Picrocrocin hydrolysis by immobilized β-glucosidase

beta -Glucosidase from sweet almond was immobilized onto a nylon support and used to hydrolyze picrocrocin, the glycoside precursor of the saffron essential volatile oil, safranal. The nylon support was derivatized as hydrazide and the enzyme attached through Schiff base to bonds. The coupling efficiency was 46.8%, the immobilization yield 29.5%, and the derivative showed 24.2 and 4.0 U/mg activity for p-nitrophenylglucoside and picrocrocin, respectively, as substrates. Kinetic parameters of the immobilized derivative were determined, with picrocrocin as substrate, showing K sub(M) = 7.2 mM and V sub(max) = 4,0 U/mg. Glucose behaved as a competitive inhibitor (K sub(i) = 95.0 mM). The immobilized derivative was thermally stable up to 45 degree C; from that temperature onwards thermoinactivation occurred. The operational deactivation showed a biphasic pattern, t sub(1/2) being 4.2 days for the first hour days of continuous operation, and 20.1 days from that point on. The immobilized enzyme lost a 50% of its initial activity after 30.7 days of storage at 4 degree C.