Determination of mono- and oligosaccharides in fermentation broths by liquid chromatographic separation and amperometric detection using immobilized enzyme reactors and a chemically modified electrode

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Abstract
Liquid chromatographic (LC) determinations of mono- and disaccharides in complex fermentation broths and beverage samples are disturbed by the presence of interfering matrix components. High selectivity can be performed by coupling of LC to immobilized enzyme reactors and amperometric detection. The carbohydrates eluting from the column are first introduced into a reactor containing immobilized amyloglucosidase which hydrolyses the oligosaccharides into glucose. A second reactor follows the first one and contains coimmobilized glucosedehydrogenase and mutarotase. The monosaccharides eluting from the first reactor are mixed with a make-up flow consisting of a nicotineamide adenine dinucleotide (NAD+) buffer. The carbohydrates are oxidized in an equivalent amount of reduced coenzyme (NADH) which is detected electrochemically by using an electrode modified with a phenoxazine derivative. The postcolumn system was applied to a high-energy soft drink, malt beer, and fermentation broths from the penicillin industry.