A highly efficient hydrophobic interaction chromatographic absorbent improved the purification of hepatitis B surface antigen (HBsAg) derived from Hansenula polymorpha cell |
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| Institution: | 1. National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100080, PR China;2. Department of Environmental Science, Beijing University of Science and Technology, Beijing 100083, PR China;1. Institute of Pharmaceutical Science, King''s College London, Franklin-Wilkins Building, 150 Stamford Street, London SE1 9NH, UK;2. Randall Division of Cell & Molecular Biophysics, King''s College London, New Hunt''s House, London SE1 1UL, UK;3. Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe 657-8501, Japan;4. Electron Microscopy for Materials Science (EMAT), University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp, Belgium;5. Centre for Molecular Oncology, Bart''s Cancer Institute, Queen Mary University of London, London EC1M 6BQ, UK;1. Department of Stomatology & Medical Research, Tung’s Taichung Metroharbor Hospital, Taichung, Taiwan;2. Institute of Molecular Biology, National Chung Hsing University, Taichung 402, Taiwan;3. Department of Medical Research, Taichung Veterans General Hospital, Taichung 407, Taiwan;4. Rong Hsing Research Center for Translational Medicine, National Chung Hsing University, Taichung 402, Taiwan;5. Ph. D Program in Translational Medicine, National Chung Hsing University, Taichung 402, Taiwan;6. Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung 912, Taiwan;7. Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA;8. The iEGG and Animal Biotechnology Center, National Chung Hsing University, Taichung 402, Taiwan;9. Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan;10. Depertment of Nursing, Jen-Teh Junior College of Medicine and Management, Hou-Loung Town, Taiwan;1. Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy, Sichuan University, Chengdu 610041, China;2. State Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China;3. University of Chinese Academy of Sciences, Beijing 100049, PR China;1. Beijing Laboratory of Biomedical Materials, College of Materials Science and Engineering, Beijing University of Chemical Technology, Beijing 100029, PR China;2. College of Pharmacy, Weifang Medical University, Weifang 261053, Shandong, PR China;3. Department of Dermatology, China-Japan Friendship Hospital, East Street Cherry Park, Chaoyang District, Beijing 100029, PR China;4. Key Laboratory of Biomedical Materials of Natural Macromolecules, Beijing University of Chemical Technology, Ministry of Education, Beijing 100029, PR China |
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| Abstract: | To improve the purification efficiency of recombinant hepatitis B surface antigen derived from Hansenula polymorpha (Hans-HBsAg), a serial of absorbents for hydrophobic interaction chromatography with the controllable ligand density and spacer arm were synthesized, then developed and further applied to purify Hans-HBsAg. The absorbent, Butyl-S QZT with the ligand density of 25 μmol/(g wet gel) and spacer arm of 3C, was screened out and its physical and chemical properties were evaluated. High rigidity and low backpressure (<0.06 MPa) were obtained at the flow rate up to 20 ml/min. Moreover, it has the stable chemical characteristics of subjecting to high concentrations of acid, alkali and detergents. This HIC absorbent was further applied to purify Hans-HBsAg with the recovery 94% and purification-fold 9 under the optimized operation condition at pH 6.5 and concentration of ammonium sulfate 7.5%. Finally, the HIC adsorbent of Butyl-S QZT was applied in the integrated three-step chromatographic purification process to purify Hans-HBsAg. About 140 mg of purified Hans-HBsAg was obtained from 1 l cell disruption supernatant at the total recovery of 27% and the purification-fold of 151.8. Based on the assay of SDS-PAGE and SEC-HPLC, the purity of the purified HBsAg was over 99% to meet the requirement for the further inoculation use. |
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