Abstract

Background: Polyphenol oxidase (PPO) has applications in pharmaceutical, environmental, and food industries. However, conventional techniques for its purification are considered costly and laborious. Alternative methods of purification are needed.

Methods: This study explored the use of an alcohol/salt-based Aqueous Two-Phase System (ATPS) to partially purify PPO from Mango Seed Kernel (MSK). Specifically, this study determined the effects of the 1-propanol/potassium phosphate buffer mass ratio on the partition parameters such as partition coefficients, activity recovery, and purification fold.

Results: The enzyme partition coefficient (KE) was greater than 1, which signified that more PPO was partitioned in the top phase due to its hydrophobic nature. Specifically, the 3.22 1-propanol/ PPB mass ratio had the highest K E of 1.91 ± 0.16 and 1.46 ± 0.16 for runs 1 and 2, respectively. Moreover, the same mass ratio led to the highest activity recovery in the top phase reaching up to 73.99 ± 1.56% for run 1 and 64.27 ± 2.48% for run 2. In terms of purification fold, runs 1 and 2 had PF values of 0.55 ± 0.05 and 0.70 ± 0.01, respectively. Since the values obtained were low due to the large number of contaminants extracted together with the target enzyme in the top phase, the use of a back-extraction step recovered the PPO in the bottom phase and resulted in an activity recovery of 82.94 ± 1.05% and a purification fold of 7.01 ± 0.95.

Conclusion: Results have shown that varying the mass ratio had significant effects on the partition parameters.

Keywords: Polyphenol oxidase, ATPS, partial purification, mango seed kernel, crude enzyme extract, PPO.

Graphical Abstract