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peak areas due to NADH were observed at pH 8.0 when the pH of the carrier consisting of piperazine- 1,4-bis(2-ethanesulfonic acid) (PIPES) buffer ranged from 7.0 to 8.5. Various buffer types were also ex- amined as carrier media at pH 8.0, and PIPES buffer showed the maximum peak area. When the carrier composed of PIPES buffer (0.1M, pH 8.0) contain- ing 1mM NAD+ and 5mM MgCl2 was used, the cali- bration curve for D-malate was linear in the range of 0.02-50µ M( r =1 .000). The detection limit (S/N = 3) was 0.01µM. Relative standard deviations of the peak area at 1µ Ma nd 10µ Mw ere 1.6% (n =7 ) and 0.48% (n =7 ), respectively. This method was applied to the analysis of D-malate in several beverages, and the recovery test of the added D-malate to samples was also carried out to afford good results.

Determination of D-Malate Using Immobilized D-Malate Dehydrogenase in a Flow System and its Application to Analyze the D-Malate Content of Beverages