Polysaccharides reduction with borohydride

In most cases, uronic acids are liberated from acidic polysaccharides by hydrolysis leading to irreproducible concomitant formation of lactones. Several methods to circumvent this problem have been published describing conversion of the uronic acid into methyl esters followed by reduction with borohydride or borodeuteride reagents and subsequent hydrolysis and GC-MS detection [129]. Other techniques are based on the liberation and quantification of carbon dioxide. Direct determination of uronic acid residues in hydrolyzates has frequently been performed according to colorimetric assays, which are rather insensitive and have thus mostly been replaced by high-performance anion exchange chromatography (HP-AEC) methods [130-132]. 

SXIA contains phosphorus, galactose, and glucose in the ratio 1 2 1, in addition to 0-acetyl groups (12%). Sequential reduction with borohydride and alkaline hydrolysis yield glycerol phosphate and a reduced polysaccharide, which, on acidic hydrolysis, yields only galactose and glucose, in the ratio 2 1. 

Much attention has been given to finding efficient procedures for the dismantling of oxidized polysaccharides, and also to methods for the separation and characterization of product mixtures. Of the fragmentation procedures employed (see Section VI. 1), that " in which the dialdehyde is reduced, and the polyol formed is subjected to total acid hydrolysis, is the most widely applied. (Although the procedure is sometimes referred to as the Smith degradation , this term is more commonly reserved for selective hydrolysis of the polyol under milder acidic conditions (see Section VI.4.) In one variation,the dialdehyde is methylated prior to reduction with borohydride, which differentiates free hydroxyl groups in the oxidized polymer from those formed upon reduction of the aldehydes. 

The oligosaccharides (11) and (12) have been isolated from the degradation products obtained from cellulase treatment of the L-arabino-D-xylo-D-glucan from tobacco leaves, Their structures were determined from the results of methylation analysis both before and after reduction with borohydride ion as well as by and n.m.r. spectroscopies. Present in the hemicellulose fraction from the endosperm cell walls of rice was a D-galacto-D-xylo-D-glucan. This polysaccharide was based on a (1-> 4)-j3-D-glucan backbone to which were attached residues of D-xylose and D-galactose at 0-6. 

The Smith degradation4 involves reduction of the periodate-oxidized polysaccharide with borohydride, followed by mild hydrolysis with acid. (The periodate oxidation has already been treated in Section 111,1, p. 200.) The modified sugar residues contain acyclic acetal groupings, which are hydrolyzed much faster than the glyco-sidic linkages. The product therefore contains small fragments (such... 

The peeling of xylans is basically similar to that of cellulose (Fig. 2). Extended alkali treatment of a rye flour arabinoxylan at room temperature resulted in only a 29% reduction in molecular weight [265]. As anticipated, the polysaccharide, after reduction with sodium borohydride, was completely stable to alkali. 

The most widely applied method for the transformation of the polyaldehydes furnished by the above oxidation process, is the Smith-degradation involving successive oxidation with periodate in aqueous medium, removal of the iodate and periodate ions and subsequent reduction with sodium borohydride. The resulting polyalcohol is then hydrolyzed and the products in the hydrolysate are analyzed. Of the polysaccharides amylopectin (4) and cellulose (5) have been first analyzed by this method as demonstration of the usefulness of the procedure which has emerged recently as a generally employed way of analysis of polysaccharides. 

The oxidation of polysaccharides with periodate is inhibited, to varying extents, by the formation of hemiacetals, so that further oxidation can be performed only after the removal (by reduction with sodium borohydride) of the protecting hemiacetals. Theoretical analysis of the kinetics of such repeated oxidations predicts that, for complete protection, the oxidation limit of the second oxidation, as a function of the extent of the first oxidation, lies between 41.1 and 43.2 %. This prediction is supported by values found experimentally for alginate. 

An efficient procedure for converting polysaccharides into their monosaccharide constituents is by hydrolysis at 121° for 1 hr with 2N tri-fluoroacetic acid. Trifluoroacetic acid is volatile and can be easily removed from the hydrolyzed samples by evaporation in a stream of air. The monosaccharides are converted to the corresponding alditols by reduction with sodium borohydride and the alditols are acetylated with acetic anhydride in the presence of a weak base, such as sodium acetate. The steps involved in converting a polysaccharide into its constituent alditol acetates are summarized in Fig. 1. 

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