固相萃取-超高效液相色谱-串联质谱法同时测定人尿液中40种抗生素的残留量  

Simultaneous Determination of Residues of 40Antibiotics in Human Urine by Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry with Solid Phase Extraction

作　　者：徐新 孙文芳[1] 徐杰 霍翔[2] 刘祥萍[1] XU Xin;SUN Wenfang;XU Jie;HUO Xiang;LIU Xiangping(Department of Physical and Chemical Analysis,Nanjing Center for Disease Control and Prevention,Nanjing210003,China;Jiangsu Provincial Center for Disease Control and Prevention,Nanjing210009,China)

机构地区：[1]南京市疾病预防控制中心理化检验科,南京210003 [2]江苏省疾病预防控制中心,南京210009

出　　处：《理化检验（化学分册）》2023年第5期591-598,共8页Physical Testing and Chemical Analysis(Part B:Chemical Analysis)

基　　金：江苏省卫生健康委科研项目(M2021078)。

摘　　要：取人尿液样品1.00 mL,加入1.0 mg·L^(-1) 13C-红霉素-d3和阿莫西林-d4的混合内标溶液10μL和乙二胺四乙酸二钠(Na2EDTA)-Mcllvaine缓冲液(pH 3.5±0.05) 0.30 mL,涡旋30 s,离心5 min,将上清液以流量1.0 mL·min^(-1)过活化好的Oasis HLB固相萃取柱,抽干柱子后用甲醇3 mL以1.0 mL·min^(-1)流量洗脱柱子,收集全部洗脱液,氮吹至近干,用10%(体积分数)甲醇溶液稀释至0.5 mL,涡旋混匀后过0.22μm有机滤膜,滤液供超高效液相色谱-串联质谱仪分析。40种抗生素(8种磺胺类、6种大环内酯类、8种β-内酰胺类、4种四环素类、9种喹诺酮类、3种氯霉素类、1种林可酰胺类和1种喹喔啉类抗生素)在Kinetex?F5 100A色谱柱(50 mm×3.0 mm, 2.6μm)上用不同体积比的0.1%甲酸(体积分数,下同)溶液和含0.1%甲酸的乙腈溶液的混合溶液进行梯度洗脱分离,用电喷雾离子源负(3种氯霉素类抗生素)、正离子模式电离,用多反应监测(MRM)模式检测,其中红霉素和阿莫西林采用内标法和基质匹配法定量,其他抗生素采用基质匹配法定量。结果表明,红霉素和青霉素V工作曲线的线性范围为1.0~100.0μg·L^(-1),其余38种抗生素工作曲线的线性范围为1.0~200.0μg·L^(-1),检出限(3S/N)为0.01~1.25μg·L^(-1);按标准加入法进行回收试验,回收率为51.0%~128%,测定值的相对标准偏差(n=5)为1.2%~14%。方法用于10份成年人尿液样品的分析,其中9份样品中检出了9种抗生素,检出量为0.09~322.86μg·L^(-1),其中阿莫西林检出率高达50%,检出量高达322.86μg·L^(-1)。An aliquot(1.00mL)of human urine sample was taken,10μL of 1.0mg·L^(-1) mixed internal standard solution(containing 13 C-erythromycin-d3and amoxicillin-d4)and 0.30mL of Na2EDTA-Mcllvaine buffer solution(pH 3.5±0.05)were added.The mixed solution was vortexed for 30sand centrifuged for 5min.The supernatant was passed through Oasis HLB solid phase extraction column at flow rate of 1.0mL·min^(-1).The column was drained,and eluted by 3mL of methanol at flow rate of 1.0mL·min^(-1).The eluate was collected,blown to near dryness by nitrogen,and diluted to 0.5 mL by 10%(volume fraction)methanol solution.The resulting solution was vortexed and passed through a 0.22μm organic filter membrane,after which the filtrate was analyzed by ultra-high performance liquid chromatograph-tandem mass spectrometer.Forty antibiotics(8 sulfonamides,6macrolides,8β-lactams,4tetracyclines,9quinolones,3chloramphenicols,1lincosamide and 1 quinoxaline antibiotics)were separated by gradient elution with mixed solutions composed of 0.1%formic acid(volume fraction,the same below)solution and acetonitrile solution containing 0.1%formic acid at different volume ratios on KinetexⓇF5100Acolumn(50mm×3.0mm,2.6μm),ionized by negative(3chloramphenicol antibiotics)and positive ion mode of electrospray ion source,and detected by MRM mode.Erythromycin and amoxicillin were quantified by internal standard and matrix matching method,and other antibiotics were quantified by matrix matching method.As shown by the results,linear ranges of working curves of erythromycin and penicillin V were 1.0-100.0μg·L^(-1),and linear ranges of working curves of the remaining 38antibiotics were 1.0-200.0μg·L^(-1),with detection limits(3S/N)in the range of 0.01-1.25μg·L^(-1).Test for recovery was made by standard addition method,giving recoveries in the range of 51.0%-128%,and RSDs(n=5)of the determined values ranged from 1.2%to 14%.The proposed method was applied to the analysis of 10adult urine samples,and 9antibiotics were detected in 9 samples,with detection amo

关 键 词：固相萃取 超高效液相色谱-串联质谱法(UHPLC-MS/MS) 抗生素 残留量 人尿液 

分 类 号：O657.63[理学—分析化学]