Sialic acid methanolysis

The neuraminic acids (also called sialic acids), which occur in the terminal positions of carbohydrate moieties of many glycoproteins and ganglio-sides, must be hydrolyzed under very mild conditions. For example, hydrolysis with 0.01 M hydrochloric acid for 30 min at 100° causes 20% decomposition of N-acetylneuraminic acid. Sialic acids are more tolerant to methanolysis they may also be removed enzymically. 

The position of the bond between the sialic acids was determined by g.l.c.-m.s. of the acetates of partially methylated sialic acid derivatives obtained after methanolysis of the methylated sialoglycolipid, and was confirmed by the periodate-oxidation data. The (2- 9) bond between... 

In the methanolysis procedure, as used for the standard quantitative monosaccharide GLC analysis of glycoconjugates [240,241], 1 M methanolic HCl (24 h, 85 C) is applied. However, under these conditions released sialic acids are completely de-Af,0-acylated, which makes this approach unsuitable for the characterization of different types of sialic acid. It is, however, a reliable approach for the determination of the total amount of a mixture of (0-acetylated) A-acylneuraminic acids. When using a milder methanolysis procedure (0.05 M methanolic HCl, 1 h, 80 C) the de-iV-acylation but not the de-O-acylation is strongly reduced [242]. 

Methanolysis in anhydrous methanolic HCI, followed by N-acetylation of amino sugars and sialic acid, and derivatization of the methyl glycosides formed to volatile trimethylsilyl ethers (TMS) or trifluoracetyl derivatives are widely used (17,18). 

Analytical procedures used in the routine analysis of glycoproteins often involve methanolysis and subsequent N-acetylation of the liberated amino-sugars. However, it has been demonstrated that partial 0-acetylation of alditols (commonly employed as internal standards) accompanies the N-acetylation of amino-sugars. It was also shown that a hot neutral solution of methyl chloride in methanol releases L-fucose, sialic acid, and some D-galactose residues from aj-acid... 

An alternative method for the preparation of sialic acids is methanolysis of crude complex carbohydrate preparations with hydrochloric acid, which leads to the formation of neuraminic acid-p-methylglycoside (Neu- -Me) from all natural N- and 0-acyl derivatives of neuraminic acid (Schauer and Buscher 1974, ScHAUER 1978). This method can easily be employed on a large scale, e.g. yielding 8.8 g Neu-p-Me from 100 g of edible bird s nest substance. It allows preparation of various homogeneous N-acyl derivatives of neuraminic acid on re-N-acetylation and acid removal of the -methyl group, with or without a radioactive label (Schauer et al. 1970, Schauer and Buscher 1974). 

---