Application Of Ionic Liquids-Based Three-phase Microemulsion In The Enzymatic Catalysis

Ionic liquids have many unique properties, such as: almost negligible vapor pressure, super dissolvingcapacity, strong designability and the possible of forming some three-phase solution with some solvents.All these properties open the road to a wide range of applications including chemical synthesis, catalysisand extraction.In this paper, a three-phase microemulsion, which named water/microemulsion/lipophilic ionicliquids, has been formed by using lipophilic ionic liquid1-butyl-3-methylimidazolium hexafluorophosphateand with the aid of mixed surfactants. The ternary diagram was also studied with surfactants, ionic liquidand salt solution as the three components to discuss the range of each component in the formation of three-phase microemulsion. The results indicate that there is a big area of continuous phase microemulsionamong our research scope. Through studying the influence of lipase, substrate and product on the volumeof each phase, the three-phase microemulsion whose volume change least was employed as the reactionmedia. In this media, the enzymatic hydrolysis of4-nitrophenyl butyrate was conducted to investigate theimpact of reaction condition on the reaction, among which the temperature, pH and time has significantinfluence on the activity of lipase. What’s more, when the pH of buffter solution was8and temperature was45℃, the reaction rate was greatest. Under the optimal condition, the kinetic study of the enzymaticreaction in the three-phase and two-phase microemulsion was conducted, in which the value of VmaxandKmwas acquired based on Michaelis-Menten equation. The Vmaxvalue of three-phase microemulsion wasfive times of two-phase microemulsion which efficiently demonstrates the advantage of three-phasemicroemulsion on enzymatic hydrolysis. To further prove the superiority of three-phase microemulsion onenzymatic catalysis, the stability of lipase in the media was studied, which demonstrate that the residualactivity of lipase can still reach70%after incubated for72h, far higher than the two-phase microemulsion.Conformational studies via Fourier transform-infrared (FT-IR) spectroscopy indicated that lipasesentrapped in the three–phase system retain their native structure compared to biphasic media, whichefficiently explains the excellent catalytic properties.In order to further discuss the unique advantage of three-phase microemulsion on the scope of enzymatic catalysis, the research of enzymatic resolution of chiral drugs was also conducted byesterification reaction in which Candida rugosa lipase was the main enzyme and ibuprofen was thesubstrate. Through high performance liquid chromatography, the method to detection R-ibuprofen, S-ibuprofen and ibuprofen ester was established, among which R-ibuprofen and S-ibuprofen could beefficiently separated, while ibuprofen ester failed to separate. The comparation of enzymatic resolutionreaction of ibuprofen between three-phase and two-phase microemulsion illustrates that theenantioselectivity and activity of Candida rugosa lipase in three-phase microemulsion was far higher thanthe other medium. The superior of three-phase microemulsion has been testified once again. What’s more,in this research, the impact of some factors on the enzymatic esterification reaction has been focused,including the concentration of substrate, alcohol salt and total surfactant, the type of alcohol, temperatureand the mole ratio between substrate and alcohol. The result illustrated that the type of alcohol, the moleratio between substrate and alcohol and the concentration of alcohol has significant influence on thereaction.