Covalent attachment of acetylcholinesterase on electrospun poly vinyl alcohol nanofiber:potential use in pesticide biosensor

Poly vinyl alcohol (PVA) nanofibers could be fabricated by electrospinning, and provide huge surface area for enzyme immobilization and catalytic reactions. In this study, acetylcholinesterase from Drosophila melanogaster (AChE) (E.C 3.1.1.7) has been successfully immobilized on poly vinyl alcohol (PVA) nanofibers via electrospinning of a mixture of acetylcholinesterase, bovine serum albumin (BSA) as an enzyme stabilizing additive and poly vinyl alcohol (PVA). The surface of nanofibrous membranes were covalently bonded with BSA and AChE via glutaraldehyde vapor. Immobilized enzyme activities were assayed by spectrophotometric technique and showed that immobilized AChE retained 40% of the free enzyme activity. The kinetic parameters of the free and immobilized AChE, Km and Kcat, were also determined. Moreover, the enzyme-nanofiber was found to be very stable, since the immobilized AChE retained >70% of its initial activity when stored in buffer at 25 ºC for 53 days, and 90% at 4 °C for 60 days. immobilized enzymes retained 90% of its initial activity after 10 consecutive reactor batch cycles. This compartment could be used for biosensor design for pesticide determination