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Polarographic measurement of oxygen uptake using lipoxygenase in reverse micelles
Author(s) -
Hochkoeppler Alejandro,
Palmieri Sandro
Publication year - 1990
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.260360704
Subject(s) - chemistry , micelle , lipoxygenase , polarography , clark electrode , chromatography , oxygen , enzyme , electrode , biochemistry , inorganic chemistry , organic chemistry , aqueous solution , electrolyte
Lipoxygenase‐catalyzed linoleic acid peroxidation was chosen as a model system to study the applicability of oxygraphy to monitor the oxygen uptake in organic solvents containing reverse micelles. Care was taken to control the oxygen back transfer from the atmosphere to the sample micellar solution, resulting in a significant improvement of electrode response. Under these conditions, lipoxygenase activity was linear up to 100 μg of enzyme. Given the quality of the calibration curve and the good correlation between lipoxygenase and ascorbate oxidase, the described technique is proposed as an alternative method for determining lipoxygenase activity in reverse micelles. The reliability of this technique was confirmed by the good agreement between polarography and classic spectrophotometry in kinetic studies. Preliminary experiments carried out on soybean cells solubilized in a Tween 85‐isopropylpalmitate system demonstrated that a light‐dependent oxygen uptake can be measured. The authors propose that the Clark‐type electrode be employed to study both the activity of oxidasic enzymes in reverse micelles and cell viability and physiology in organic solvents.