Design of an immobilized enzyme system for naringin hydrolysis at high-pressure

Abstract

The effect of pressure was studied in an enzymatic reaction with an immobilized biocatalyst. Naringinase immobilized by entrapment in calcium alginate beads was the biocatalyst used to catalyze, at high-pressure, the hydrolysis of naringin to naringenin. These molecules have great potential in the pharmaceutical industry due to their recognized anti-oxidant, anti-inflammatory, anti-carcinogenic, anti-hypertensive and hypocholesterolemic effects. At high-pressure, the influence of relevant parameters on naringinase catalytic activity such as temperature, substrate concentration, and biocatalyst reuse was studied. At 160 MPa, naringinase entrapped in Ca-alginate beads displayed higher activity, namely in the range of 35–40 °C, whereas the optimum, at atmospheric pressure, was 35 °C. The immobilized naringinase presented a Michaelis–Menten kinetic, with a 65% higher maximum initial rate ( V max ap = 0.069 mM mi n − 1 ) , and a 70% lower K M ap ( 0.097 mM ) at 160 MPa, as compared to kinetic parameters, at atmospheric pressure ( V max ap = 0.042 mM min − 1 and K M ap = 0.303 mM ) . A positive effect of pressure on naringin hydrolysis by immobilized naringinase in Ca-alginate beads was confirmed with a negative activation volume (Δ V ≠) of −9 mL mol −1. The stability of immobilized naringinase was also evaluated at high-pressure.