Enzyme Immobilization onto Ultrahigh Specific Surface Cellulose Fibers via Amphiphilic (PEG) Spacers and Electrolyte (PAA) Grafts

Abstract

Our group has investigated the incorporation of proteins in fibers by two approaches. One involves physical encapsulation of proteins in the bulk of ultrafine and porous fibers and the other bound proteins to fiber surfaces with reactive spacers and functional grafts. This paper reports two examples from the second approach. Both involve the addition of covalently bonded polymeric chains to ultra-fine cellulose fiber surfaces. One tethers enzyme proteins by covalently bonds via amphiphilic PEG spacers that carry reactive end groups. The other adds polyelectrolyte PAA grafts that are sufficiently polar to attract enzymes via secondary forces. Both surface polymer systems are compatible with aqueous and organic media. Lipase (E.C. 3.1.1.3, from Candida rugosa) enzyme was used. PEG was introduced by reacting cellulose with PEG diacylchloride followed by amide covalent bond formation between COOH of PEG and amine (NH 2 ) of the lipase. PAA (0.76-40.9 mmol of COOH per g cellulose) was grafted via ceric ion initiated polymerization. On the PEG attached cellulose, reactive COOH end groups ranging from 0.2 to 1.0 mmol per g cellulose. The adsorbed lipase on the PAA grafted cellulose fibers exhibited significantly higher immobilization efficiency, i.e., up to 391 Unit/mmol COOH, than the covalently bound lipase via PEG spacers. The immobilized lipases via both methods possessed much superior retention of catalytic activity following exposure to hydrocarbons, including cyclohexane, toluene, and hexane, than the free lipase. The covalently bound lipase exhibited significantly higher catalytic activity retention levels at elevated temperatures than the adsorbed and free form. In addition, both adsorbed and covalently bound lipases can be repeatedly used for 4 cycles.