ABSTRACT
In this study, acetylcholinesterase (AChE) and choline oxidase (ChO) were co-immobilized on poly(2-hydroxyethyl methacrylate) (pHEMA) membranes with the aim of using them in biosensor construction. pHEMA membranes were prepared with the addition of different salts in different HEMA: aqueous solution ratios and characterized in terms of porosity, thickness. permeability, and mechanical properties. Membranes prepared in the presence of SnCl4 were found to be superior in terms of porosity and permeability and were chosen as the immobilization matrix. Immobilization of the enzymes was achieved both by entrapment and surface attachment via epichlorohydrin (Epi) and Cibacron Blue F36A (CB) activation. The effect of immobilization on enzyme activity was evaluated by the comparison of Km and Vmax values for the free and immobilized bi-enzyme systems. The increase in Km was negligible (1.08-fold) for the bi-enzyme system upon immobilization on surface but was 2.12-fold upon entrapment. Specific activity of the free enzyme system was found to be 0.306 mV s(-1) microg(-1) ChO while it was 0.069 (4.43-fold decrease) for entrapped and 0.198 (1.54 fold decrease) for CB-Epi immobilized enzymes. The performance of immobilized enzymes in different buffer types, pH, and temperature conditions were evaluated. The best enzyme activity was obtained at pH 9.0. Activity of the enzymes was found to increase with increasing temperature (in the range 25-40 degrees C).
Subject(s)
Acetylcholinesterase/chemistry , Alcohol Oxidoreductases/antagonists & inhibitors , Biosensing Techniques , Cholinesterase Inhibitors/pharmacology , Enzyme Inhibitors/pharmacology , Membranes, Artificial , Polyhydroxyethyl Methacrylate/chemistry , Hydrogen-Ion Concentration , Kinetics , Microscopy, Electron, Scanning , Models, Chemical , Permeability , Temperature , Time Factors , Tin CompoundsABSTRACT
Aldicarb loaded microspheres of carboxymethyl cellulose, CMC, of various compositions were prepared with the crosslinking action of Al(III). Into these microspheres, lignin was introduced as a filler to further regulate the rate of release. The encapsulation efficiency was low (4.15-13.00%) due to the high water solubility of aldicarb. Release into aqueous media was quite rapid and almost complete within 24h. Initial stages of the release (60% of AS released) was investigated to determine the release mechanism and it was found that release from the microspheres was governed by Fickian diffusion. Among the various parameters, the ratio of lignin to CMC and aldicarb to CMC were found to be influential on the rate of release from the crosslinked CMC-lignin microspheres. The soil behaviour confirmed the delayed release effect of the controlled release formulation compared to Temik (a commercial preparation of aldicarb), especially when lignin was incorporated.
