The Ecological Effect Of Parathion Pollution On Soil FDA Hydrolase And Its Mechanism

Parathion（PTH）,as one of organophosphorus pesticides with broad spectrum,high efficiency and low cost,has played an important role in ensuring crop harvest and food security in agricultural production.Due to its excessive application in the field,landfill of waste products and inadvertent spillage of containers,PTH could accumulate in soil environment for considerable long time and cause soil pollution,which has posed serious threats to human and animal health and soil ecology security.Therefore,evaluating the ecotoxicity of soil contaminated by PTH and sustainable utilization of soil resources after its remediation have become a hotspot of current research.Soil enzymes and microorganisms are important components of soil,which can reflect the stress responses of soil to exogenous environment pollution and play significant roles in evaluating the ecotoxicity of contaminated soil.However,the current ecological risks of PTH mainly focus on its fate and transformation and bioaccumulation in the environment,there is a lack of deep understanding of the toxic effects of soil ecological functions,especially soil enzymes and their toxic response mechanisms.Additionally,the potential environmental risks to soil ecology induced by PTH degradation process were also ignored in previous studies.This paper systematically investigated the toxic effects of PTH pollution on soil ecological function through laboratory simulation experiment,the main contents of this paper were to（ⅰ）investigate the effects of PTH on the activity of sensitive soil enzyme-FDA hydrolase and its kinetics and thermodynamic characteristics under acute stress,（ⅱ）analyze the mechanism of PTH on the inhibition of FDA hydrolase activity and its regulation factors of ecotoxicity,（ⅲ）investigate the contribution of soil microorganisms and chemical hydrolysis to the degradation of PTH through natural and abiotic experiments,and reveal the effects of PTH and its degradation process on soil ecological function and kinetics of FDA hydrolase,.（ⅳ）elucidate the toxicity evolution characteristics of soil microbial activities including FDA hydrolase activity,microbial biomass carbon（MBC）,basal respiration（BR）and microbial metabolic entropy（q CO2）during PTH degradation based on ecological models and integrated biomarker response（IBR/n）index,which could provide rich microbial theory for evaluating the ecological toxicity and environmental risk of PTH contaminated soil.The main research contents and results of this paper are as follows:（1）The toxic effect of PTH on soil FDA hydrolase activity was investigated through acute contamination experiment.It was found that PTH significantly inhibited the activity of FDA hydrolase,which was negatively correlated with PTH concentration,indicating that FDA hydrolase was very sensitive to PTH pollution.Based on the relative enzyme activity of FDA hydrolase,the ecological dose ED₁₀ and ED₅₀values of PTH acute pollution were calculated by dose-response model,which were ranged from 36.1 to 136.6 mg kg^-1 and from 324.7 to1229.7 mg kg^-1,respectively.There were small toxic effects of PTH on FDA hydrolase in Jiangsu paddy soil,Yunnan lateritic red earths and Jilin black soil,indicating that soil with higher fertility could buffer the detrimental effects of PTH on FDA hydrolase.Stepwise regression analysis revealed that soil organic matter（SOM）and available potassium（AK）were significantly positively correlated with the ED value of FDA hydrolase relative activity,and the regression coefficient of the former was larger than the latter,suggesting that SOM was the main key soil factor affecting the toxicity of PTH on FDA hydrolase.（2）The inhibitory mechanism of PTH on the activity of FDA hydrolase was investigated through kinetic and thermodynamic analysis.Based on the changing characteristics of kinetic parameters along with pollution concentration,it was deduced that the inhibition types of PTH on soil FDA hydrolase activity belonged to noncompetitive inhibition,uncompetitive inhibition and linear mixed inhibition that the competitive inhibition type was the main type.The competitive inhibitory constants K_ic and noncompetitive inhibitory constants K_iu of FDA hydrolase under PTH stress ranged from 0.064 to 0.447 m M and 0.209 to 0.723 m M,respectively,which were larger than the K_m in values.This indicated that the affinity of PTH to FDA hydrolase and the binding strength of PTH to the enzyme-substrate complex were lower than that of the enzyme to substrate.Thermodynamic analysis showed that increasing temperature could promote the reaction rate constant k of FDA hydrolase and alleviate the toxicity of PTH on FDA hydrolase.The thermodynamic parameters E_a,ΔG*,ΔH*andΔS*of soil FDA hydrolase exhibited a linear increase with the increase of PTH concentration.The deceleration process of soil FDA hydrolase enzymatic reaction induced by PTH was mediated by enthalpy control process.Soil properties played important roles in affecting the mechanism and ecotoxicity of PTH on FDA hydrolase,and SOM and total nitrogen（TN）are the main factors influencing the PTH toxicity.（3）The degradation characteristics of PTH in red soil and coastal solonchaks under natural and abiotic conditions were studied.It was found that the maximum degradation percentage of PTH in red soil was 50.1%～75.5%for natural condition and 25.0%～69.8%for abiotic condition,and the t_1/2was 54.6-117.5 and 66.6-147.8 days under natural and abiotic conditions,respectively.PTH was completely degraded at 30,45,60,90 and 120 days under two conditions in coastal solonchaks,which corresponded to the pollution concentration of 50,100,250,500 and 1000 mg kg^-1,respectively.And the degradation half-life(t_1/2)of PTH in coastal solonchaks ranged from 5.6 to 56.8 days and 12.1 to 130.8 days for natural and abiotic conditions,respectively.There were great differences in the percentage of PTH degradation that influenced by soil types under natural and abiotic environments,showing that the chemical stability of PTH in red soil was stronger than that in coastal solonchaks.Besides,the chemical hydrolysis was the main transformation behavior leading to the PTH degradation in the tested soils,and the capacity of biodegradation of coastal solonchaks was stronger than that of red soil.（4）The effects of PTH and its degradation process on microbial activity of red soil and coastal solonchaks were investigated to obtain the environmental risk evolution characteristics of PTH during degradation on the tested soils.Results showed that although the PTH residues in two soils decreased along with incubation time under nature and abiotic process,the activities of FDA hydrolase for tested soils declined on the pollution concentration gradient.In addition,there were obvious differences in the changes of FDA hydrolase activity on the time scale for two soils.For instance,the FDA hydrolase activity for red soil maintained steady in the early stage and increased slowly in the late stage overall,while the trend for coastal solonchaks was opposite to that of red soil.Furthermore,different concentrations of PTH inhibited MBC in both tested soils during natural degradation,the BR and q CO₂ decreased significantly with the increase of incubation time.There was no significant increase in MBC under abiotic condition.Correlation analysis showed that the PTH residue was negatively correlated with FDA hydrolase activity and MBC in coastal solonchaks during natural degradation,while the relationship was not observed for red soil.Meanwhile,the FDA hydrolase activities for two soils were significantly positively correlated with MBC,BR and q CO₂ under the same time at different PTH concentrations,MBC and BR exhibited significantly positive correlation overall.Two-way ANOVA analysis demonstrated that microbial activities were negatively affected not only by PTH dose and incubation time but also by their interactions.The variation range of IBR/n index varied from 0 to 3.1 for red soil and 0.46 to 5.90 for coastal solonchaks on the time scale,indicating that the toxicity of PTH to soil microorganisms during its natural degradation gradually decreased along with the time.However,the toxicity of PTH and its metabolic risk substances to the two soils at the end of experiment was stronger than that at the initial time,and the toxicity and environmental risk of PTH to coastal solonchaks were higher than that of red soil.（5）The functional adaptability characteristics of two soils were reflected by the kinetic parameters of FDA hydrolase during PTH degradation.The kinetic parameters K_m,V_max and V_max/K_m of FDA hydrolase decreased significantly with the increase of PTH concentration under natural and abiotic conditions in two soils,indicating that the inhibition mechanism of PTH on the activity of FDA hydrolase was the same in the degradation process,belonging to the uncompetitive inhibition type.There were slow increases in K_m and V_max of FDA hydrolase for tested soils during PTH natural degradation on the time scale.This suggested that soil microorganisms secreted isoenzymes in response to the toxic effect during PTH degradation,thus increasing the potential amount of enzymes in soil.Correlation analysis showed that there were significant positive correlations between FDA hydrolase activity and MBC and V_max,as well as the kinetic parameters K_m and V_max/K_m,which confirmed the increase of enzyme amount in soils during PTH degradation was caused by microbial proliferation and secretion.The dynamic changes of the K_m,K_iu and ED values of FDA hydrolase for two soils could reflect the ecological functions adaptability of two soils during PTH degradation,and showed that the capacity of coastal solonchaks to PTH toxicity was better than the red soil,and the microorganisms played a vital role in the soil quality recovery ability under PTH pollution.To sum up,the activity of soil FDA hydrolase was inhibited by PTH pollution under acute stress through three forms,and the deceleration step in the enzymatic reaction was regulated by enthalpy or entropy process,the chemical hydrolysis of PTH transformation in soil was stronger than that of microbial degradation,and the ecotoxicity of PTH to soil remained severe during PTH degradation.This study not only expanded the knowledge of the responses and mechanisms of soil enzymes to PTH pollution,but also provided rich soil enzymology theory for the risk assessment of soil PTH pollution.