Nanofibrous membranes with an average diameter of 100 and 180nm were fabricated from poly(acrylonitrile-co-maleic acid) (PANCMA) by the electrospinning process. These nanofibrous membranes contain reactive groups which can be used to covalently immobilize biomacromolecules. Two natural macromolecules, chitosan and gelatin, were tethered on these nanofibrous membranes to fabricate dual-layer biomimetic supports for enzyme immobilization in the presence of 1-ethyl-3-(dimethyl-aminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxyl succinimide (NHS). Lipase from Candida rugosa was then immobilized on these dual-layer biomimetic supports using glutaraldehyde (GA), and on the nascent PANCMA fibrous membrane using EDC/NHS as coupling agent, respectively. The properties of the immobilized lipases were assayed. It was found that there is an increase of the activity retention of the immobilized lipase on the chitosan-modified nanofibrous membrane (45.671.8%) and on the gelatin-modified one (49.771.8%), compared to that on the nascent one (37.671.8%). The kinetic parameters of the free and immobilized lipases, Km and Vmax, were also assayed. In comparison with the immobilized lipase on the nascent nanofibrous membrane, there is an increase of the Vmax value for the immobilized lipases on the chitosan- and gelatin-modified nanofibrous membranes. Results also indicate that the pH and thermal stabilities of lipases increase upon immobilization. The residual activities of the immobilized lipases are 55% on the chitosan-modified nanofibrous membrane and 60% on the gelatin-modified one, after 10 uses.

Novel nanofibrous membranes containing reactive carboxyl groups were fabricated from poly- (acrylonitrile-co-maleic acid) (PANCMA) by the electrospinning process. The morphology and fiber diameter were analyzed with field emission scanning electron microscopy. It was found that the fiber diameter could be varied from 100 to 600 nm by changing the solution concentration. Lipase from Candida rugosa was covalently immobilized onto the membrane surface via the activation of carboxyl groups in the presence of 1-ethyl-3- ((dimethylamino)propyl)carbodiimide hydrochloride/N-hydroxylsuccinimide. The properties of the immobilized lipases on the nanofibrous and hollow fiber PANCMA membranes were measured. It was found that, compared with the hollow fiber membrane, the enzyme loading and the activity retention of the immobilized lipase on the nanofibrous membrane increase from 2.36 ( 0.06 to 21.2 ( 0.7 mg/g and from 33.9 to 37.6%, respectively. The kinetic constants of the free and immobilized lipases, Km and Vmax, were assayed. Results indicate that the Vmax values are similar for both immobilized enzymes, while the Km value of the immobilized enzyme decreases from 1.36 on the hollow fiber membrane to 0.98 on the nanofibrous membrane. The studied lipase-immobilized nanofibers can be used as biocatalysts for polyester synthesis and/or in situ formation of nanofiber reinforcement composites.