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目的建立一种快速、灵敏的同时测定人乳中多种B族维生素的超高压液相色谱-串联质谱法。方法样品经甲醇沉淀蛋白、乙醚去除非极性杂质,UPLC HSS T3色谱柱分离后采用保留时间和多反应离子检测(MRM)定性、定量,同位素内标校正。结果维生素B1(硫胺素)、维生素B2(核黄素)和维生素B6(吡哆醛)的定量下限(LOQ)均为0.2μg/100 g,维生素B3(烟酰胺)和维生素B5(泛酸)的LOQ均为0.3μg/100 g。标准加入法测定所有被测物的高、中、低三个浓度加标水平的回收率在93.1%~101.1%之间,相对标准偏差(RSD)在2.8%~6.2%之间。5种水溶性维生素在各自浓度范围内线性良好,R2均大于0.998。结论本文建立了同时测定人乳中5种B族水溶性维生素的超高压液相色谱-串联质谱法。方法前处理简单、快速,样品需要量少;采用质谱检测器同位素内标一对一校正定量,检测结果准确、灵敏。方法可以为人乳中维生素含量的检测、调查、研究提供技术支持。
Objective To establish a rapid and sensitive method for the simultaneous determination of B vitamins in human milk by ultra-high performance liquid chromatography-tandem mass spectrometry. Methods The samples were precipitated with methanol and ether to remove nonpolar impurities. After UPLC HSS T3 column was separated, the retention time and MRM were used for qualitative, quantitative and isotopic calibration. Results The LOQ values of vitamin B1 (thiamine), vitamin B2 (riboflavin) and vitamin B6 (pyridoxal) were 0.2 μg / 100 g, and vitamin B3 (niacinamide) and vitamin B5 The LOQs were 0.3 μg / 100 g. The standard addition method was used to determine the recoveries of all analytes at the spiked levels of 93.1% -101.1%, and the relative standard deviations (RSDs) ranged from 2.8% to 6.2%. The five water-soluble vitamins had good linearity in their respective concentrations, with R2 greater than 0.998. Conclusion This article established a simultaneous determination of five kinds of water-soluble vitamins in human milk by ultra-high performance liquid chromatography - tandem mass spectrometry. The pretreatment method is simple, rapid, and requires less sample. The mass spectrometry detector is used to calibrate the internal standard one by one. The detection results are accurate and sensitive. The method can provide technical support for the detection, investigation and research of vitamin content in human milk.