Flow-injection determination of 3-hdroxybutyrate in serum with an immobilized 3-hydroxybutyrate dehydrogenase reactor and chemiluminescence detection |
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Authors: | Nobutoshi Kiba Hidekazu Koemado and Motoshisa Furusawa |
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Institution: | Department of Applied Chemistry and Biotechnology, Faculty of Engineering, Yamanashi University, Kofu 400, Japan |
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Abstract: | A flow-injection system for the determination of 3-hydroxybutyrate in serum is described. 3-Hydroxybutyrate dehydrogenase is immobilized on poly(vinyl alcohol) beads and incorporated in a flow-injection system. 1-Methoxy-5-methylphenazinium methylsulphate reacts with enzymatically generated NADH to give H2O2, which is detected chemiluminometrically with the reaction of luminol and hexacynoferrate(III). Serum is diluted and filtered through an ultrafiltration membrane. The system responds linearly to injected samples (80 μl) in the concentration range 0.5–300 μM; the detection limit is 0.1 μM. The within-day relative standard deviation (n = 90) for 58 μM 3-hydroxybutyrate in serum is 0.8%. The maximum throughout is 20 samples per hour. The immobilized enzyme is stable for at least 1 month. |
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Keywords: | Chemiluminescence Flow injection enzyme reactor Hydroxybutyrate Serum |
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