studies on atrophic changes in lympho-hematopoietic organs of rats treated with piperonyl butoxide or subjected to dietary restriction.

Changes observed in lympho-hematopoietic organs in rats given piperonyl butoxide may be attributable either to direct toxic effects or to undernutrition. Male F344 rats were therefore fed diet containing 2.5% piperonyl butoxide or subjected to a 64% restriction of food intake for 2 weeks. Marked inhibition of body weight gain, decreased white blood cell count, depletion of T/B lymphocytes in lymphoid tissues, hypoplasia of the bone marrow, and decreased proliferating cell nuclear antigen (PCNA) labeling indices in these tissues were seen in both dietary restriction and 2.5% piperonyl butoxide groups. The depletion of T lymphocytes in the thymus and spleen was stronger in the 2.5% piperonyl butoxide group, as indicated by PCNA labeling indices and image analysis of T lymphocyte areas of the spleen, however, the toxicological profile observed for the chemically treated group was essentially the same as for animals on the restricted diet. These results suggest that the lympho-hematopoietic findings in rats receiving 2.5% piperonyl butoxide are probably due to undernutrition resulting from a reduced food intake.

The effect of pesticides on carp (Cyprinus carpio L). Acetylcholinesterase and its biochemical characterization.

The activity and molecular forms of acetylcholinesterase (AChE) were characterized in tissues of the carp (Cyprinus carpio). Tissue AChE activity was determined in response to specific inhibitors (ethopropazine, BW 284 C51) or pesticides (CuSO4, paraquat (PQ), methidathion (MD)). The highest AChE activity was found in the serum (878 +/- 100 U/liter), followed by the brain (113 +/- 12 U/liter), heart (89 +/- 6 U/liter), and trunk muscle (35 +/- 5 U/liter). Experiments with specific choline esterase inhibitors revealed a very low amount of pseudocholinesterase in all tissues studied. The ratio of the membrane-bound to the cytoplasmic-free AChE molecular forms was increased in the order of brain, trunk muscle, and heart. In sera of fish treated with MD (2 ppm) there was an 80% inhibition of AChE lasting for 2 weeks. Treatment with CuSO4 or PQ (both 5 ppm) led to a 50% decrease in the serum AChE activity followed by a transient increase over the control level. After 2 weeks of chronic treatment, AChE activity in fish exposed to CuSO4 returned to the control level, whereas in fish treated with PQ an elevated level (130% when compared to the control level) of enzyme activity was found. Our present experimental data indicate that pesticides occurring in natural waters not only inhibit AChE activity in fish but may influence the resynthesis of the enzyme as well.

[Synergism of selected pesticides. 1. Acute oral toxicity in combined administration]. / Untersuchungen zum Koergismus ausgewählter Pestizide 1. Mitt. Akute orale Toxizität bei kombinierter applikation.

Comparative studies with male rats were performed to investigate the coergism of single oral doses of selected pesticides, the comparison being based on the lethal effects observed. The quantitative analysis of the dose-mortality relationship and the determination of the coergistic index evidenced a slight increase of the lethal effect with lindane-phosmet and lindane-carbaryl combinations. The combined administration of lindane and ethylenethiourea resulted in an additive lethal effect.