在线固相萃取-液相色谱-串联质谱法测定血液和尿液中31种有毒植物化学组分的含量
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摘要
取血液或尿液样品0.20mL,加入乙腈0.60mL,涡旋振荡1min,并离心10min,取上清液0.40mL,加入10mol·L~(-1)乙酸铵溶液(pH 7.0)1.1mL,振荡1min,用0.22μm滤膜过滤后,溶液经HLB在线固相萃取柱富集纯化,所用流动相A为水,B为甲醇,C为乙腈。所得淋出液经Poroshell 120EC-C18色谱柱并用流动相A 10mol·L~(-1)乙酸铵溶液和流动相B乙腈(上述二流动相中均含φ0.1%甲酸)进行梯度洗脱,使所测定的31种有毒植物的化学组分达到分离,并进行质谱测定。所测定的31种目标物的质量浓度在一定范围内与其峰面积之间呈线性关系,其检出限(3S/N)在0.01～5.00μg·L~(-1)之间。分别以上述两种样品为基体,用标准加入法进行回收试验,测得31种化合物在血液样品中的回收率在90.4%～107%之间,在尿液样品中的回收率在90.0%～103%之间。测定值的相对标准偏差(n=6)依次在0.2%～2.6%,0.3%～2.9%之间。
0.20 mL of the sample of blood or urine was taken and shaked vortically for 1 min with 0.60 mL of acetonitrile,and then centrifuged for 10 min.A portion(0.40 mL)of the supernatant was taken and diluted with1.1 mL of 10 mol·L~(-1) CH_3COONH_4 solution(pH 7.0);after shaking for 1 min,the solution was filtered through0.22μm filtering membrane and the filtrate was passed through the HLB online solid phase extraction column for enriching and purifying the 31 analytes(i.e.,the constituents of poisonous plants)by elution with mobile phases of(A)water,(B)methanol and(C)acetonitrile.The eluates obtained were then separated by gradient elution using Poroshell 120 EC-C18 column as stationary phase and(A)10 mol·L~(-1) CH_3COONH_4 solution,(B)acetonitrile as mobile phases,both of which contained 0.1%(φ)of formic acid.The 31 analytes in the eluates were then determined by tandem MS.Linear relationships were obtained between values of peak areas of the 31 analytes and their mass concentrations in definite ranges,with detection limits(3 S/N)in the range of 0.01-5.00μg·L~(-1).Test for recovery was made by standard addition on the base of blood sample or urine sample as matrixes separately,giving results of recovery of the 31 compounds in the range of 90.4%-107% for blood sample and of 90.0%-103%for urine sample.Values of RSDs(n=6)ranged from 0.2% to 2.6% and from 0.3% to 2.9% for the 2 kinds of samples respectively.
0.20 mL of the sample of blood or urine was taken and shaked vortically for 1 min with 0.60 mL of acetonitrile,and then centrifuged for 10 min.A portion(0.40 mL)of the supernatant was taken and diluted with1.1 mL of 10 mol·L~(-1) CH_3COONH_4 solution(pH 7.0);after shaking for 1 min,the solution was filtered through0.22μm filtering membrane and the filtrate was passed through the HLB online solid phase extraction column for enriching and purifying the 31 analytes(i.e.,the constituents of poisonous plants)by elution with mobile phases of(A)water,(B)methanol and(C)acetonitrile.The eluates obtained were then separated by gradient elution using Poroshell 120 EC-C18 column as stationary phase and(A)10 mol·L~(-1) CH_3COONH_4 solution,(B)acetonitrile as mobile phases,both of which contained 0.1%(φ)of formic acid.The 31 analytes in the eluates were then determined by tandem MS.Linear relationships were obtained between values of peak areas of the 31 analytes and their mass concentrations in definite ranges,with detection limits(3 S/N)in the range of 0.01-5.00μg·L~(-1).Test for recovery was made by standard addition on the base of blood sample or urine sample as matrixes separately,giving results of recovery of the 31 compounds in the range of 90.4%-107% for blood sample and of 90.0%-103%for urine sample.Values of RSDs(n=6)ranged from 0.2% to 2.6% and from 0.3% to 2.9% for the 2 kinds of samples respectively.
引文
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[10]应剑波,王芳琳,栾玉静,等.固相支撑液液萃取-高效液相色谱-串联质谱法测定血中欧夹竹桃苷和欧夹竹桃苷乙的含量[J].理化检验-化学分册,2017,53(2):125-129.
[11]USUI K,HAYASHIZAKI Y,HASHIYADA M,et al.Simultaneous determination of 11aconitum alkaloids in human serum and urine using liquid chromatography-tandem mass spectrometry[J].Legal Medicine,2012,14(3):126-133.
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[13]PIETSCH J,GNTHER J,HENLE T,et al.Simultaneous determination of thirteen plant alkaloids in a human specimen by SPE and HPLC[J].Journal of Separation Science,2008,31(13):2410-2416.
[14]YU Z H,WU Z P,GONG F J,et al.Simultaneous determination of six toxic alkaloids in human plasma and urine using capillary zone electrophoresis coupled to time-of-flight mass spectrometry[J].Journal of Separation Science,2012,35(20):2773-2780.
[15]冯祖飞,赵颖娟,昌征.磁性修饰羟基化多壁碳纳米管萃取人血浆中马钱子生物碱的方法研究[J].分析试验室,2017,36(4):379-383.
[16]朱定姬,卢敏萍,黄克建,等.在线固相萃取结合液相色谱-线性离子阱多级质谱法同时检测生物样品中7种乌头类生物碱[J].色谱,2016,34(3):249-257.
[17]ZHANG N,LI J,SONG Q Q,et al.Direct stability characterization of aconite alkaloids in different media by autosampler-mediated incubation-online solid phase extraction-LC-MS/MS[J].Analytical Methods,2016(8):1942-1949.
[18]刘晓锋,黄克建,林翠梧,等.在线固相萃取-液相色谱-质谱法测定血液中9种局部麻醉药[J].理化检验-化学分册,2015,51(2):245-247.
[19]黄克建,卢敏萍,周哲,等.在线固相萃取/液相色谱-线性离子阱质谱法同时定性定量检测全血和尿液中7种抗凝血杀鼠药[J].色谱,2015,33(7):691-698.
[20]卢敏萍,黄克建,周哲,等.在线固相萃取/液相色谱-线性离子阱质谱法同时检测生物样品中18种氨基甲酸酯类农药[J].分析测试学报,2016,35(7):777-784.
[2]吴惠勤,张春华,黄晓兰,等.气相色谱-串联质谱法同时检测尿液中15种有毒生物碱[J].分析测试学报,2013,32(9):1031-1037.
[3]CHENG M,LIU R,WU Y,et al.LC-MS/MS determination and urinary excretion study of seven alkaloids in healthy Chinese volunteers after oral administration of Shuanghua Baihe tablets[J].Journal of Pharmaceutical and Biomedical Analysis,2016,118:89-95.
[4]WU Y,LIU R J,GU P,et al.Highly sensitive method for simultaneous determination of nine alkaloids of Shuanghua Baihe tablets in human plasma by LC-MS/MS and its application[J].Journal of Chromatography B,2015,1007:81-92.
[5]WU H Q,XIONG X T,HUANG X L,et al.Simultaneous determination of 17toxic alkaloids in human fluids by liquid chromatography couples with electrospray ionization tandem mass spectrometry[J].Journal of Liquid Chromatography&Related Technologies,2013,36(9):1149-1162.
[6]ZHAI J X,SHEN M,LIU W.Simultaneous screening for 45poisonous alkaloids in blood by LC-MS/MS[J].Journal of Forensic Medicine,2015,31(1):28-33.
[7]刘伟,沈敏,沈保华,等.LC-MS/MS-MRM筛选血液中22种有毒生物碱成分[J].法医学杂志,2007,23(5):349-352.
[8]张春华,吴惠勤,黄晓兰,等.液相色谱-电喷雾串联质谱同时检测尿液和胃液中12种有毒生物碱[J].分析化学,2012,40(6):862-869.
[9]张盼盼,张福成,王朝虹,等.超高效液相色谱-串联质谱法同时测定尿样中4种痕量的乌头类生物碱[J].色谱,2013,31(3):211-217.
[10]应剑波,王芳琳,栾玉静,等.固相支撑液液萃取-高效液相色谱-串联质谱法测定血中欧夹竹桃苷和欧夹竹桃苷乙的含量[J].理化检验-化学分册,2017,53(2):125-129.
[11]USUI K,HAYASHIZAKI Y,HASHIYADA M,et al.Simultaneous determination of 11aconitum alkaloids in human serum and urine using liquid chromatography-tandem mass spectrometry[J].Legal Medicine,2012,14(3):126-133.
[12]BEYER J,PETERS F T,KRAEMER T,et al.Detection and validated quantification of toxic alkaloids in human blood plasma-comparison of LC-APCI-MSwith LC-ESI-MS/MS[J].Journal of Mass Spectrometry,2007,42(5):621-633.
[13]PIETSCH J,GNTHER J,HENLE T,et al.Simultaneous determination of thirteen plant alkaloids in a human specimen by SPE and HPLC[J].Journal of Separation Science,2008,31(13):2410-2416.
[14]YU Z H,WU Z P,GONG F J,et al.Simultaneous determination of six toxic alkaloids in human plasma and urine using capillary zone electrophoresis coupled to time-of-flight mass spectrometry[J].Journal of Separation Science,2012,35(20):2773-2780.
[15]冯祖飞,赵颖娟,昌征.磁性修饰羟基化多壁碳纳米管萃取人血浆中马钱子生物碱的方法研究[J].分析试验室,2017,36(4):379-383.
[16]朱定姬,卢敏萍,黄克建,等.在线固相萃取结合液相色谱-线性离子阱多级质谱法同时检测生物样品中7种乌头类生物碱[J].色谱,2016,34(3):249-257.
[17]ZHANG N,LI J,SONG Q Q,et al.Direct stability characterization of aconite alkaloids in different media by autosampler-mediated incubation-online solid phase extraction-LC-MS/MS[J].Analytical Methods,2016(8):1942-1949.
[18]刘晓锋,黄克建,林翠梧,等.在线固相萃取-液相色谱-质谱法测定血液中9种局部麻醉药[J].理化检验-化学分册,2015,51(2):245-247.
[19]黄克建,卢敏萍,周哲,等.在线固相萃取/液相色谱-线性离子阱质谱法同时定性定量检测全血和尿液中7种抗凝血杀鼠药[J].色谱,2015,33(7):691-698.
[20]卢敏萍,黄克建,周哲,等.在线固相萃取/液相色谱-线性离子阱质谱法同时检测生物样品中18种氨基甲酸酯类农药[J].分析测试学报,2016,35(7):777-784.