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Liver alcohol dehydrogenase immobilized on polyvinylidene difluoride
Author(s) -
Roig M. G.,
Bello J. F.,
Moreno Vega M. A. De,
Cachaza J. M.,
Kennedy J. F.
Publication year - 1990
Publication title -
journal of chemical technology and biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.64
H-Index - 117
eISSN - 1097-4660
pISSN - 0268-2575
DOI - 10.1002/jctb.280490202
Subject(s) - alcohol dehydrogenase , diffusion , adsorption , substrate (aquarium) , chemistry , kinetic energy , immobilized enzyme , membrane , work (physics) , enzyme , chromatography , thermodynamics , organic chemistry , biochemistry , physics , oceanography , quantum mechanics , geology
Abstract A physical method for immobilization of liver alcohol dehydrogenase (ADH) by hydrophobic adsorption onto a supporting membrane of polyvinylidene difluoride (PVDF) was performed. Simultaneously, a physicochemical characterization of the immobilized enzyme regarding its kinetic behaviour was performed. The activity/pH profile observed points to an effect of pH on activity that is completely different from the case of ADH in solution. The disturbance in the typical bell‐shaped profile owing to the fact that the enzyme was immobilized is explained on the basis of a potent limitation to the diffusion of the protons in the support. The findings of the present work also reveal the existence of an effect that limits free external diffusion of the substrate towards and/or the product from the support; this effect seems to be the determinant of the overall rate of the enzymatic reaction and is thus of great importance in the effective kinetic behaviour (v([S])) of immobilized ADH, whose kinetic behaviour is complex (non‐Michaelian), as may be seen from the lack of linearity observed in the corresponding double reciprocal and Eadie‐Hofstee plots. By non‐linear regression numerical analysis of the v([S]) data and application of the F‐ test for model discrimination, the minimum rate equation necessary to describe the intrinsic kinetic behaviour of PVDF‐immobilized ADH proved to be one of the polynomial quotient type of degree 2:2 (in substrate concentration).