Partial purification of a banana polyphenol oxidase using triton X-114 andPEG 8000 for removal of polyphenols

Banana pulp polyphenol oxidase was partially purified in a latent form usin g sequential aqueous two-phase systems based on Triton X-114 and PEG 8000/p hosphate. The purification achieved from a crude extract of banana pulp was 5-fold, with 50% recovery of the activity. The (poly)phenols, including ta nnins, were also reduced to 6% of the original, avoiding the postpurificati on tanning of the enzyme and rendering only one enzymatic form in contrast to the many previously shown for this enzyme in both electrophoretic and is oelectrofocusing methods. The enzyme was kinetically characterized with its natural substrate (dopamine) and tert-butylcatechol in the presence and in the absence of the main activating agent, sodium dodecyl sulfate. In addit ion, the effect of several inhibitors was also tested, and the K-s, values of the two most effective substrate analogues (tropolone and kojic acid) we re determined.