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

Réka Balogh, Péter Jankovics, S. Béni

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

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. μg/mL while LNB concentration decreased from 26. μg/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.

Original languageEnglish
Pages (from-to)140-146
Number of pages7
JournalJournal of Chromatography A
Volume1422
DOIs
Publication statusPublished - Nov 27 2015

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High performance liquid chromatography
Human Milk
Tandem Mass Spectrometry
Oligosaccharides
Mass spectrometry
Carbon
High Pressure Liquid Chromatography
Chemical analysis
Sugar Alcohols
Size exclusion chromatography
Medicine
Solid Phase Extraction
Lactation
Gel Chromatography
Limit of Detection
N-acetyllactosamine
Monitoring
Guidelines

Keywords

  • HPLC-MS/MS
  • Human milk oligosaccharide
  • Lacto-N-biose
  • N-Acetyllactosamine
  • Porous graphitic carbon

ASJC Scopus subject areas

  • Analytical Chemistry
  • Organic Chemistry
  • Biochemistry

Cite this

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title = "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",
abstract = "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. μg/mL while LNB concentration decreased from 26. μg/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.",
keywords = "HPLC-MS/MS, Human milk oligosaccharide, Lacto-N-biose, N-Acetyllactosamine, Porous graphitic carbon",
author = "R{\'e}ka Balogh and P{\'e}ter Jankovics and S. B{\'e}ni",
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T1 - 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

AU - Balogh, Réka

AU - Jankovics, Péter

AU - Béni, S.

PY - 2015/11/27

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N2 - 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. μg/mL while LNB concentration decreased from 26. μg/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.

AB - 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. μg/mL while LNB concentration decreased from 26. μg/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.

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