title: Entrapment of α-Amylase in Agar Beads for Biocatalysis of Macromolecular Substrate.
abstract: Attempts have been made to optimize immobilization parameters, catalytic property, and stability of immobilized α-amylase in agar. The work compares natural entrapment efficiency of agar with the ionotropically cross-linked agar hydrogel, with the advantage of easy scale-up and cost and time effectiveness. Beads prepared with 3% (w/v) agar and 75 mM calcium chloride and hardened for 20 minutes were selected for further studies on the basis of entrapment efficiency (80%) and physical stability. Following entrapment, pH and temperature optima of enzyme were shifted from 6 to 6.5 and 50 to 55°C, respectively. Michaelis constant (K m ) for both free and entrapped enzymes remained the same (0.83%) suggesting no change in substrate affinity. However, V max⁡ of entrapped enzyme decreased ~37.5-fold. The midpoint of thermal inactivation for entrapped enzyme increased by 8 ± 1°C implying its higher thermal stability. The entrapped enzyme in calcium agar bead had an Ea value of 27.49 kcal/mol compared to 17.6 kcal/mol for free enzyme indicating increased stability on entrapment. Half-life of enzyme increased ~2.2 times after entrapment in calcium agar at 60°C indicating stabilization of enzyme. The reusability of beads was size dependent. Beads with diameter <710 μm were stable and could be reused for 6 cycles with ~22% loss in activity.
authors.affiliation: Department of Pharmacy, Banasthali University, Bansthali 304022, India.
authors.forename: Manu
authors.initials: M
authors.lastname: Sharma
authors.affiliation: Department of Bioscience and Biotechnology, Banasthali University, Banasthali 304022, India.
authors.forename: Vinay
authors.initials: V
authors.lastname: Sharma
authors.affiliation: Department of Pharmaceutics, Delhi Institute of Pharmaceutical Sciences and Research, Formerly College of Pharmacy, University of Delhi Pushp Vihar, Sector III, New Delhi 110017, India.
authors.forename: Dipak K
authors.initials: DK
authors.lastname: Majumdar