Kinetic studies and analytical application of cholesterol oxidase and peroxidase immobilized to synthetic polymer |
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Authors: | Lyubov K Yotova Ivan P Ivanov |
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Institution: | (1) Department of Biotechnology, University of Chemical Technology and Metallurgy, 1756 Sofia, Bulgaria;(2) Department of Chemistry and Biochemistry, Medical University, Zdrave Str. 2, 1431 Sofia, Bulgaria |
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Abstract: | A method for individual and simultaneous covalent immobilization of cholesterol oxidase and peroxidase to copolymer of acrylonitrile
with acrylamide is described. The effect of immobilization on the catalytic properties of the covalently bound enzymes was
studied. The immobilized enzymes showed no change in pH optima and an increase in temperature optima, activation energy, and
K
m
, compared to data received from experiments with soluble enzymes. A small glass column packed with immobilized multienzyme
complex was used to develop a method for manual determination of cholesterol in foodstuffs (e.g., in mayonnaise “Olinease”).
The method was characterized by high analytical precision (coefficient of variation = 2.67%). The results show high correlation
with those obtained by the Kageyama method (r=0.986). The method is economical (the enzyme-carrier conjugate may be used more than 300 times), precise, easy to perform,
and less time-consuming than the manual methods utilizing soluble enzymes. The established manual method can be proposed for
cholesterol determination in foodstuffs. |
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Keywords: | Immobilization simultaneous immobilization cholesterol measurement cholesterol oxidase peroxidase copolymer of acrylonitrile with acrylamide |
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