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Automatic determination of Michaelis—Menten constants by the variable flow-rate technique

✍ Scribed by Juliana Marcos; Angel Ríos; Miguel Valcárcel

Publisher: Elsevier Science
Year: 1993
Tongue: English
Weight: 772 KB
Volume: 283
Category: Article
ISSN: 0003-2670
DOI: 10.1016/0003-2670(93)85254-h

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✦ Synopsis

An automatic method for the determination of Michaelis-Menten constants of enzymatic reactions in unsegmented flow systems was developed using the variable flow-rate technique, allowing the substrate concentration to be changed during the experiments in order to obtain different reaction rates. The proposed method can be applied both to enzymes in solution and immobilized, in this case on controlled-pore glass, and packed in a micro-reactor. Various flow-rate-time patterns were used for this purpose and the results were compared. Values of 1 X lo-* and 3 x 10e4 M were found for Michaelis-Menten constants corresponding to the reaction between /3-o-glucose and glucose oxidase by using dissolved and immobilized enzyme, respectively, with standard deviations of kO.1 x lo-' (dissolved enzyme) and kO.1 X 10d3 (immobilized enzyme).

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