Uronic acid quantitation

Polysaccharides which contain uronosyl residues, such as the acidic pectic polysaccharides and the glucuronoarabinoxylans, pose a particular problem in quantitative determinations of glycosyl compositions. Urono-sidic bonds are resistant to acid hydrolysis. Conditions sufficiently harsh to hydrolyze the uronosidic bonds often result in significant degradation of the uronic acids as well as degradation of some of the neutral sugars. Uronic acids, even when converted into their monomeric form, do not form stable alditol acetates as the reduced form of an uronic acid is an aldonic acid which cannot be acetylated by the standard procedures. 

The problems associated with the presence of uronosyl residues can be bypassed by converting the uronosyl residues to the corresponding hexosyl residues. This reduction is most successfully carried out by converting the uronosyl residues into lactones by reaction with water soluble carbodiimides. The lactones are reduced with sodium borodeuteride (124). It is important to use sodium borodeuteride rather than sodium 

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Blumenkrantz, N. Asboe-Hansen G. 1973. New method for quantitative determination of uronic acids. Anal. Biochem. 54 484-489. 

In model experiments,205 it was found that concentration of an aqueous solution of the aldonic acids in the presence of hydrochloric acid gives exclusively the 1,4-lactones, which, on subsequent trimethyl-silylation, give only one peak on the chromatogram. This method was successfully employed for the separation of D-galacturonic, D-glucuronic, and D-mannuronic acids. Of the 1,4-lactones examined, only the trimethylsilyl ether of D-mannono-1,4-lactone was obtained in crystalline form. However, all of the derivatives showed characteristic differences in their infrared spectra in the range of 1500 to 600 cm"1. When this method was applied to the determination of uronic acids in a variety of polysaccharides,205 it was impossible to find any hydrolytic conditions under which the uronic acids were quantitatively released and then reduced, a problem experienced by other workers.20 The method was, however, successful in affording a qualitative, microscale procedure for the identification of hexuronic acids (which otherwise are difficult to detect). 

The quantitative determination of uronic acids in polysaccharides is complicated by lactonization, and these problems have been carefully examined by Blake and Richards.26,98 In the course of these studies, D-glucuronic acid was reduced to D-glucitol, which was characterized as the hexaacetate.98 As uronic acid residues in a polysaccharide may be reduced relatively easily,465,466 determination as the alditol provides an alternative method of analysis.461,468 This procedure is most valuable when different uronic acids are present in... 

Infrared spectra of glycosaminoglycans in D20 and DC1 were used165 for quantitative evaluation of the uronic acid and acetamido groups in this class of polysaccharide. The results showed that i.r. spectroscopy of compounds in D20 solution provides a simple, quantitative basis for examination of ras(COJ) and amide I bands in heparin and chondroitin 4-sulfate. In DC1, analysis was based on p(C02H) and amide I bands which were well resolved. The apparent acid-dissociation constants of the studied polysaccharides were estimated from the absorbance of p(C02) or p(C02H) bands, or both, at different pH (pD) values. 

Traces of the complex carbohydrates contained in the juice survive the processing and appear in the molasses analytical data indicate that they probably consist largely of pectins and pentosans.10 Quantitative estimations based on the ash-free solids of Louisiana molasses revealed the presence of 2% uronic acids and 0.5% methoxyl.16 Heat-modified... 

The characterization and quantitative determination of uronic acid components in polysaccharides faces the problem of complete release of the uronic acids without accompanying decomposition. This is a difficult task because of the acid resistance of the glycosiduronic bond. From a comparison of several methods, it was shown that methanolysis combined with trifluoroacetic acid hydrolysis is the best for the liberation of uronic acids.204 The identification can be performed by gas chromatography of the trimethylsilyl derivatives.205... 

The reducing group in uronic acids can be oxidized by nitric acid or bromine to yield aldaric acids. Uronic acids, as salts, can be quantitatively reduced to aldonic acids by borohydride.239 Lactones are readily reduced, as in the controlled conversion of D-glucurono-6,3-lactone into D-gluco-hexodialdose. The carboxyl group of glucuronic acids may be reduced to a primary alcoholic group. 

A. Haug and B. Larsen, Quantitative determination of the uronic acid composition of alginates, Acta. Chem. Scand., 16 (1962) 1908-1918. 

Fazio, S.A., Uhlinger, D.L., Parker, J.H., and White, D.C. (1982) Estimations of uronic acids as quantitative measures of extracellular and cell wall polysaccharide polymers from environmental samples. Appl. Environ. Microbiol. 43, 1151-1159. 

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