Butanolysis Derivatization: Improved Sensitivity in LC-MS/MS Quantitation of Heparan Sulfate in Urine from Mucopolysaccharidosis Patients

Paul J. Trim, John J. Hopwood, Marten F. Snel

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Abstract

Heparan sulfate (HS) is a complex oligosaccharide that is a marker of a number of diseases, most notably several of the mucopolysaccharidoses (MPS). It is a very heterogeneous compound and its quantification at physiological concentrations in patient samples is challenging. Here, we demonstrate novel derivatization chemistry for depolymerization/desulfation and alkylation of HS based on butanolysis. The resultant alkylated disaccharides are quantifiable by LC-MS/MS. This new method is at least 70-fold more sensitive than a previously published methanolysis method. Disaccharide yield over time is compared for methanolysis, ethanolysis, and butanolysis. Maximum disaccharide concentration was observed after 2 h with butanolysis and 18 h with ethanolysis whereas a maximum was not reached over the 24 h of the experiment with methanolysis. The sensitivity of the new technique is illustrated by the quantification of HS in 5 μL urine samples from MPS patients and healthy controls. HS was quantifiable in all samples including controls. Disaccharide reaction products were further characterized using exact mass MS/MS.

LanguageEnglish
Pages9243-9250
Number of pages8
JournalAnalytical Chemistry
Volume87
Issue number18
DOIs
Publication statusPublished - 15 Sep 2015

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

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title = "Butanolysis Derivatization: Improved Sensitivity in LC-MS/MS Quantitation of Heparan Sulfate in Urine from Mucopolysaccharidosis Patients",
abstract = "Heparan sulfate (HS) is a complex oligosaccharide that is a marker of a number of diseases, most notably several of the mucopolysaccharidoses (MPS). It is a very heterogeneous compound and its quantification at physiological concentrations in patient samples is challenging. Here, we demonstrate novel derivatization chemistry for depolymerization/desulfation and alkylation of HS based on butanolysis. The resultant alkylated disaccharides are quantifiable by LC-MS/MS. This new method is at least 70-fold more sensitive than a previously published methanolysis method. Disaccharide yield over time is compared for methanolysis, ethanolysis, and butanolysis. Maximum disaccharide concentration was observed after 2 h with butanolysis and 18 h with ethanolysis whereas a maximum was not reached over the 24 h of the experiment with methanolysis. The sensitivity of the new technique is illustrated by the quantification of HS in 5 μL urine samples from MPS patients and healthy controls. HS was quantifiable in all samples including controls. Disaccharide reaction products were further characterized using exact mass MS/MS.",
author = "Trim, {Paul J.} and Hopwood, {John J.} and Snel, {Marten F.}",
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TY - JOUR

T1 - Butanolysis Derivatization

T2 - Analytical Chemistry

AU - Trim, Paul J.

AU - Hopwood, John J.

AU - Snel, Marten F.

PY - 2015/9/15

Y1 - 2015/9/15

N2 - Heparan sulfate (HS) is a complex oligosaccharide that is a marker of a number of diseases, most notably several of the mucopolysaccharidoses (MPS). It is a very heterogeneous compound and its quantification at physiological concentrations in patient samples is challenging. Here, we demonstrate novel derivatization chemistry for depolymerization/desulfation and alkylation of HS based on butanolysis. The resultant alkylated disaccharides are quantifiable by LC-MS/MS. This new method is at least 70-fold more sensitive than a previously published methanolysis method. Disaccharide yield over time is compared for methanolysis, ethanolysis, and butanolysis. Maximum disaccharide concentration was observed after 2 h with butanolysis and 18 h with ethanolysis whereas a maximum was not reached over the 24 h of the experiment with methanolysis. The sensitivity of the new technique is illustrated by the quantification of HS in 5 μL urine samples from MPS patients and healthy controls. HS was quantifiable in all samples including controls. Disaccharide reaction products were further characterized using exact mass MS/MS.

AB - Heparan sulfate (HS) is a complex oligosaccharide that is a marker of a number of diseases, most notably several of the mucopolysaccharidoses (MPS). It is a very heterogeneous compound and its quantification at physiological concentrations in patient samples is challenging. Here, we demonstrate novel derivatization chemistry for depolymerization/desulfation and alkylation of HS based on butanolysis. The resultant alkylated disaccharides are quantifiable by LC-MS/MS. This new method is at least 70-fold more sensitive than a previously published methanolysis method. Disaccharide yield over time is compared for methanolysis, ethanolysis, and butanolysis. Maximum disaccharide concentration was observed after 2 h with butanolysis and 18 h with ethanolysis whereas a maximum was not reached over the 24 h of the experiment with methanolysis. The sensitivity of the new technique is illustrated by the quantification of HS in 5 μL urine samples from MPS patients and healthy controls. HS was quantifiable in all samples including controls. Disaccharide reaction products were further characterized using exact mass MS/MS.

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U2 - 10.1021/acs.analchem.5b01743

DO - 10.1021/acs.analchem.5b01743

M3 - Article

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SP - 9243

EP - 9250

JO - Analytical Chemistry

JF - Analytical Chemistry

SN - 0003-2700

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