Toxic Effects Of Diclofenac And Simvastatin On Daphnia Magna

As emerging environmental contaminants,pharmaceutical and personal care products?PPCPs?are widely used in our daily life.Diclofenac?DCF?and simvastatin?SV?,two typical PPCPs,have been frequently detected in various environmental media such as municipal wastewater.However,there is little research about the potential impact of DCF and SV exposure on aquatic invertebrates.In this study,daphnia magna,a typical crustacean invertebrate,was employed to study the toxic effects of diclofenac?DCF?and simvastatin?SV?on aquatic organisms.We investigated the toxic effects of diclofenac?DCF?and simvastatin?SV?on the physiological parameters?e.g.,survival,growth rate,and reproduction etc.?of daphnia magna,via a 21-d chronic toxicity test.We also evaluated the effects of diclofenac?DCF?and simvastatin?SV?on the expression of the genes related to the detoxification metabolism,growth,development and reproduction?e.g.,Cu/Zn SOD,CAT,MRP4 and MRP5?in acute exposure?up to 96 h?by RT-PCR.The LC₅₀ value at 48 h of diclofenac and simvastatin was 47 mg/L,and 435?g/L for daphnia magna,respectively.It showed that simvastatin has acute toxicity to daphnia magna.Effects of diclofenac on D.magna in a 21 days chronic toxicity showed that the molting frequency,days to the first brood,days to the first egg production and death rates?offspring?displayed a parabolic trend with the increase of diclofenac concentration.Eggs amount in the first production?n?and intrinsic growth rate decreased with the increase of chemical concentration.Days to the first brood and days to the first egg production under 50?g/L diclofenac exposure was significantly longer than the control group?p<0.05?,which means that diclofenac has an effect on the growth and multiplication of D.magna.We also found that the parameters of days to the first brood and days to the first egg production were more sensitive to the exposure of diclofenac,which could be used as a sensitive biomarker to characterize the chronic toxicity of diclofenac on daphnia magna.Simvastatin exposure in 21 days chronic toxicity showed that the sensitivity of D.magna in different concentration groups to simvastatin exposure was different.The days to the first egg production,total egg production number per individual,total number of broods per individual?n?,death rates?offspring?and intrinsic growth rate were more sensitive to simvastatin exposure than others,which could be used as sensitive parameters to characterize the chronic toxicity of simvastatin on D.magna.At high concentration of simvastatin?50?g/L?,the growth and reproduction capability of D.magna were significantly inhibited.In addition,with the increase of simvastatin concentration,days to the first egg production was delayed and the death rates?offspring?was increased.On contrary,eggs amount in the first production?n?,total number of broods per individual and intrinsic growth rate were decreased.It suggested that high concentration of simvastatin has toxic effects on the growth and reproduction of D.magna.The expression of Cu/Zn SOD CAT,MRP4 and MRP5 mRNA in D.magna showed that diclofenac could interfere with the normal functions of antioxidant enzymes and detoxification metabolic system in different concentrations of diclofenac.The expressions of Cu/Zn SOD and CAT genes are more sensitive to diclofenac and can reflect the toxic effect of diclofenac to some extent,and can be used as toxicity biomarkers of diclofenac exposure.The expression of Cu/Zn SOD,CAT,MRP4 and MRP5 genes mRNA in D.magna were significantly inhibited or induced by different concentrations of simvastatin for different time,and showed obvious dose-effect and time-effect relationship,indicating that simvastatin exposure can interfere with the antioxidant enzymes detoxification metabolic functions.It also showed that simvastatin was an inducer of SOD gene in D.magna,SOD and CAT genes are more sensitive to simvastatin than MRP4 and MRP5 genes do and can reflect the toxic effects of simvastatin to a certain extent.In this study,the toxic effects of diclofenac and simvastatin on the growth and reproduction of D.magna were studied in individual level.At the same time,the effects of diclofenac and simvastatin on the antioxidant defense and detoxification-related genes expression were investigated in molecular level.The quantitative assessment of the transcriptional expression of Cu/Zn SOD,CAT,MRP4 and MRP5 genes in D.magna was established.This study provides a scientific basis for evaluating the ecological risk assessment of diclofenac and simvastatin to aquatic organisms,especially aquatic invertebrate crustaceans.