| More than 30% of the current used organophosphorus (OPs) insecticides are chiral. The studies of enantioselectivity in the aquatic toxicity of chiral OPs have been steadily increasing, however, research concerning enantioselectivity in the toxicity of OPs to nontarget and nonmammalian species, particularly soil organisms, is still inadequate. In this study, the acetylcholinesterase (AChE) activity and coelomocytes lysosomal membrane stability of earthworm (Eisenia fetida) were selected as biomarkers, and the enantioselective effects of three chiral OPs (methamidophos, malaoxon, and isomalathion) were evaluated. Related research content and results are as follows:(1)The inhibition of acetylcholinesterase (AChE) activity by the chiral OPs all showed significant enantioselectivity. In the vivo research, the IC50 value of (±)-methamidophos on the inhibition of the AChE was 0.0281μg/cm2, the value of (+)-methamidophos was 0.0860μg/cm2,with 3.1 fold differences between them. The most toxic of malaoxon to AChE was (+)-malaoxon, the IC50 value was 0.0014μg/cm2, the value of the weakest ((-)-malaoxon)was 0.0058μg/cm2, with 4.1 fold differences. The most obvious difference in the three chiral OPs was 1R,3R-isomalation with (±)-isomalation, they were 331 fold differences. From the value of inhibition kinetic constants we also could find enantioselective difference, the value of ki expression the degree of AChE inhibition, the value of ki of (-)-methamidophos was 5 times higher than (+)-methamidophos, and (+)-malaoxon was 3.23 times higher than (-)-malaoxon, as to the four stereoisomers of isomalation the order of toxicity was 1R,3R-isomalation> 1S,3R-isomalation>1S,3S-isomalation>1R,3S-isomalation,with 1.412.12 fold differences. The relationship between the concentratin of organophosphorus pesticide and AChE inhibtion showed obvious dose effect relationship, with the increase in inhibitory concentration, result in stronger inhibition of enzyme activity. The enzyme inhibition dynamic assays further indicated that both the spatial orientations and phosphorylation properties of different stereoisomers towards AChE were different.(2)The research of using neutral red retention time (NRRT) evaluation lysosomal membrane stability of coelomocytes showed organophosphorus pesticide contamination could injure lysosomal membrane stability of earthworm. As the concentration of (±)-methamidophos was 0.0016μg /cm2, the value of NRRT was 76.88min, with the concentration increase to 0.16μg/cm2, the value of NRRT reduced to 29.78min, the value of NRRT of (±)-malaoxon dropped from 61.13min to 34.13min, (±)-isomalation's from 47.60min to 34.32min. The shorter neutral red retention time was, the worse the stability of lysosomal membrane, the more serious damage of lysosomal membrane. Also these researches showed enantioselective effect, but no significant difference contrast to acetylcholine activity study, the differences were maintained at 2 times under. Earthworms as part of invertebrates, its coelom is an important component of the immune system, illustrated that organophosphorus pesticide-contaminated will result in immune injury of earthworms.(3)The recovery of AChE of the earthworm after treated by malathion, was studied that malathion inhibited acetylcholine of earthworms is a reversible process, after 48h exposure treatment, AChE activity of earthworms was inhibited 80%,when transfer to a clean soil within 7 days after,sustained inhibition enhanced,after, AChE activity gradually recovered,when detectioned in the 30th day, AChE activity was only inhibited 27.7%,AChE activity recovered 70%.(4)The comet assay was used to evaluate the damagement of coelomocytes DNA of earthworm by malaoxon, the result showed that malaoxon cause very serious DNA damage, when at the lowest expose concentration of 0.0016μg/cm2, Very obvious smearing was observed, and on the control group also have a little degree of injury, We believed that high levels of genetic damage in itself was a universal feature of earthworms.
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