ISOLATION AND PURIFICATION OF POLYPHENOL OXIDASE FROM A NEW VARIETY OF POTATO

Polyphenol oxidase (E.C. 1.10.3.1) was extracted from potato (Solanum tuberosum var. Tebere) tissue and purified using the techniques of ammonium sulphate fractionation, DEAE-cellulose chromatography and filtration through Sephadex G-200. Four isoenzymes with different relative mobilities were detected by polyacrylamide gel electrophoresis. The enzyme had a molecular weight of 112,000 as estimated by gel filtration on Sephadex G-200 and an apparent Km of 8.68 .times. 10-5M for chlorogenic acid as substrate, and pH optimum of 6.6. The heat inactivation plot at 65.degree. C at different pH values was biphasic. It was also found that the Arrhenius plot for chlorogenic acid oxidation was biphasic showing a discontinuity at 8.degree. C with an increased slope as temperatures were reduced to 1.degree. C. Cinnamic and ferulic acids were competitive inhibitors with respect to chlorogenic acid with Ki values of 2 .times. 10-5 and 5 .times. 10-6M, respectively.