Determination Of Three Tetracyclines Antibiotics And Their Metabolites In Pork Tissues And Organs By DLLME

Dispersive liquid liquid microextraction(DLLME)is a sample pretreatment technique which has developed rapidly in recent years.It has many advantages such as low pollution,low cost,high efficiency,high accuracy,simple and convenient operation and easy to combine with various detectors,etc.It is widely used in the separation analysis of chemistry,biology,medicine,environment,food and other fields.In this paper,the extraction of three kinds of tetracycline antibiotics(TC?OTC?CTC)and their metabolites(ETC?EOTC?ECTC)were studied by using two kinds of DLLME model(IL-DLLME and SO-DLLME).By comparison,selecting the best extraction model combined with HPLC to determine different pork tissues and organs.The migration of six components in tissues and organs was studied.In addition,the method of HPLC was used to detect and analysis three kinds of antibiotics(TC ?OTC?CTC)in kidney in different storage conditions.The main research contents are as follows:1.The method for the extraction of three kinds of TCs and its metablities in the sample solution by IL-DLLME was established.The optimum extraction conditions were obtained by choosing the type and the volume of extrant and dispersant,the extraction time and the salt concentration.Under the optimum conditions,two kinds of mixed standard solution with concentration of 10 mg·L-1(CTC and ECTC wre 20mg·L-1)were extracted by IL-DLLME and detected by HPLC.After calculation,the enrichment of the three TCs were: TC 10.2 was times,OTC 3.9 was times,CTC was24.3 times.The enrichment of the metabolities of three TCs were: ETC was 18.7times,EOTC was 12.5 times,ECTC was 25.2 times.2.The method for the extraction of three kinds of TCs and its metablities in the sample solution by SO-DLLME was established.The optimum extraction conditions were obtained by choosing the type of solvent,the salt concentration,the volume of extrant and the extraction time.Under the optimum conditions,two kinds of mixed standard solution with concentration of 10 mg·L-1(CTC and ECTC wre 20 mg·L-1)were extracted by SO-DLLME and detected by HPLC.After calculation,the enrichment of the three TCs were: TC was 7.3 times,OTC was 16.8 times,CTC was2.2 times.The enrichment of the metabolities of three TCs were: ETC was 21.1 times,EOTC was 6.9 times,ECTC was 5.9 times.3.The method of IL-DLLME-HPLC was adopted to determine the three kinds of TCs and its metabolitis.Extracted the six components under the optimized IL-DLLME conditions,and detected by HPLC.The results showed that there was a good linear relationship between the six components at the concentraction of 0.02~2mg·L-1(CTC and ECTC were 0.04~4 mg·L-1),the correlation coefficients were greater than 0.99;The recovery of spiked samples were between 79.2~107.5%;The limit of detection(LOD)were between 0.013~0.023 mg·L-1;The relative standard deviation were less than 5%(n=11).This method can be used to separate the six very similar components,and has high precision and low detection limit.The study of the six components in pork tissues and organs can provide reference for the pharmacokinetics of TCs in animals.4.The method of HPLC was used to study the degradation of OTC?TC?CTC in kidney sample in different storage conditions(different storage time,temperature and adding different preservatives).The results showed that:(1)In the same storage temperature,with storage time increase,the degradation of OTC?TC?CTC in kidney sample were higher.(2)In the same storage time,with the increase of temperature,the residual rate of porcine of OTC?TC?CTC in kidney samples were gradually reduced,and the degradation were higher and higher.(3)The degradation of OTC?TC?CTC in kidney sample in the storage conditions of adding preservatives of benzoic acic,sorbic acid,sodium nitrite were lower than adding nothing·(4)Under the refrigerated storage conditions,the degradation of the sample which had added sodium nitrite and sorbic acid were lower than added benzoic acic.