Flow-injection fluorometric quantification of pyruvate using co-immobilized pyruvate decarboxylase and aldehyde dehydrogenase reactor: Application to measurement of acetate, citrate and l-lactate |
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Authors: | Tsukatani Tadayuki Matsumoto Kiyoshi |
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Institution: | a Biotechnology and Food Research Institute, Fukuoka Industrial Technology Center, Kurume 839-0861, Japan b Division of Food Biotechnology, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School Kyushu University, Fukuoka 812-8581, Japan |
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Abstract: | A flow-injection system for the quantification of pyruvate based on the coupled reactions of pyruvate decarboxylase (PDC) and aldehyde dehydrogenase (AlDH) was conceived and optimized. A co-immobilized PDC and AlDH reactor was introduced into the flow line. Sample and reagent (NAD+) were injected into the flow line by an open sandwich method and the increase of NADH produced by the immobilized-enzyme reactor was monitored fluorometrically at 455 nm (excitation at 340 nm). Linear relationships between the responses and concentrations of pyruvate were observed in the ranges of 2.0 × 10−5 to 1.5 × 10−3 M at the flow rate of 1.0 ml min−1 and 5.0 × 10−6 to 1.0 × 10−3 M at the flow rate of 0.5 ml min−1. The relative standard deviation for 10 successive injections was 0.95% at the 1.0 mM level. This FIA system for pyruvate was applied to the measurement of acetate, citrate and l-lactate. |
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Keywords: | Flow-injection analysis Immobilized-enzyme reactor Fluorometry Pyruvate Pyruvate decarboxylase Aldehyde dehydrogenase |
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