title: Design of suitable carrier buffer for free-flow zone electrophoresis by charge-to-mass ratio and band broadening analysis.
abstract: In this work, charge-to-mass ratio (C/M) and band broadening analyses were combined to provide better guidance for the design of free-flow zone electrophoresis carrier buffer (CB). First, the C/M analyses of hemoglobin and C-phycocyanin (C-PC) under different pH were performed by CLC Protein Workbench software. Second, band dispersion due to the initial bandwidth, diffusion, and hydrodynamic broadening were discussed, respectively. Based on the analyses of the C/M and band broadening, a better guidance for preparation of free-flow zone electrophoresis CB was obtained. Series of experiments were performed to validate the proposed method. The experimental data showed high accordance with our prediction allowing the CB to be prepared easily with our proposed method. To further evaluate this method, C-PC was purified from crude extracts of Spirulina platensis with the selected separation condition. Results showed that C-PC was well separated from other phycobiliproteins that have similar physicochemical properties, and analytical grade product with purity up to 4.5 (A620/A280) was obtained.
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Fan-Zhi
authors.initials: FZ
authors.lastname: Kong
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.|School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Yu-Chen
authors.initials: YC
authors.lastname: He
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Guo-Qing
authors.initials: GQ
authors.lastname: Li
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Liu-Yin
authors.initials: LY
authors.lastname: Fan
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China.
authors.forename: Hua
authors.initials: H
authors.lastname: Xiao
authors.affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, P. R. China.
authors.forename: Shan
authors.initials: S
authors.lastname: Li
authors.affiliation: Laboratory of Analytical Biochemistry and Bio-separation, State Key Laboratory of Microbial Metabolism, School of Life Science and Biotechnology, Shanghai Jiao Tong University, Shanghai, P. R. China. cxcao@sjtu.edu.cn.
authors.forename: Cheng-Xi
authors.initials: CX
authors.lastname: Cao
mesh_terms: D000818:Animals; D002021:Buffers; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006454:Hemoglobins / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D010798:Phycocyanin / Q000032:analysis; D053545:Spirulina / Q000737:chemistry
chemical_list: D002021:Buffers; D006454:Hemoglobins; D010798:Phycocyanin
keywords: Band broadening; C-phycocyanin; Charge-to-mass ratio analysis; Preparative electrophoresis; Protein purification