Application of UPLC-MS/MS Method for Analyzing B-vitamins in Human Milk

作     者：REN Xiang Nan YIN Shi An YANG Zhen Yu YANG Xiao Guang SHAO Bing REN Yi Ping ZHANG Jing 

作者机构：National Institute for Nutrition and HealthChinese Center for Disease Control and Prevention Beijing Key Laboratory of Diagnostic and Traceability Technologies for Food PoisoningBeijing Research Center for Preventive Medicine Zhejiang Provincial Centre for Disease Prevention and Control 

出 版 物：《Biomedical and Environmental Sciences》 (生物医学与环境科学（英文版）)

年 卷 期：2015年第28卷第10期

页      面：738-750页

核心收录：

学科分类：1010[医学-医学技术（可授医学、理学学位）] 10[医学] 

基　　金：supported by the National High Technology Research and Development Program of China(863 Program)(No.2010AA023004) 

主　　题：B-vitamins Human milk UPLC-MS/MS 

摘      要：Objective To determine ten B-vitamins in human milk by ultra-high performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS）. Methods The pretreated human milk samples were adequately separated and quantified within 11 min by UPLC-MS/MS with an Acquity UPLC HSS T3 column （2.1×100 mm, 1.8 μm）. The mobile phase was a gradient of 2.5 mmol/L ammonium formate aqueous solution and acetonitrile at a flow rate of 0.35 mL/min. Stable isotope internal standards were used in the analysis, to correct for the method variability, including matrix and ionization effects. The homogenized human milk samples were deproteinzed using methanol, unknown contaminants were extracted with diethyl ether and hydrophobic phase was discarded. The analytes were monitored via ESl＋ionization and detected in multiple reaction monitoring （MRM） with three acquisition functions. Results Calibration curves ranged from 0.5-160 ng/mL （thiamin, riboflavin, biotin, nicotinic acid, pyridoxine, pyridoxamine, pyridoxal）, and 2.5-800 ng/mL （pantothenic acid, FAD and nicotinamide） （R^2=0.990-0.999）. The relative recovery ranged from 80.1% to 120.2%; accuracy was determined to be 98.3% to 108.0%. Intra-day and inter-day variation were 3.4%-19.9% and 5.9%-18.1%, respectively. The limit of quantification （LOQ） for all vitamins was between 0.25 and 3 lag/L. Conclusion This method was successfully applied for simultaneous analysis of ten B-vitamins in human milk.