Comparative Study on Characterization of Bromelain Extracted from the Stem and Fruit of Pineapple (Ananas comosus)

Abstract

Bromelain is a group of cysteine proteases that is found in pineapple (Ananas comosus) and has a wide range of medical and industrial applications. The present research work characterized bromelain extracted from the stem and fruit of fully ripened pineapple plants by homogenization in an aqueous buffer and then purified by ammonium sulfate precipitation and DEAE-cellulose chromatography. Enzyme activities were spectrophotometrically determined using N-α-Carbobenzoxy-Lysine p-Nitrophenyl Ester (LNPE) as substrate. The purified fractions of fruit bromelain (FBM) and stem bromelain (SBM) from the DEAE-cellulose chromatography (both at 0.05 mM NaCl conc.) were subjected to an evaluation of their temperature and pH optima, and kinetic parameters (K_m and V_max) determinations. The results obtained revealed that FBM had higher optimum temperature (45ºC) than SBM (35ºC). Both enzymes showed the same pH optima of 8.0. In addition, FBM (K_m = 0.39 mM and V_max = 0.38 μmol/min) was found to be proteolytically more active with greater affinity towards the substrate used than SBM (K_m = 1.17 mM and V_max = 0.25 μmol/min). This could serve as possible criteria for quality control/standardization protocols for both enzymes for specific applications and paves the way for exploring their potential usefulness.

Keywords: Cysteine Proteases, Bromelain, N-α-Carbobenzoxy-Lysine p-Nitrophenyl Ester, Optimum Temperature, Optimum pH, Kinetic Parameters.