OPTIMISATION OF BROMELAIN PURIFICATION FROM PINEAPPLE PEEL (Ananas comosus L.) USING RESPONSE SURFACE METHODOLOGY

Abstract

 Bromelain, a cysteine protease with broad industrial uses, can be efficiently extracted from pineapple processing residues, offering a sustainable method for waste valorisation. This study aimed to optimise the purification of bromelain from pineapple (Ananas comosus L.) peel using dialysis combined with Response Surface Methodology (RSM) based on a Box-Behnken design. A crude enzyme extract was obtained through different extraction media, followed by partial purification through ammonium sulfate precipitation and subsequent dialysis. The effects of pH (6 - 8), dialysis duration (20 - 30 h), and buffer-to-sample ratio (100:1 - 200:1, v/v) on specific activity were assessed. A quadratic regression model was applied to the bromelain purification process with statistically significant results (p < 0.01, R² = 0.9971). The optimal conditions were determined as pH 6.93, a dialysis time of 25.16 h, and a buffer-to-sample ratio of 134.50 (v/v), yielding a predicted specific activity of 235.86 U/mg protein, which closely matched the experimental value of 235.48 U/mg protein. The purified bromelain demonstrated high stability within pH 6 - 8 and at temperatures of 40 - 50 °C. SDS-PAGE analysis confirmed a predominant protein band at 25 - 30 kDa. These findings show that dialysis combined with statistical optimisation offers an effective and scalable method for bromelain purification, supporting the sustainable use of pineapple processing waste. This study provides one of the first systematic optimisations of dialysis parameters for bromelain purification from pineapple peel, highlighting its potential as a low-cost alternative to conventional chromatographic techniques. Compared with previously reported purification approaches, the proposed method demonstrates competitive specific activity while significantly reducing operational complexity and cost.