Quantification of sialic acids in red meat by UPLC-FLD using indoxylsialosides as internal standards

Glycoconj J (2016) 33:219226 DOI 10.1007/s10719-016-9659-1

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Quantification of sialic acids in red meat by UPLC-FLD using indoxylsialosides as internal standards

Hong L. Yao1,2 & Louis P. Conway1 & Mao M. Wang1 & Kun Huang1 & Li Liu1 &

Josef Voglmeir1

Received: 27 January 2016 /Revised: 23 February 2016 /Accepted: 25 February 2016 /Published online: 11 March 2016 # Springer Science+Business Media New York 2016

Abstract Herein we describe a UPLC-FLD-based method for the quantification of the sialic acid content of red meat, using a synthetic neuraminic acid derivative as an internal standard. X-Gal--2,6-N-propionylneuraminic acid was synthesized via a chemoenzymatic pathway and its hydrolytic stability was characterized. Known quantities of this compound were incubated with samples of red meat under sialic acid-releasing conditions. The released sialic acids were derivatized, analyzed by UPLC-FLD, and the Neu5Ac/ Neu5Gc content of the meat sample was determined by comparison with the internal standard. A number of red meats were analyzed by this method with the following results (Neu5Ac g/g tissue, Neu5Gc g/g tissue s.d.): pork (68 3, 15.2 0.7), beef (69 8, 36 5), lamb (46 2, 33 1), rabbit (59 2, 0.4 0.4), and hare (50 4, 1 1). We envisage that this methodology will find application in investigating the health effects of dietary Neu5Gc.

Keywords Sialic acid . Neu5Gc . N-glycolylneuraminic acid, Neu5Ac . red meat . UPLC-FLD

AbbreviationsCMP cytidyl 5-monophosphateDCC dicyclohexylcarbodimideEcNeuS Escherichia coli neuraminic acid synthaseESI electrospray ionizationFLD fluorescence detectorKdn -2,6-keto-3-deoxy-D-glycero-D-galactononic acidMALDI matrix-assisted laser desorption ionizationMS mass spectrometryNeu5Ac N-acetylneuraminic acidNeu5Gc N-glycolylneuraminic acidNeu5Pr N-propylneuraminic acidNHS N-hydroxysuccinimideNmCSS Neisseria meningitidis CMP-sialic acid synthase PdST6 Photobacterium damselae -2,6-sialyltransferase OPD o-phenylenediamineUPLC ultra high performance liquid chromatography X-Gal 5-bromo-4-chloro-indolyl--D-galactopyranoside

Introduction

Sialic acids are a family of monosaccharides with a nine carbon backbone which are commonly found as terminal residues in glycoconjugates while rarely appearing in their free form [15]. To date, more than 50 members have been identified [6], with the two best known being N-acetylneuraminic acid 1 (Neu5Ac) and N-glycolylneuraminic acid 2 (Neu5Gc) (Fig.1a) [79].

Neu5Ac 1 and Neu5Gc 2 are the most abundant members of the sialic acid family in mammals. Humans are capable of...