Studies On The Enrichment Pollution Of Mixed Pesticides In The Soil And Combined Toxicity To The Earthworms

Mixed use of pesticides is very common in agricultural production,traditional research and related standards only assess the environmental risk of a single agent,often underestimate the potential threat of mixed pesticides.To reveal the enrichment mechanism and toxicological effects of mixed pesticides in soil,in this study,the three main pesticides avermectin,imidacloprid and carbendazim of cowpea in Hainan Province were used for analysis of enrichment and degradation in soil after mixed application,acute toxicity,oxidative stress toxicity and avoidance toxicity to soil indicator organisms--earthworm under compound contamination conditions,which provide theoretical basis and technical guidance for the safe use of three pesticides in Hainan Province,the main results were as follows:1、The degradation residues of three pesticides in soil was determined by HPLC with modifications.The instrument detection methods were detector:PDA detector;chromatographic column:C₁₈（4.6×250 mm）stainless steel column;column temperature:30℃;flow rate:1.0 m L/min;wavelength:245 nm;injection:10μL;gradient elution program.The pretreatment method was soil shock extraction 3 times.The average recovery rates of three pesticides in soil using this method were from80%to 104%,the daily RSD value was between 2.8%to 12.5%,while the daytime RSD value was between 1.9%to 4.1%,respectively.All results were in accordance with the pesticide residue test guidelines.2、Through the above method,the pesticide residues were detected in soil samples of dynamic field,and the degradation and enrichment of 3 pesticides in soil after mixed spraying were obtained.Degradation half-lives of avermectin,imidacloprid and carbendazim in soil were 6.93,4.81 and 4.01 d,respectively.All of them were degradable pesticides.The degradation dynamic study found that the degradation half-life of pesticides in soil increase along with the increase of the amount of pesticides applied,while the degradation rate of avermectin,imidacloprid and carbendazim at the same time was the slowest,degradation half-life were 10.19,7.79 and 6.30 d,respectively.The avermectin,imidacloprid and carbendazim were applied separately and mixed for three times,and the residues in soil were 0.154,0.223 and 4.328 mg/kg,and 0.187,0.287 and 5.775 mg/kg at 7 d,respectively.With the increase of application times,pesticides were enriched in soil,the residue increases significantly with the increase of application times and pesticide quantity.3、Acute toxicity of three pesticide single agent and its mixture to Eisenia fetida were studied by the contact filter paper method and the artificial soil method respectively.The results of the contact filter paper method showed that the 48h-LC₅₀values of avermectin,imidacloprid and carbendazim to earthworms were 8.89×10^-3,5.56×10^-5,>10 mg/cm²,respectively.The order of toxicity was imidacloprid>avermectin>carbendazim.The artificial soil analysis showed that the 14d-LC₅₀of avermectin,imidacloprid and carbendazim were 26.81,3.18,8.48 mg/kg dw,respectively.The order of toxicity was imidacloprid>carbendazim>avermectin.Reference OECD and GBT21809-2008 standards,the toxicity grade of avermectin,imidacloprid and carbendazim to earthworms was low toxicity,middle toxicity and middle toxicity,respectively.The mixing results of both methods showed that the toxicity to earthworms increased exponentially with the number of pesticides.4、The additive index method and the combined-index method were used to evaluate the mixture interaction.The true values coincide with most of the two evaluation curves,indicating that the accuracy of the two evaluation methods is high.The 48 h of the avermectin+imidacloprid treatment group in the contact filter paper test were synergetic,the other treatment groups were additive in the additive index method.The combined-index method was the synergetic effect in 24 h of the avermectin+imidacloprid treatment group in the contact filter paper method,all the mixture combinations showed antagonistic effect in 7 d in the artificial soil method,the binary mixture of imidacloprid+carbendazim treatment group still showed antagonistic effect,the treatment group of avermectin+imidacloprid and avermectin+carbendazim showed additive effect,and the treatment group of ternary mixture of avermectin+imidacloprid+carbendazim treatment group showed synergistic effect at 14 d.The combined-index method can evaluate the combined toxicity more quickly and accurately.5、The effects of avermectin,imidacloprid and carbendazim single and mixed agents on the antioxidant enzyme system of earthworms were investigated using SOD and CAT activity.The results showed that when earthworms were subjected to avermectin,imidacloprid and carbendazim stress,the SOD activity in vivo increased significantly,reached a peak at 21 d,28 d decreased significantly but still higher than that of control,the SOD activity under mixed pesticide stress was earlier activated than that of single agent and higher than that of single agent at 28 d,which was positively correlated with the concentration.Over time,the CAT activity of earthworms increased significantly,in which imidacloprid treatment group was CAT inhibited at 7 d and significantly activated at 14 d.The CAT activity was not related to pesticide concentration.6、The avoidance behavior of earthworms against avermectin,imidacloprid,carbendazim single and its mixture agent was determined by two-compartment avoidance reaction device.The results showed that all seven treatment had avoidance effect in the set concentration range,the 48h-AC₅₀values of avermectin,imidacloprid and carbendazim to earthworm were 0.28,0.03,0.05 mg/kg dw,respectively,The sensitivity was imidacloprid>carbendazim>avermectin.With the addition of pesticide,the soil conditions were more severed for earthworms,earthworms were more prone to avoidance behavior.AC₅₀of avoidance behavior outcomes was more acute than toxicity results LC₅₀two orders of magnitude,and earthworms to pesticide avoidance trend was like acute toxicity results.These results indicated that avoidance response can be more rapid and accurate response to pesticide hazards in the environment.This study provide theoretical support for the revision of residue limit standards and the safe use of mixed pesticides in the field.