Metabolism And Residue Elimination Of Mequindox On Sea Cucumbers And Abalones

Studies have shown that toxic effect of Mequindox is closely related to its residual metabolites.Sea cucumbers and abalones,abundant in Yantai of Shandong Province,were used as objects in this paper.This study analyzed and identified metabolites of Mequindox in sea cucumbers and abalones,explored the metabolic pathways of Mequindox,and further elaborated the possible relationship between its structure and metabolism,in order to provide data support and reference for the determination of residual targets of Mequindox in aquatic animals and the formulation of resting period.The specific research contents are as follows:(1)The samples of sea cucumber and abalone were cultured in water bath with Mequindox at a concentration of 4 mg/L.Three different methods were used to extract metabolites of Mequindox from each sample,and the optimal extraction method was determined by cpmparing the peak shape and response value of the chromatographic peak of the metabolites.In the formal experiment,using organic reagents(methanol and ethyl acetate containing 0.1% formic acid),metabolites of Mequindox in sea cucumbers and abalones were extracted and purified.(2)UHPLC-Q Exactive Plus high resolution mass spectrometry was used to detect and identify the metabolites of Mequindox in sea cucumbers.Based on the above extraction and detection methods,nine metabolites of Mequindox were detected and identified in the body of sea cucumbersin this research.The specific structure of each metabolite was inferred from the secondary fragment ions of detected metabolites,named MM1-MM9.Nine metabolites were α,3-dimethyl-1,4-dioxygen-quinoxaline-2-methanol,2-ethanoyl-3-methyl-4-oxygenquinoxaline,2-ethanoyl-3-methyl 1-oxygen quinoxaline,α,3-dimethyl-1-dioxygen-quinoxaline-2-methanol,2-ethanoyl-3-methyl quinoxaline,α,3-dimethyl quinoxaline-2-methanol,3-methyl quinoxaline-2-carboxylic acid,1,4-dioxygen quinoxaline-3-methyl and4-oxygen quinoxaline-3-methyl.Different from the previous analysis and identification of Mequindox and its metabolites in mammals,there weren’t metabolites with hydroxylation reaction in sea cucumbers.And the metabolic species differences in Mequindox can be confirmed.In order to investigate the elimination of metabolites of Mequindox in sea cucumbers,the samples were divided into five experimental groups.All of them were cultured in water bath for 12 h,after which they were incubated with fresh seawater 6h,12 h,24 h,48 h and 72 h,respectively.The results showed that metabolite MM6 was relatively stable structures in sea cucumbers.72 h after the end of drug feeding,the residul amount of metabolites of Mequindox was pretty low,and even some metabolites were not detected in the tissue.MQCA(MM7),a metabolite that was previously shown to be linearly correlated with the total metabolites of Mequindox residue,was barely detectable at 72 h after the end of drug feeding.In summary,the main metabolic pathway of Mequindox in the sea cucumbers is the reduction of N→O group and carbonyl group.Mequindox and its metabolites can be eliminated quickly in sea cucumbers,and the relevant studies can provide a basis for the formulation of Mequindox in the rest period of sea cucumbers.(3)The same method was used to analyze and identify the abalone samples,and7 metabolites,named PM1-PM7,were detected.Among them,MM1 and PM1,MM3 and PM3,MM6 and PM6,MM9 and PM7 are of the same lineage.In addition,the hydroxyl substitution product of Mequindox(PM2)was found in abalones.Compared with metabolites in sea cucumbers,the response values of metabolites of Mequindox in abalone were higher.Data showed that the most of metabolites of Mequindox could be metabolized out in 48 h after the end of drug feeding.Metabolites involving N→O group reduction accounted for 54.6% of the total metabolites content(in terms of response values),carbonyl reduction accounted for 90.3%,aceyl reduction and hydroxyl substitution accounted for 0.3% and 3.1%,respectively.Date showed that carbonyl reduction and N → O group reduction were the most important metabolic pathways in abalones.