Extraction and Purification of Sialic Acid from Submandibular Mucin of Cattle and Goat

Extraction and Purification of Sialic Acid from Submandibular Mucin of Cattle and Goat.

ABSTRACT

In this study, Sialic acid (Neuraminic acid) was extracted, purified and partially characterized from submandibular mucin of cattle and goat.  The extraction involved homogenization, treatment with enzymes (phospholipase and lipase) and acid hydrolysis. Ion exchange chromatography was used to purify the sialic acids.

The partial characterization involved thin- layer chromatography and Fourier transform infrared spectroscopy (FTIR). Treatment of the submandibular mucin with phospholipase led to an increase in release of sialic acid with optimum release of sialic acids at 90 minutes while treatment of the homogenate with lipase led to a slight increase in sialic acids released.

Acid hydrolysis of the homogenized submandibular glands led to a 15 fold increase in sialic acids released. The purification fold after carrying out ion exchange chromatography on the sialic acid extracts of the cattle and goat submandibular mucin was 16.59 and 12.11 respectively.

Thin-layer chromatography revealed that both sialic acid extracts moved at the same pace with a standard sialic acid from Sigma. The FTIR spectra of the purified sialic acid and that of the standard sialic acid had wavenumbers that are indicative of alkyl halide, alkenes, allene, alkane and carboxylic acid which are the functional groups that are found in sialic acids.

The results indicated that the sialic acid contained in goat submandibular gland is more and of similar quality with that from cattle submandibular gland. The results have indicated that sialic acids are extractible from the submandibular mucins with the aid of phospholipase and lipase.

INTRODUCTION

Sialic acid is a derivative of a nine-carbon monosaccharide. It comprises of a large family of N– and O-substituted neuraminic acids. The amino group of sialic acid is linked to either an N-acetyl or N-glycolyl group, which yields N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc) respectively. The hydroxyl groups of both sialic acids are often modified.

It is a naturally widespread carbohydrate with numerous biological functions, including blood protein half-life regulation, variety of toxin neutralization, starting reagent of biochemical derivatives for the synthesis of pharmaceuticals, cellular adhesion and glycoprotein lytic protection (Varki et al., 2009; Schauer, 2004a).

Sialic acid is covalently bound to the side chains of mucin by a 2-6′ glycosidic bond; however, the amount and types of sialic acids present in mucin vary by species of animal and system of isolation. For example, 22% of the dry weight of bovine submaxillary mucin is sialic acid, but the vast majority of the sialic acid is N-acetylneuraminic acid, with minor amounts of N-glycolylneuraminic acid (Mizan et al., 2000).

Their first level of diversity results from the different alpha linkages that may be formed between the Carbon-2 (C-2) of sialic acid and underlying sugars by specific sialyltransferases, using cyclic monophosphate bound to sialic acid (CMP-Sias) as high-energy donors. The most common linkages are to the C-3 or C-6 positions of galactose residues or to the C-6 position of N-acetylgalactosamine residues (Varki and Schauer, 2009).

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