Determination of 2-keto- l-gulonic, 2-keto- d-gluconic and 2,5-diketo- d-gluconic acids by capillary zone electrophoresis

Abstract

During the biosynthetic processing of 2-keto- l-gulonic acid (2-KLG) from 2,5-diketo- d-gluconic acid (2,5-DKG) by Corynebacterium sp., 2-keto- d-glutonic acid (2-KDG) is produced as a byproduct. These organic acids have been analyzed by high-performance liquid chromatography (HPLC) on an Aminex HPX-87H column, but the resolution was not good enough for quantitative analysis. We investigated the quantitation of 2-KLG, 2-KDG and 2,5-DKG using capillary electrophoresis (CE) and the results were compared with those of HPLC. With CE, in contrast to HPLC, good resolution, efficiency and rapid analysis were demonstrated as well as low consumption of solvent and samples. The CE system was applied at 15 kV with UV detection at 195 nm using 100 m M sodium borate (pH 8.4) as an electrolyte. The results were shown within 5 min with efficiency approaching 100 000 theoretical plates. The relative standard deviations of migration time and peak area were less than 0.9% and 1.6%, respectively. The detection limits for quantitative determination were 0.5–1.3 μM level. The above compounds, in fermentation broth, were analyzed under the optimum conditions. Considering the results of our study, the CE method should be highly suitable for the separation of 2-KLG, 2-KDG and 2,5-DKG in the fermentation broth.