反相高效液相色谱-串联质谱法测定母乳及奶粉中核苷和核苷酸的含量  

Determination of Nucleosides and Nucleotides in Breast Milk and Infant Formula by Reversed Phase Liquid Chromatography-Tandem Mass Spectrometry

作　　者：张婷婷 曾明飞 金燕 刘绿叶 

机构地区：[1]赛默飞世尔科技有限公司上海应用实验室,上海201203

出　　处：《分析化学》2014年第4期585-591,共7页Chinese Journal of Analytical Chemistry

摘　　要：建立了反相高效液相色谱-串联质谱法测定母乳和婴幼儿奶粉中5种核苷和5种核苷酸含量的方法.样品经温水溶解后,选择0.5％乙酸沉淀蛋白,并用超滤离心管离心过滤大分子物质;采用键合较长碳链的Acclaim C30色谱柱为分析柱,以20 mmol/L醋酸铵和乙腈为流动相,梯度洗脱分离,流速0.4 mL/min,温度15℃,紫外检测波长260 nm,采用电喷雾正离子模式进行电离,选择反应监测模式进行检测,外标法定量分析.结果表明,核苷和核苷酸在12 min内达到基线分离,在0.02～ 100 mg/L范围内,线性关系好,相关系数R＞0.999,方法检出限0.1 ～0.2 ng,其中CMP,UMP,GMP,U,AMP在1,10和50 mg/L加标水平下的平均回收率在85.2％～ 104.7％之间,峰面积RSD在0.7％～2.0％之间（n=6）,核苷C,I,G,A和核苷酸IMP在0.5,5和10 mg/L加标水平下的平均回收率在85.08％～ 103.7％之间,峰面积RSD在0.6％～1.7％之间（n=6）.应用本方法定性和定量分析母乳和婴幼儿奶粉中核苷和核苷酸的含量,并且对紫外和质谱测定的结果进行比较,基本一致.Abstract A method of reversed phase liquid chromatography-tandem mass spectrometry （HPLC-MS/MS） was established for the simultaneous determination of five kinds of Nucleotides and five kinds of Nucleosides in breast milk and infant formula. The samples were dissolved in hot water and pretreated by adding 0.5 % acetic acid to precipitate the proteins. Before injection, the samples were centrifuged with ultrafiltration centrifugal tube. The separation was performed on an Acclaim C30 column （150 mmx2.1 mm, 3 μm） with 20 mmol/Lammonium acetate and acetonitrile gradient solution as the mobile phase at a flow rate of 0.4 mL/min and 15 ℃. The detection wavelength was at 260 nm and the sample injection volume was 10 μL. The electrospray source was operated in the positive ion mode, and monitored in the selective reaction monitoring （ SRM ） mode. The concentration of nucleotides and nucleosides in the range of 0.02-100 mg/L was linearly correlated to the peak area, with correlation coefficients greater than 0. 999. The recoveries of nucleotides and nucleosides from infant formula were from 85.08% to 104.7% with the relative standard deviations （RSD） of 0.64 % - 1.96 % （ n = 6）. The proposed method was applied in the detection of Nucleotides and Nucleosides in breast milk and infant formula.

关 键 词：液相色谱 质谱 核苷 核苷酸 母乳 奶粉 

分 类 号：O657.63[理学—分析化学] TS252.7[理学—化学]