Uronic acids, detection

Fig. 2. HPLC ion exchange gradient separation of samples (25 ug of uronic acid) of smaller (G7) citrus pectin oligomers (a), larger (G12) citrus pectin oligomers (b), and the B. fruit extract (c). Detection was by PAD. For each sample the peak co-eluting with the standard galacturonate oc-tamer was designated peak 8, and the remaining peaks were numbered consecutively.

The identification of L-iduronic acid as the major glycuronic acid constituent of heparin proved to be a much slower process than the identification of the amino sugar residue. Although this compound was detected in acid hydrolyzates of heparin116117 and heparin oligosaccharides,118 its yield was usually poor, because of the drastic conditions used for the acid hydrolysis (which are known to lead to extensive destruction of uronic acid).119120 Also, L-iduronic acid escaped detection as L-idose in the hydrolyzates of carboxyl-reduced heparin, probably because L-idose is readily converted into 1,6-anhydro-L-idose under the usual hydrolytic conditions. 

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). 

Uronic acid in the form of galacturonic acid is a major component of the pectic polysaccharide rhamnogalacturonan, which is present in large amounts in the cell walls of most fruits and vegetables. Small amounts of glucuronic acid and 4-O-methylglu-curonic acid have also been detected in cell walls. 

The mucilage consists mainly of D-galactose, the content of which is about 70% on an ash-free basis. L-Galactose is considered to be absent because the benzimidazole procedure of Link and coworkers48 failed to detect it. While carrageenin is believed to contain no uronic acid, Dilsea mucilage contains 9.6-11.0% of it. The nature of the uronic acid is... 

The dinitrophenylhydrazones of the fission products of sugars are detectable with sodium hydroxide.160 Also, a wide variety of sugars and their acetates, and uronic acid lactones may be detected with sodium hydroxide followed by heating,161 Sensitivity is poor, however. After paper electrophoresis, chondroitinsulfate may be distinguished from proteins by means of Toluidine Blue.162... 

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]. 

It is interesting that polysaccharides obtained from O inagglutinable, capsular strains (having K antigens) were reported to contain uronic acid neutral sugars were also present, however, and 0 antigen specificity was also detected in the products. 

Non-endospermic xylans have substituent groups of D-glucopyrano-syluronic acid, or its 4-methyl ether, or both. Individual molecules may lack such residues, but, almost certainly, other xylan molecules in the total hemicelluloses from each grass have both groups.137,157 Uronic acid groups may escape detection or identification in hydrolyzates of xylans studied by paper chromatography, and in hydrolyzates and methanolyzates of methylated xylans studied by g. l.c. after conversion of the products into volatile derivatives. 

Crook and Johnston and Martin and Adams were unable to find uronic acids in the cell walls of fungi. However, 2 % of a glucuronic acid was detected in the dry cells of Dactylium dendroides, and lesser proportions in Aspergillus niger (0.1 %), Penicillium album (0.2%), a Fusarium sp. (0.5%), and an Alternaria sp. (0.7%). Glucuronic acid residues are apparently present in the cell walls of Neurospora crassa. ... 

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