Abstract
A major problem in determining the composition of plant cell wall polysaccharides has been the lack of a suitable method for accurately determining the amounts of galacturonic and glucuronic acids in such polymers. A gas chromatographic method for aldose analysis has been extended to include uronic acids. Cell wall polysaccharides are depolymerized by acid hydrolysis followed by treatment with a mixture of fungal polysaccharide-degrading enzymes. The aldoses and uronic acids released by this treatment are then reduced with NaBH(4) to alditols and aldonic acids, respectively. The aldonic acids are separated from the alditols with Dowex-1 (acetate form) ion exchange resin, which binds the aldonic acids. The alditols, which do not bind, are washed from the resin and then acetylated with acetic anhydride to form the alditol acetate derivatives. The aldonic acids are eluted from the resin with HCl. After the resin has been removed, the HCl solution of the aldonic acids is evaporated to dryness, converting the aldonic acids to aldonolactones. The aldonolactones are reduced with NaBH(4) to the corresponding alditols, dried and acetylated. The resulting alditol acetate mixtures produced from the aldoses and those from the uronic acids are analyzed separately by gas chromatography. This technique has been used to determine the changes in composition of Red Kidney bean (Phaseolus vulgaris) hypocotyl cell walls during growth, and to compare the cell wall polysaccharide compositions of several parts of bean plants. Galacturonic acid is found to be a major component of all the cell wall polysaccharides examined.
Highlights
A major problem in determining the composition of plant cell wall polysaccharides has been the lack of a suitable method for accurately determining the amounts of galacturonic and glucuronic acids in such polymers
The derivative is recorded on magnetic tape for subsequent electronic integra- formed from 4-O-methyl-D-glucuronic acid, 4-0-methyl-sorbitol tion of peak areas
Duplicate 10-mg samples of cell wall material from hypocotyl tissue of Small White, Red Kidney, and Pinto bean plants were analyzed as described under "Materials and Methods." The percentage yields for each of the polysaccharide constituents are given on the ordinate axis
Summary
A major problem in determining the composition of plant cell wall polysaccharides has been the lack of a suitable method for accurately determining the amounts of galacturonic and glucuronic acids in such polymers. The resulting alditol acetate mixtures produced from the aldoses and those from the uronic acids are analyzed separately by gas chromatography. This technique has been used to deter. Methods for determining polysaccharide composition have, in general, relied upon the separation of the monosaccharide components of hydrolytic digests by means of paper chromatography and electrophoresis [51, 64] and identification of these components by a variety of color reactions [20]. Such methods are both laborious and subject to errors of considerable magnitude. In this case both D-galacturonic and D-glucuronic acids are oxidized, and what is determined is the sum of the concentrations of the two acids
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