Abstract
A post-column enzyme reactor with co-immobilized glucoamylase and glucose oxidase has been used for the detection of α-glucans after HPLC separation of native and NaBH4-reduced oligosaccharides. Oligomers eluting from the column were cleaved into monomers which were oxidized with production of hydrogen peroxide, which was measured amperometrically at a platinum electrode. Detection of α-glucans proved to be highly selective and sensitive. The enzyme reactors are very stable under the conditions used. Linear calibration graphs were observed over 3 decades, and the detection limit for various oligosaccharides is into the lower ng range. Efficient separations were obtained on reversed phase columns using gradient elution with acetonitrile. Reduced oligomers were separated up to a degree of polymerization >40. The influence of temperature, pH, reaction time and sodium borohydride concentration was evaluated for the reduction of α-glucans. Reduction is complete within 2 minutes at 80°C and within 20 minutes at room temperature.
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