Thermal and pH Stabilities of Partially Purified Polyphenol Oxidase Extracted from Solanum melongenas and Musa sapientum Fruits

Kinetic Studies of Polyphenol Oxidase from White Yam (Dioscorea rotundata Poir) Tuber

“…The activity profile of DR-PPO under varied temperature and pH conditions conformed to the inverted bell-shaped curve as earlier described [13,17,27,52]. The pH versus DR-PPO activity profile displayed peak values of enzyme activities at pH ≈ 8.5 units, pH ≈ 10.5 units and pH ≈ 11.5 units ( Figure 2), which were obvious indication of the presence of isoforms of DR-PPOs, based on previous propositions [21,27,38,[52][53][54][55][56][57]. The study by Altunkaya and Gökmen, [21] showed the presence of several PPO isoforms that have been implicated in browning reactions.…”

“…Studies have established the fact that variations in pH optimum of PPO activity depended largely on the plant maturity and cultivar, [40,49,55] in concert with experimental factors such as extraction methods and purities of enzyme, presence of activators/ inhibitors in addition to the experimental buffers and substrates used for analyses [9,49]. In general, most plants exhibit maximum PPO activity near neutral pH values [13,27,60]. depending on the nature of the experimental substrate used for the enzyme assay [10,40,55] as well as physiologic characteristics such as the cultivar, stage of maturation and presence of isoenzyme [53].…”

“…Extraction and partial purification of PPO was according to the methods of Madani et al, [26] with minor modifications as previously reported by Chikezie et al [27] Ten grams (10 g) each of the 6 sample was homogenized in external ice bath, using Ultra-Turrax T25 (Janke and Kunkel, Staufen, Germany) homogenizer set in 80 mL of 0.20 M K 2 PO 4 /KHPO 4 (phosphate buffer); pH=7.5 units containing 10 mM ascorbic acid (Darmstadt, Germany) for 180 sec at intervals of 15 sec. The homogenate was quickly mixed with 200 mL of acetone (Sigma-Aldrich, St. Louis, USA) (-20°C) to eliminate phenolic compounds [13] and squeezed through two layers of clean cheese cloth into a beaker kept in ice.…”

“…PPO activity was measured immediately after the extract was partially purified. Enzyme assay was according to the methods of Qudsieh et al, [30] with minor modifications according to Chikezie et al [27] Enzyme activity was determined by a measure of increase in absorbance the analyte using a spectrophotometer (U-2000 Hitachi, Japan) at 25 ± 5ºC. The reaction mixture contained 3.5 mL of 0.20 M phosphate buffer (pH=7.5 units), 1.0 mL of each serial dilutions of 4-16 mM catechol, and 0.5 mL of enzyme solution in a final volume of 5.0 mL.…”

“…The effect of temperature and pH on PPO activity as a measure of enzyme stability was ascertained according to the methods of Chikezie et al [27] Purified enzyme solutions extracted from D. rotundata tuber were stored in capillary tubes (1.0 mm i.d., 200 μL total volume) and held on ice until pre-incubated in a circulating water bath (Model 20B, Julabo, Allentown, PA) [19] at temperatures of 50, 60 and 70°C. At regular time intervals of 5, 10 and 15 min, aliquots of the enzyme solution was withdrawn and assayed for PPO activity.…”