Characterization of polyphenol oxidase from butter lettuce (Lactuca sativa var. capitata L.)

Polyphenol oxidase (PPO) was isolated from butter lettuce (Lactuca sativa var. capitata L.) grown in Poland and its biochemical characteristic were studied. PPO from butter lettuce showed a higher affinity to 4-methylcatechol than to catechol. The KM and Vmax values were: 3.20 ± 0.01 mM and 4081 ± 8 U/ml min−1 for catechol and 1.00 ± 0.09 mM and 5405 ± 3 U/ml min−1 for 4-methylcatechol. The optimum pHs of the enzyme were found to be 5.5 using catechol and 6.8 using 4-methylcatechol as substrate. The enzyme had a temperature optimum of 35 °C. The enzyme was relatively stable at 30 °C and 40 °C. The times required for 50% inactivation of activity at 50 °C, 60 °C and 70 °C were found to be about 30, 20 and 5 min, respectively. Inhibitors used for investigation in this study were placed in relative order of inhibition: p-hydroxybenzoic acid > glutathione ≈ ascorbic acid > l-cysteine > EDTA > citric acid. The enzyme eluted in the chromatographic separations was analyzed electrophoretically under denaturating conditions. The analysis revealed a single band on the SDS–PAGE which corresponded to a molecular weight of 60 kDa.