Polymer Immobilized Alkylimidazolium Ionic Liquids In Extraction And Purification Of Proteins

Ionic liquids have been attracted many attentions in analysis of various matters. The functional groups contained in the structure endow the ionic liquid with a high efficiency in separation and enrichment. Meanwhile, their proper nonvolatility brings on an environmental friendly pretreatment process. As a result, an ionic-liquid-based extraction technique has been established for separation and purification of proteins. However, the ionic liquids normally have high viscosity and some of them are miscible with water, which result in slow mass transport processes, low reaction rate to reach the equilibrium as well as difficult phase separations for the extraction. To solve these problems, the ionic liquids were immobilized chemically or physically on a surface of solid phase substances. Compared to that of the pure ionic liquids, the surface of the immobilized ionic liquids are more adjustable for providing more active specific properties. The so called solid phase extraction has been proposed to separate metal ions, organic compounds and medicines effectively. The superior physicochemical properties of ionic liquids, especially compared to organic solvents, have been attracted many attentions on extraction and separation of biological samples such as proteins.In this work, polymer immobilized ionic liquids were used to separate and enrich proteins from complex samples. The utilization of the ionic liquids was improved and the quality of the ionic liquids was reduced. The interactions between the eluent, the surfactant sodium dodecyl sulfate (SDS) and the proteins were studied in order to provide a mechanism for better understanding. The proposed methods have been used to separate hemoglobin (Hb) from the complex practical sample of the human blood.Imidazolium cations were grafted on the surface of chloromethyl polystyrene, termed as PS-CH2-[MIM]+CF. This modified polymer material was applied to selectively extract protein Hb. The prepared extractant PS-CH2-[MIM]+Cl-, containing 2 mmol of immobilized imidazole groups per gram polymer, was characterized by FT-IR, surface charge analysis and chemical element analysis, respectively. An adsorption efficiency of 91% was achieved. The sorption capacity of the PS-CH2-[MIM]+Cl- to Hb was 23.6 μg·mg-1 and 80% of the retained Hb could be readily recovered using 0.5%(m/v) SDS aqueous solution as the eluate. The activity of the eluted Hb was maintained about 90%. The prepared imidazolium containing solid phase polymer was directly used to adsorb Hb at certain conditions without using any other solid matrix as the support of the ionic liquid. This material was practically used to isolate Hb from human whole blood.An imidazolium containing sulfonated polyetheretherketone (SPEEK) composite, termed as SPEEK-Bmim, was prepared from SPEEK and l-butyl-3-methylimidazolium hexafluorophosphate. This composite material was characterized with FT-IR, surface charge analysis and elemental analysis. The prepared material could selectively adsorb protein Hb. The adsorption efficiency was about 90%. The sorption capacity of the SPEEK-Bmim to Hb was 31.6μg mg-1, and 84% of the retained Hb could be readily recovered by elution with 0.5%(m/v) aqueous SDS solution. The activity of the eluted Hb was about 91%. This ionic liquid-modified SPEEK composite was also practically used to isolate Hb from human whole blood.The interactions between SDS and Hb were investigated by the fluorescence quenching technique. The results of fluorescence titration revealed that SDS could strongly quench the intrinsic fluorescence of Hb through a static quenching procedure. The apparent binding constants K and number of binding sites n of SDS with Hb were obtained accordingly. The thermodynamic parameters were calculated. The change in enthalpy (△H) was found to be minus, and the change in entropy (AS) was Slightly greater than zero, which indicated that the interaction of SDS with Hb was driven mainly by electrostatic interaction. Moreover, the change in Gibbs free energy was minus indicating a spontaneous binding process of SDS and Hb during the elution. The results of synchronous fluorescence, three-dimensional fluorescence and circular dichroism (CD) spectra showed that the SDS induced conformational changes of Hb.