STUDIES ON KINETIC PARAMETERS AND BIOCHEMICAL CHARACTERISTICS OF POLYPHENOL OXIDASE PURIFIED FROM JACKFRUIT (ARTOCARPUS HETEROPHYLLUS) WASTE

  • Geetha Rani Y Department of Biotechnology, K. S. Rangasamy College of Technology, Tiruchengode- 637215, Tamil Nadu
  • Arulvel R Department of Biotechnology, K. S. Rangasamy College of Technology, Tiruchengode- 637215, Tamil Nadu
  • Selvarajan E School of Bioscience and Technology, Vellore Institute of Technology, Vellore, Tamil Nadu.

Abstract

Objectives: Polyphenol oxidase activity was extensively studied in jackfruit for its role in enzymatic browning. PPO and the phenolic compound play a vital role in defensive mechanism against pest and diseases. Thus, to facilitate further studies in jack fruit waste, Polyphenol oxidase [PPO] was purified and characterized.

Methods: Partial Purification of PPO from waste done through a sequential process of ammonium sulfate precipitation, dialysis and ion-exchange chromatography [DEAE- Cellulose]. Then the partially purified PPO was subjected to check various parameters like molecular weight and kinetic activity, the following characteristics of enzyme are checked: SDS-PAGE, pH, temperature, thermal stability, heat inactivation, metal ions, surfactants and inhibitor.

Results: Purified PPO resulted in ~23 folds enriched in the specific activity of 1360 [µkat/mg] and it was found to be the monomer with a molecular weight of 63 kDa revealed by Coomasie Brilliant Blue staining. PPO exhibited optimum activity at pH 7.0 and temperature 20oC. PPO showed the maximum stability between pH 6.4- 7.6 at 10 oC - 40 oC. PPO showed the enzyme activity towards Diphenol> Triphenol> Monophenol, the substrate specificity was especially high towards the catechol at 0.1 M. The PPO activity was activated by Mn2+, Triton X- 100, EDTA, Sorbic acid and Citric acid, but inhibited by L- cysteine, Ascorbic acid, SDS, Cetyl trimethyl ammonium bromide [CTAB], K+, Zn2+, Ca2+ and Mg2+. Kinetic constant for PPO was found to be km= 15.82 mM and Vmax= 2182 U/ml min using catechol as substrate.

Conclusion: Partial Purification of PPO from waste done through a sequential process of ammonium sulfate precipitation, dialysis and ion-exchange chromatography [DEAE- Cellulose]. The best substrate for PPO was identified as catechol [diphenol] and best inhibitor was L-cysteine and ascorbic acid.

 

Keywords: Polyphenol oxidase, PPO purification, Catechol, Ascorbic acid and L- Cysteine.

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Y, G. R., A. R, and S. E. “STUDIES ON KINETIC PARAMETERS AND BIOCHEMICAL CHARACTERISTICS OF POLYPHENOL OXIDASE PURIFIED FROM JACKFRUIT (ARTOCARPUS HETEROPHYLLUS) WASTE”. International Journal of Pharmacy and Pharmaceutical Sciences, Vol. 6, no. 9, 1, pp. 473-8, https://innovareacademics.in/journals/index.php/ijpps/article/view/2346.
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