Determination of Florfenicol Enantiomers and Their Metabolite Florfenicol Amine in Pork by Ultra-Performance Convergence Chromatography with Solid Phase Extraction
-
摘要: 在10.0 g猪肉样品中加入含3%(体积分数)氨水的乙酸乙酯溶液25 mL,涡旋1 min混匀,离心5 min,所得沉淀用上述方法再重复提取一次。合并两次提取液,于40℃减压浓缩至近干,用10 mL磷酸盐缓冲溶液(pH 6.0)溶解残渣。所得溶液过活化好的HLB固相萃取柱,用10 mL水淋洗,真空抽柱1 min,用6 mL甲醇洗脱。收集洗脱液,于40℃氮吹至近干,加入体积比8∶2的正庚烷-异丙醇混合溶液1 mL,涡旋3 min溶解残渣,过0.22 μm有机滤膜,滤液供超高效合相色谱仪分析。采用CHIRALPAK AD-3手性色谱柱作固定相,设置柱温40℃,系统背压13.8 MPa,以不同体积比的超临界二氧化碳和含0.5%(体积分数)氨水的甲醇溶液的混合溶液为流动相进行梯度洗脱,在检测波长224 nm处用外标法测定(-)-氟苯尼考、(+)-氟苯尼考及它们的代谢物氟苯尼考胺的含量。结果显示:3种目标化合物的质量浓度在0.50~20.00 mg·L-1内与其对应的峰面积呈线性关系,测定下限为0.05 mg·kg-1[(-)-氟苯尼考和(+)-氟苯尼考]和0.1 mg·kg-1(氟苯尼考胺)。以阴性猪肉样品为基质进行3个浓度水平的加标回收试验,3种目标化合物的回收率为81.2%~107%,测定值的相对标准偏差(n=6)为5.0%~9.0%。方法用于20份猪肉样品的分析,仅在1份样品中检出(-)-氟苯尼考(248 μg·kg-1)。Abstract: Ethyl acetate solution (25 mL) containing 3% (φ) aqueous ammonia was added into 10.0 g of pork sample, the mixture was vortexed for 1 min, centrifuged for 5 min, and the precipitation was extracted once again as the method described above. The two extracts were combined and evaporated to nearly dry under vacuum at 40℃, and the residue was redissolved with 10 mL of phosphate buffer solution (pH 6.0). The HLB solid phase column activated beforehand was used for loading the solution, washed with 10 mL of water, drained by vacuum for 1 min, and eluted with 6 mL of methanol. The eluate was collected and dried by nitrogen at 40℃ to nearly dry. The residue was dissolved by vortex for 3 min with 1 mL of mixed solution of n-heptane and isopropanol at volume ratio of 8:2, filtered through a 0.22 μm filter membrane and the filtrate was analyzed by ultra-performance convergence chromatography. CHIRALPAK AD-3 chiral column was used as the stationary phase with column temperature of 40℃ and the system back pressure of 13.8 MPa. The mixture of supercritical carbon dioxide and methanol solution containing 0.5% (φ) aqueous ammonia at different volume ratios was used as the mobile phase for gradient elution. (-)-Florfenicol, (+)-florfenicol and their metabolite florfenicol amine were determined by external standard method at wavelength of 224 nm. It was shown that the mass concentrations of the 3 targets were linearly related with their corresponding peak areas in the range of 0.50-20.00 mg·L-1, and the lower limits of determination were 0.05 mg·kg-1[for (-)-florfenicol and (+)-florfenicol] and 0.1 mg·kg-1 (for florfenicol amine). Test for the spiked recovery was made on the negative pork samples at the 3 concentration levels, giving recoveries of the 3 targets in the range of 81.2%-107%, and RSDs (n=6) of the determined values ranged from 5.0% to 9.0%. The proposed method was used for the analysis of 20 pork samples, and (-)-florfenicol (248 μg·kg-1) was detected in one sample.
-
-
[1] SHIROMA L S, QUEIROZ S C N, JONSSON C M, et al. Extraction strategies for simultaneous determination of florfenicol and florfenicol amine in tilapia (oreochromis niloticus) muscle:Quantification by LC-MS/MS[J]. Food Analytical Methods, 2020,13(1):291-302.
[2] 李广.氟苯尼考及氟苯尼考胺药物残留检测方法及应用[J].辽宁农业科学, 2019(5):59-62. [3] 李涛,杨伟,杨丽芬,等.高效液相色谱-紫外法检测畜禽肉中氟苯尼考残留[J].现代食品, 2020(9):130-132. [4] 李然.兽药氟苯尼考及其代谢物与甲砜霉素残留免疫分析方法的研究[D].广州:华南农业大学, 2018. [5] 文丽,吴泽华,李仕超.氟苯尼考手性特性及在临床上的应用[J].猪业科学, 2016,33(8):70-71. [6] 肖建森,刘在青,宋凤香,等.一种动物专用复方左旋氟苯尼考注射液及其制备方法:201310322969.6[P].(2014.01.15)[2021.11.14]. [7] 赵梅仙,张传良,马佳颖,等.氟苯尼考对映异构体手性拆分及其光学纯度的测定[J].分析测试学报, 2013,32(4):459-462. [8] 吴东林,王铭宏.一种测定氟苯尼考中手性活性成分含量的方法:201310244316.0[P].(2013.10.02)[2021.07.14]. [9] 中华人民共和国农业农村部,中华人民共和国国家卫生健康委员会,国家市场监督管理总局.食品安全国家标准食品中兽药最大残留限量:GB 31650-2019[S].北京:中国标准出版社, 2019. [10] 何红梅,徐秀珠,潘春秀,等.氯霉素类药物的高效液相色谱手性分离研究[J].分析化学, 2005,33(2):165-168. [11] RAJ T J S, SRINIVAS N, PRASAD C H S, et al. Validated chiral liquid chromatographic method for the enantiomeric separation of florfenicol[J]. Journal of Liquid Chromatography&Related Technologies, 2007,31(2):231-239.
[12] 王宇,王炎,叶晶晶,等.氟苯尼考光学异构体的分离测定方法研究[J].化工管理, 2021(24):136-137. [13] 张文华,谢文,侯建波,等.超高效合相色谱法对6种三唑类农药对映体的拆分及其在黄瓜中的残留分析[J].色谱, 2019,37(12):1356-1362. [14] JIANG H, YANG L, XING X D, et al. Development of an analytical method for separation of phenolic acids by ultra-performance convergence chromatography (UPC2) using a column packed with a sub-2-μm particle[J]. Journal of Pharmaceutical and Biomedical Analysis, 2018,153:117-125.
[15] YU W S, LIU X, ZHANG Y Z, et al. Simultaneous determination of pigments in tea by ultra-performance convergence chromatography (UPC2)[J]. Analytical Letters, 2020,53(10):1654-1666.
[16] CHANG X Q, SUN P, MA Y, et al. A new method for determination of thymol and carvacrol in thymi herba by ultraperformance convergence chromatography (UPC2)[J]. Molecules (Basel, Switzerland), 2020,25(3):502-514.
[17] LUCKEY B, MEKILE V. Vitamin A analysis by ultra-performance convergence chromatography[J]. Journal of AOAC International, 2019,102(2):659-661.
[18] 宋菲,黄瑶,张利明,等.猪肉中氟苯尼考残留量高效液相色谱-串联质谱检测方法的建立[J].国外畜牧学(猪与禽), 2021,41(2):89-93. [19] 刁志祥,张培杨,王旭堂,等.加速溶剂萃取/超高效液相色谱-荧光法检测猪肉中氟苯尼考和氟苯尼考胺残留[J].中国畜牧兽医, 2019,46(9):2753-2759. [20] YANG J J, SUN G Z, QIAN M R, et al. Development of a high-performance liquid chromatography method for the determination of florfenicol in animal feedstuffs[J]. Journal of Chromatography B, 2017,1068/1069:9-14.
[21] 张守明.高效液相色谱法检测猪肉中氟苯尼考及其代谢物氟苯尼考胺的残留[D].长春:吉林大学, 2010. [22] 刘柏林,谢继安,赵紫微,等.固相萃取-超高效液相色谱-串联质谱法同时测定动物源性食品中11种禁限兽药及代谢物[J].色谱, 2021,39(4):406-414. [23] LI J, GONG J Y, YUAN H N, et al. Determination of chloramphenicol,thiamphenicol and florfenicol in Chinese gelatin medicines using dispersive solid-phase extraction coupled with ultra high-performance liquid chromatography-mass spectrometry[J]. Journal of Chromatographic Science, 2020,58(5):471-476.
[24] RASHEED C M, FAKHRE N A. Derivative spectrophotometric determination of florfenicol in chicken samples[J]. Zanco Journal of Pure and Applied Sciences, 2018, 29(6):49-58.
[25] 国家药典委员会.中华人民共和国药典2020年版[M].北京:中国医药科技出版社, 2020. [26] 国家质量监督检验检疫总局,中国国家标准化管理委员会.实验室质量控制规范食品理化检测:GB/T 27404-2008[S].北京:中国标准出版社, 2008.
计量
- 文章访问数: 14
- HTML全文浏览量: 1
- PDF下载量: 0
下载:
