| Meloxicam is a new kind of non-steroidal anti-inflammatory drugs,from the enol acid derivatives.It is widely used in veterinary clinical medicine because it has better curative effect on anti-inflammatory,analgesic,antipyresis and higher gastrointestinal tolerance.In recent years,because the animal husbandry tends to intensive cultivation and large-scale operations,and the phenomenon of abusing the veterinary drugs and breaching the withdrawal time hasexisted,which causes the problem of animal food safety.This study developed a method for the confirmation and quantification of meloxicamresiduein edible tissues(includingmuscle,liver,kidneyand milk)of porcine and bovine byliquid chromatography tandem mass spectrometry(LC-MS/MS)in order to provide technical detection of meloxicam residues in animal-derived food.Two methodswere used to pretreat samples,because the tremendous differences between the tissue and milk in the components and structures.The samples of liver,kidney and muscle which containing meloxicam and meloxicam-d3 were extracted by acetonitrile-methanol(95:5,V/V)solution,then the supernatant was complicated with a constant volume and placed in the freezer(-20?)for 60 min.The upper of the extract(6 m L)was moved and evaporated to dryness in the nitrogen stream,reconstituted in 2 m L of acetonitrile-water(1:1,V/V)solution and stirred vigorously.The sample solution was defatted by n-hexane,and again after high-speed centrifuge to remove n-hexane.The lower phase was transferred and filtrated by 0.22 ?m filterable membrane.The samples of milk which containing meloxicam and meloxicam-d3 were extracted by methanol,after mechanical rotary shaker and stirred vigorously.Then the supernatant was complicated with a constant volume and the extract(5 m L)was moved and evaporated to dryness in the nitrogen stream,reconstituted in 1 m L of 0.1% formic acid in acetonitrile-methanol(90:10,V/V)solution,high-speed centrifuge to remove impurities,following filtrated by 0.22 ?m filterable membrane.The filtrates were analyzed by LC-MS/MS under a gradient elution program with the mobile phase of 0.1% formic acid solution and acetonitrile.In this experiment,the limit of detection(LOD)in the muscle,liver and kidney of porcine was 2.5 ?g/kg,5.0 ?g/kg and 5.0 ?g/kg,respectively.The LOD in the muscle,liver and kidney of bovine was 2.5 ?g/kg,5.0 ?g/kg and 5.0 ?g/kg,respectively.The LOD in milk was 1.0 ?g/kg.The limit of quantification(LOQ)in the muscle,liver and kidney of porcine was 5.0 ?g/kg,10.0 ?g/kg and 10.0 ?g/kg,respectively.The LOQ in the muscle,liver and kidney of bovine was 5.0 ?g/kg,10.0 ?g/kg and 10.0 ?g/kg,respectively.The LOQ in milk was 1.5 ?g/kg.The linear range of meloxicam was 1.0~60.0 ?g/kg,100.0~200.0 ?g/kg and 100.0~200.0 ?g/kg,respectively in the muscle,liver,and kidney tissues of porcine,the correlation coefficient was more than 0.998 and the linearity was good.The linear range of meloxicam was 1.0~60.0 ?g/kg,100.0~200.0 ?g/kg,100.0~200.0 ?g/kg and 1.5~45.0 ?g/kg,respectively in the muscle,liver,and kidney tissues of bovine,the correlation coefficient was more than 0.997 and the linearity was good.Blank samples(including liver,kidney,muscle and milk)were spiked with meloxicam at concentrations corresponding to LOQ,0.5MRL,1.0MRL and 2.0MRL,in order to investigate parameters such as repeatability and reproducibility.The average recov ery within run of meloxicam in the porcine tissue(muscle,liver,kidney)was92.00%~109.81%,101.96%~108.13% and 94.04%~110.75%,respectively.The coefficient of variation(C V)within run was2.79%~7.59%,1.60%~4.09% and 0.98%~4.93%,respectively.The CV between run was 4.96%~7.28%,2.71%~4.24% and 3.78~7.37%,respectively.The average recovery within run of meloxicam in the bovine tissue(muscle,liver,kidney,and milk)was94.95%~111.36%,100.28%~120.33%,86.78%~116.07% and 98.40%~ 115.20%,respectively.The CV within run was 1.67%~7.11%,1.16%~4.81%,0.39%~3.87% and 0.78%~3.90%,respectively.The CV between run was4.24%~6.28%,3.24%~6.85%,7.86%~12.34% and 2.87%~5.41%,respectively.The results showed that the LC-MS/MS method for the confirmation and quantification of meloxicam residuein edible tissues of porcine and bovine had good reproducibility and high sensitivity,that meets the demands of the analysis of veterinary drug residues. |
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