Microbial Production of Glucose Oxidase

Productivity of extracellular glucose oxidase was examined for various microorganisms and it was found in strains belonging to genus Penicillium except one species of Tallalomyces.

As the best glucose oxidase producer, Penicillium purpurogenum No. 778 was isolated from natural source. This microorganism produced 32,000 units per ml broth of glucose oxidase in a simple medium containing beet molasses, NaNO³ and KH²PO⁴ by submerged culture for 3 days. That value was about 10-times of that of Penicillium amagasakiense which has been known as an excellent glucose oxidase producer.

Culture conditions for glucose oxidase production were examined, which were extremely different among microbial species. In the case of Penicillium chrysogenum AJ 7007 and Penicillium purpurogenum No. 778, the effects of aeration and carbon sources were remarkably different from each other.

Penicillium purpurogenum No. 778 produces catalase sufficiently in a culture broth for glucose oxidase application in food industry.

Glucose oxidase was purified about 25-fold from culture supernatants of Penicillium purpurogenum No. 778, and some properties of the enzyme were examined. The optimum temperature and pH for the activity were 35°C and 5.0, respectively. The enzyme was stable at pH 5.0 to 7.0 when it was incubated at 40°C for 2 hr, while it was stable at temperature lower than 50°C when incubated at pH 5.6 for 15 min. The enzyme was specific for d-glucose and apparent Michaelis constant for d-glucose was 12.5 mm. The enzyme was inhibited by 1 mm of HgCl², CuSO⁴, NaHSO⁴ and phenylhydrazine, but not inhibited by 1 mm of p-hydroxy-mercuribenzoate, EDTA, hydroxylamine and dimedone. Four percents NaCl inhibited the activity about 50%, while the addition of ethanol (from 0 to 16%) increased oxygen uptake more than that expected from the peroxidase activity of catalase.