title: Extraction and separation of proteins by ionic liquid aqueous two-phase system.
abstract: A satisfactory protocol of protein extraction and separation has been established based on the ionic liquid aqueous two-phase system (IL-ATPS) for the purification of bioactive substances. Compared with the effects of eight different ionic liquids, 1-octyl-3-methylimidazolium bromide ([omim][Br]) was selected as the suitable ionic liquid. Based on the single-factor experiment, an initial serial investigative test was used to identify the optimal conditions of the extraction. Owing to their different isoelectric points, bovine serum albumin (BSA), hemoglobin (Hb) and lysozyme (Lys) were used to determine the effect of pH value on the protein extraction. Trypsin (Try) was used to confirm the protein activity. The linearity for analyzing BSA, Hb, Try and Lys was in the concentration range of 0.05-1.00 mg ml(-1), 0.025-1 mg ml(-1), 0.01-1.00 mg ml(-1) and 0.01-1.00 mg ml(-1), respectively, with a correlation coefficient of between 0.9985 and 0.9999. Limits of detection (LODs) were 16.47-7.02 μg ml(-1) and RSDs of inter-day stability were less than 2.9%. Repeatability and precision were respectively lower than 5.3% and 1.1%. Under the optimum conditions, the average recoveries of BSA, Hb, Try and Lys were 90.5%, 94.5%, 92.7% and 93.8% and the obtained RSDs were 1.19%, 1.23%, 1.34% and 1.04%, respectively. According to UV spectra, conductivity, dynamic light scattering (DLS), and transmission electron microscope (TEM) images, the cluster phenomenon originating from IL itself or combined with protein was evaluated. As the driving forces which are involved in the partitioning of protein between the IL-rich phase and the phosphate phase, the cluster phenomenon could, in principle, be applied to a variety of different samples and exhibited potential value.
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082, P.R. China. wyzss@hnu.edu.cn.
authors.forename: Xiao
authors.initials: X
authors.lastname: Lin
authors.forename: Yuzhi
authors.initials: Y
authors.lastname: Wang
authors.forename: Qun
authors.initials: Q
authors.lastname: Zeng
authors.forename: Xueqin
authors.initials: X
authors.lastname: Ding
authors.forename: Jing
authors.initials: J
authors.lastname: Chen
mesh_terms: D000818:Animals; D002417:Cattle; D006454:Hemoglobins / Q000032:analysis; D006801:Humans; D052578:Ionic Liquids / Q000737:chemistry*; D059627:Liquid Phase Microextraction / Q000379:methods*; D009113:Muramidase / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D012710:Serum Albumin, Bovine / Q000032:analysis
chemical_list: D006454:Hemoglobins; D052578:Ionic Liquids; D011506:Proteins; D012710:Serum Albumin, Bovine; D009113:Muramidase