We report here a two-step procedure preparing lipase nanogel. The first step is to introduce vinyl groups onto enzyme surface and the second step is in-situ polymerization using acrylamide as monomers. The nanogel network, while enhancing the stability of encapsulated enzyme via multiple covalent attachments, generates minor additional resistance for mass transport. The greatly enhanced durability against harsh conditions, e.g. high temperature and the presence of organic solvent, indicates a great potential for industrial process of many kind.

       Lipase (EC 3.1.1.3) is extensively studied for potential application in synthesis of surfactants and pharmaceuticals, production of biodiesel, resolution of racemic mixtures, and detergent industry. The production of biodiesel from plant oil can be efficiently catalyzed by lipase in a water-containing system without an organic solvent. However, this process is hindered by methanol induced deactivation of lipase.

       In this work, lipase nanogel was prepared using above mentioned procedure. Transmission electron microscope (TEM), size exclusion chromatography (SEC), SDS-PAGE, FT-IR were used to characterize the enzyme nanogel. Using p-Nitrophenylburate as substrate, Km and Vmax of lipase nanogel was 0.29 mM, 51 Umg-1 while their respective counterpart of free lipase was 0.28 mM, 87 Umg-1. The half-life time of native lipase was 50 min at 50°C while almost no visible loss of lipase activity was observed in the 500 min for the nanogel. After incubation in 40% methanol aqueous solution, lipase nanogel preserved about 80% of its hydrolytic activity while native lipase reserved only 10%. All these are of great interest for further exploration of the potential application of lipase nanogel for the production of biodiesel.