Quantification and characterisation of polyphenol oxidase from vanilla bean

Polyphenol oxidase (PPO) of Vanilla planifolia Andrews beans was extracted and purified through ammonium sulphate precipitation, dialysis, and gel filtration chromatography. PPO activity was measured by improved UV technique using 4-methylcatechol and catechol as substrates increasing substantial sensitivity of previous procedure. The optimum pH and temperature for PPO activity were found to be 3.0 and 3.4 and 37 °C, respectively. Km and Vmax values were found to be 10.6 mM/L and 13.9 OD300 min−1 for 4-methylcatechol and 85 mM/L and 107.2 OD300 min−1 for catechol. In an inhibition test, the most potent inhibitor was found to be 4-hexylresorcinol followed by ascorbic acid. The thermal inactivation curve was biphasic. Activation energy (Ea) and z values were calculated as 92.10 kJ mol−1 and 21 °C, respectively.