Tetravclines Transformation,Accumulation And In Vitro Diagnosis Model In Soil-vegetable System

Tetracyclines?TCs?is widely used drug in the world,and there is more and more research concern on their ecotoxicity.Soil types are very complex in China,it is important to understand TCs transformation,accumulation and ecotoxicity in soil-vegetable system,this can establish the model of health risk assessment in TCs contaminated soil-vegetable system.In present sdudy,both hydroponic and soil experiments were carried out and several advanced techniques,such as ELISA,TEM and FCM were used to study the TCs transformation,accumulation and ecotoxicity in soil-vegetable system,and establish the in vitro diagnosis modle of TCs pollution. The main coclusions are as follows:1.Greenhouse soil culture and hydroculture experiments were conducted to study the degradation of tetracycline?TC?,oxytetracycline?OTC?and chlortetracycline?CTC?in different types of soils and subsequent effects on germination,root growth and biomass yield of pakchio?Brassica campestris L.?.The results showed that root length was most sensitive to the antibiotics,followed by seed germination and biomass yield.In the hydroculture experiment,the sensivtivity of root length was in order of CTC>OTC>TC.The effect of tetracyclines on root growth of pakchoi varied among the soil types in the order of dark red soil>red soil>purplish clay soil>black soil,whereas the degradation rate of TCs varied in different types of soils in the order of black soil>purplish clay soil>red soil>dark red soil. In addition,differential degradation of the antibiotics in soil was noted and their degradation rate in the same types of soil changed in the order:OTC>CTC>TC. There were significant negative correlations between TCs residues and organic matter,NPK contents,or cation exchange capacity of the soils.The antibiotics degradation and phytotoxicity varied with soil type,which may be related to soil chemical and biochemical properties.2.Combined HL-7702 model,transmission electron microscope?TEM?and flow cytometry?FCM?technologies,in vitro revealed that the cytotoxicity of TCs to the human liver cell line HL-7702 was mainly due to the impairment of liver function?ALT and AST activities increase?,oxidative stress?SOD activity decrease,POD activity,H₂O₂ and MDA contents increase?,mitochondrial and rough endoplasmic reticulum lesions?MC and RER swell?and late cell apoptosis in liver cells that were exposed to the three antibiotics at concentrations ranging from 0 to 50 mg L-1,the sensitivity to HL-7702 cells was in order of CTC>OTC>TC.The results suggested that oxidative stress,mitochondrial and rough endoplasmic reticulum lesions cell apoptosis were the main reason for liver injury to TCs.The liver cytotoxicity bioassay can serve as an efficient tool for monitoring TCs contamination.Specifically,extracellular ALT,AST,SOD,POD,H₂O₂,MDA and cell apoptosis of HL-7702 cells can be used as biomarkers in ecotoxicologic tests of TCs pollution.3.ELISA detection technology,greenhouse soil culture was carried out to eastablish the risk assement of TCs in soil-vegetable sysetem.The results showed that three antibiotics?TC,OTC and CTC?concentrations in two types of soils were determined.TC,OTC,and CTC can enter into pakchoi root through soil,and only TC and CTC can transfer into pakchoi leaf,and the accumulation was in order of TC>CTC>OTC.The accumulaition of TC and CTC in root and leaf of pakchoi increased with increase in soil TCs,and TC and CTC accumulation in leaf was higer than that in root,and the accumulation of TC and OTC in root and leaf of pakchoi were higher in purplish clay than that in black soil after long time treatment.Comparing with safety intake levels for these TCs recommended by Food and Agriculture Organization of the United Nations?FAO?,the hazard quotient?HQ?values for single antibiotic for all three antibiotics were far below 1,and in the order of TC>CTC>OTC,indicating no chronic-toxic risks for adult who intake the pakchoi grown on single TCs contaminated soils.4.Using in vitro digestion method combined with HL-7702 model and TEM,FCM and ELISA detection technologies,we can estabilish cell dinogosis of TCs contamination in soil-vegetable system.The results showed that TC and CTC concentrations in the leaf of pakchoi grown on the purplish clay were higher than that of the black soil.The in vitro study revealed that the cytotoxicity of in vitro digestion of pakchoi grown on two contaminated soils to the human liver cell line HL-7702 was mainly due to the oxidative stress,mitochondrial and rough endoplasmic reticulum lesions,impairment of liver function.The final cell apoptosis in liver cells that was due to exposure of in vitro digestion of pakchoi grown on two contaminated soils with the TC and CTC at concentrations ranging from 0 to 20 mg kg-1.After long time treatment,the cytotoxicity was in the order:OTC?CTC>OTC,raw pakchoi>cooked pakchoi,purplish clay>black soil.The results demonstrated that the toxic effects of TCs in pakchoi on liver cell varied in different soils and different eating ways of pakchoi,and the cytotoxicity is aggravating with the TCs concentration increase.The preliminary results also suggest that the more fertilizer existed in the soil,the less TCs toxicity happened to human liver cell;moreover,it is suggested that under the condition of low leaf TCs concentration,cooked vegetable was much safer to human consumption than raw vegetable.So HL-7702 cells can be used as a tool to diagnose TCs pollution in soil-vegetable system.