D-Xylulose and d-Xylose: Determination with d-Xylose Isomerase

Abstract

This chapter elaborates the enzymatic determination of D-xylulose and D-xylose with D-xylose isomerase. The cysteine–carbazole reaction, in conjunction with D-xylose isomerase, can be employed for the determination of D-xylose and D-xylulose. The enzyme is readily purified from extracts of Lactobacillus plantarum. A correction is necessary if the sample contains L-arabinose or L-ribulose because even the best D-xylose isomerase preparations still have L-arabinose isomerase activity. The enzymatic method is based on the principle that D-xylose isomerase catalyzes the reaction, as given in this chapter. The equilibrium lies in favor of the aldopentose because the equilibrium constant is 4.55 at 23°C. In the presence of excess isomerase, 82% of the D-xylulose is converted to D-xylose. D-xylose does not react in the cysteine-carbazole test. Therefore, if the color test is carried out before and after incubation with D-xylose isomerase, the difference in color intensities is equivalent to 82% of the D-xylulose present in the sample. The method is standardized with crystalline ribulose-o-nitrophenylhydrazone, which reacts quantitatively as ketopentose in the color test. A similar procedure is used for the determination of D-xylose, except that the reaction is carried out in borate buffer at pH 8.2 instead of tris buffer pH 7.5.