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The development of an immobilized lactate oxidase system for lactic acid analysis
Author(s) -
Can John J.,
Chen LiFu,
Flickinger Michael C.,
Tsao George T.
Publication year - 1984
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260260209
Subject(s) - chemistry , immobilized enzyme , cofactor , chromatography , lactic acid , fermentation , biochemistry , enzyme , biology , bacteria , genetics
An enzyme designated as lactate oxidase was purified from Acetobacter peroxydans by using the partition methods of separation. A DE‐52 cellulose column was used for the primary purification of lactate oxidase, and the purified enzyme was covalently bound to a porous cellulose bead matrix in which benzoquinone was used as the coupling reagent. The physicochemical properties of the native and immobilized enzymes were determined including molecular weight, cofactor requirements, and optimal reaction conditions. Lactate oxidase was shown not to be subject to product inhibition, and to require Mg 2+ as a metal cofactor. Analysis of an immobilized lactate oxidase packed‐bed reactor indicated that this system may not be subject to internal diffusional limitations. Molecular oxygen appeared to be a cosubstrate of the enzyme, and a reaction mechanism was postulated to predict the kinetic behavior of the immobilized reactor system. Applications of the immobilized lactate oxidase reactor for the pulse‐flow analysis of lactic acid in whole milk and in a yeast fermentation system were considered.