Qualitative and quantitative analysis of N-acetyllactosamine and lacto-N-biose, the two major building blocks of human milk oligosaccharides in human milk samples by high-performance liquid chromatography-tandem mass spectrometry using a porous graphitic carbon column
Qualitative and quantitative analysis of N-acetyllactosamine and lacto-N-biose, the two major building blocks of human milk oligosaccharides in human milk samples by high-performance liquid chromatography-tandem mass spectrometry using a porous graphitic carbon column
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DOI：
10.1016/j.chroma.2015.10.006
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发表时间：
2015-11-27
影响因子：
4.1
通讯作者：
Beni, Szabolcs
中科院分区：
文献类型：
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作者：
Balogh, Reka;Jankovics, Peter;Beni, Szabolcs
本研究提出了一种基于多孔石墨碳固定相的LC-MS/MS方法，用于鉴定和定量乳酸- n -二糖（LNB）和n -乙酰乳胺（LacNAc）。这些化合物是人乳低聚糖的主要组成部分，然而，到目前为止，还没有研究过人乳中是否存在它们的非结合形式。在适当的样品制备（包括排色层析和固相萃取）后，通过梯度LC法和多反应监测（MRM）分析完成了这些高度相关结构的糖醇形式的分离。组分的基线分离为该方法提供了选择性。根据欧洲药品管理局（EMA）指南进行验证，发现该方法是精确和准确的。使用我们开发和验证的方法，我们首次能够识别和量化人乳中的两种糖类。根据我们的结果，LacNAc浓度在6.7-31 μ g/mL范围内，LNB浓度从泌乳第一周的26 μ g/mL下降到低于检测限。母乳中LNB和LacNAc的存在也暗示了新的生物学功能，这可以使我们更接近于理解这种复杂生物流体的各种功能。(C) 2015 Elsevier B.V.版权所有
This study presents a validated, porous graphitic carbon stationary phase-based LC-MS/MS method for the identification and quantification of lacto-N-biose (LNB) and N-acetyllactosamine (LacNAc). These compounds are the major building blocks of human milk oligosaccharides, however the presence of their unbound form in human milk has not been examined so far. The separation of these highly related structures in their alditol form was accomplished by a gradient LC method and multiple reaction monitoring (MRM) analysis after appropriate sample preparation including size-exclusion chromatography and solid-phase extraction. Baseline separation of the components provides the selectivity for the method. Validation was performed according to the European Medicines Agency (EMA) Guidelines and the method was found to be precise and accurate.Using our developed and validated method we were able to identify and quantify both saccharides in human milk for the first time. Based on our results the LacNAc concentration is in the range of 6.7-31 mu g/mL while LNB concentration decreased from 26 mu g/mL below the detection limit during the first week of lactation. The presence of LNB and LacNAc in human milk also implies new biological functions which can lead us closer to the understanding of the various functions of this complex biofluid. (C) 2015 Elsevier B.V. All rights reserved.