新型超大孔纤维素介质的研制及其在蛋白质色谱中的应用
|
摘要
为了制备一种可以在高操作流速下保持高效率的新型色谱介质,我们开展了本研究,制备了一种超大孔球形纤维素介质,测试了介质的传质性能,并将其应用于实际物系中蛋白质的分离纯化。
首先,以固体碳酸钙颗粒为致孔剂,采用热熔胶转化再生纤维的方法制备了超大孔纤维素介质。介质内含有超大孔和微孔结构。对牛血清白蛋白的相对孔隙率和有效孔扩散系数都得到了提高。其最高操作流速与刚性大孔介质相仿。孔内传质以对流传质为主。与微孔介质填充柱相比,超大孔介质填充柱的动态吸附容量和柱效都得到了提高,并且不受流速的影响。说明其在高速蛋白质色谱中具有很好的应用前景。
其次,为提高超大孔纤维素介质的吸附容量,采用硝酸铈胺(Ce~(4+))引发的方法,将甲基丙烯酸缩水甘油酯(GMA)接枝到超大孔纤维素介质的骨架上,制备了触角式超大孔纤维素介质。介质经接枝后,其静态吸附容量比接枝前介质的高2倍。接枝后,介质的孔径减小,孔内对流传质的程度降低。但是,触角式超大孔介质具有较高的静态吸附性能,因此仍然拥有较高的动态吸附容量。
为进一步研究超大孔纤维素介质的传质性能,在二维电场电色谱柱中,考察了电场对其传质性能的影响。电场的存在促进了介质孔内的传质,并且二维电场对传质的促进作用最强。与电场对超大孔介质填充柱传质性能的影响相比,电场对微孔介质的影响更显著,说明超大孔介质的传质性能比较好。并且实验还发现,随着流速的增大,二维电场对超大孔介质传质的促进作用增强,说明超大孔介质电色谱柱更适合在高流速下操作。
在超大孔纤维素介质的应用方面,开展了分子伴侣纯化的研究。超大孔纤维素填充柱对分子伴侣具有较高的吸附容量。流速对超大孔介质的分离能力的影响很小,在高流速下仍然可以分离得到纯度较高的目标产物。结果进一步证明该超大孔纤维素色谱介质可用于高速蛋白质色谱分离纯化。
The thesis focuses on the fabrication of superporous cellulose bead for high-speed protein chromatography and its application in the purification of proteins. The details in this work are summarized as follows:
A novel superporous cellulose (SC) matrix has been fabricated by water-in-oil emulsification-thermal regeneration using granules of calcium carbonate as porogenic agents. There were more irregular macropores and micropores scattering on the surface of the bead. The pore diffusivity of bovine serum albumin (BSA) in the diethylaminoehtyl-superporous cellulose (DEAE-SC) bead was two to three times larger than that in the diethylaminoehtyl-microporous cellulose (DEAE-MC) bead. In addition, the column packed with the DEAE-SC showed lower backpressure, higher column efficiency and dynamic binding capacity than the column packed with the DEAE-MC at a flow rate range of 150 to 900 cm/h.
To enhance the adsorption capacity of BSA to DEAE-SC adsorbents, we fabiricated the grafted superporous cellulose beads using ceric ammonium nitrate as initiator. Infrared spectral analysis confirmed the grafting reaction. The adsorption capacity of BSA to the diethylaminoehtyl-grfted- superporous cellulose (DEAE-G-SC) was over two times increased as compared to the ungrafted DEAE-SC and no nonspecific adsorption of BSA was observed. The grafted polymer chains led to the decrease of flowthrough pores, so the convective mass transport in the particles was significantly reduced, leading to the decrease of column efficiency with flow velocity. Despite this unfavorable effect, however, due to the high adsorption capacity of the grafted beads, the DEAE-G-SC column could still be operated at a higher DBC than the normal DEAE-SC column in the range of 150 to 600 cm/h.
The DEAE-SC beads and DEAE-MC beads were packed in a novel preparative electrochromatographic column to investigate the effect of elecric field on mass transfer. The preparative electrochromatographic column is constitutive with seven compartments which can provide two-dimensional (2D) electric field. The results showed that the two columns possessed higher dynamic binding capacity in the presence of 2D electric field, indicating which promote the mass transfer more efficiently. However, the effect of electric field on DEAE-MC column was more notable than DEAE-SC column. It is speculated that the mass transfer in DEAE-SC beads is superior compared with DEAE-MC beads.
To determine the property of the DEAE-SC matrix, a purification of GroEL from the cell debris of recombinant E. coil was carried out by the column packed with DEAE-SC beads and DEAE-MC beads, respectively. The results of breakthrough experiments showed that the dynamic binding capacity of the DEAE-SC column was higer than that of DEAE-SC column. Moreover, the purity of GroEL by DEAE-SC column with stepwise gradient elution was higher than that by DEAE-MC column, and dese not affect by the mobile phase velocity.
All the results exhibited the superior characteristics of the cellulose beads containing superpores as a potential medium for high-speed chromatography.
首先,以固体碳酸钙颗粒为致孔剂,采用热熔胶转化再生纤维的方法制备了超大孔纤维素介质。介质内含有超大孔和微孔结构。对牛血清白蛋白的相对孔隙率和有效孔扩散系数都得到了提高。其最高操作流速与刚性大孔介质相仿。孔内传质以对流传质为主。与微孔介质填充柱相比,超大孔介质填充柱的动态吸附容量和柱效都得到了提高,并且不受流速的影响。说明其在高速蛋白质色谱中具有很好的应用前景。
其次,为提高超大孔纤维素介质的吸附容量,采用硝酸铈胺(Ce~(4+))引发的方法,将甲基丙烯酸缩水甘油酯(GMA)接枝到超大孔纤维素介质的骨架上,制备了触角式超大孔纤维素介质。介质经接枝后,其静态吸附容量比接枝前介质的高2倍。接枝后,介质的孔径减小,孔内对流传质的程度降低。但是,触角式超大孔介质具有较高的静态吸附性能,因此仍然拥有较高的动态吸附容量。
为进一步研究超大孔纤维素介质的传质性能,在二维电场电色谱柱中,考察了电场对其传质性能的影响。电场的存在促进了介质孔内的传质,并且二维电场对传质的促进作用最强。与电场对超大孔介质填充柱传质性能的影响相比,电场对微孔介质的影响更显著,说明超大孔介质的传质性能比较好。并且实验还发现,随着流速的增大,二维电场对超大孔介质传质的促进作用增强,说明超大孔介质电色谱柱更适合在高流速下操作。
在超大孔纤维素介质的应用方面,开展了分子伴侣纯化的研究。超大孔纤维素填充柱对分子伴侣具有较高的吸附容量。流速对超大孔介质的分离能力的影响很小,在高流速下仍然可以分离得到纯度较高的目标产物。结果进一步证明该超大孔纤维素色谱介质可用于高速蛋白质色谱分离纯化。
The thesis focuses on the fabrication of superporous cellulose bead for high-speed protein chromatography and its application in the purification of proteins. The details in this work are summarized as follows:
A novel superporous cellulose (SC) matrix has been fabricated by water-in-oil emulsification-thermal regeneration using granules of calcium carbonate as porogenic agents. There were more irregular macropores and micropores scattering on the surface of the bead. The pore diffusivity of bovine serum albumin (BSA) in the diethylaminoehtyl-superporous cellulose (DEAE-SC) bead was two to three times larger than that in the diethylaminoehtyl-microporous cellulose (DEAE-MC) bead. In addition, the column packed with the DEAE-SC showed lower backpressure, higher column efficiency and dynamic binding capacity than the column packed with the DEAE-MC at a flow rate range of 150 to 900 cm/h.
To enhance the adsorption capacity of BSA to DEAE-SC adsorbents, we fabiricated the grafted superporous cellulose beads using ceric ammonium nitrate as initiator. Infrared spectral analysis confirmed the grafting reaction. The adsorption capacity of BSA to the diethylaminoehtyl-grfted- superporous cellulose (DEAE-G-SC) was over two times increased as compared to the ungrafted DEAE-SC and no nonspecific adsorption of BSA was observed. The grafted polymer chains led to the decrease of flowthrough pores, so the convective mass transport in the particles was significantly reduced, leading to the decrease of column efficiency with flow velocity. Despite this unfavorable effect, however, due to the high adsorption capacity of the grafted beads, the DEAE-G-SC column could still be operated at a higher DBC than the normal DEAE-SC column in the range of 150 to 600 cm/h.
The DEAE-SC beads and DEAE-MC beads were packed in a novel preparative electrochromatographic column to investigate the effect of elecric field on mass transfer. The preparative electrochromatographic column is constitutive with seven compartments which can provide two-dimensional (2D) electric field. The results showed that the two columns possessed higher dynamic binding capacity in the presence of 2D electric field, indicating which promote the mass transfer more efficiently. However, the effect of electric field on DEAE-MC column was more notable than DEAE-SC column. It is speculated that the mass transfer in DEAE-SC beads is superior compared with DEAE-MC beads.
To determine the property of the DEAE-SC matrix, a purification of GroEL from the cell debris of recombinant E. coil was carried out by the column packed with DEAE-SC beads and DEAE-MC beads, respectively. The results of breakthrough experiments showed that the dynamic binding capacity of the DEAE-SC column was higer than that of DEAE-SC column. Moreover, the purity of GroEL by DEAE-SC column with stepwise gradient elution was higher than that by DEAE-MC column, and dese not affect by the mobile phase velocity.
All the results exhibited the superior characteristics of the cellulose beads containing superpores as a potential medium for high-speed chromatography.
引文
[1] 孙彦,生物分离工程,北京:化学工业出版社,1998.
[2] 熊宗贵,生物技术制药,北京:高等教育出版社,1999.
[3] 徐炎华,欧阳平凯,韦萍,我国生物分离纯化技术现状及发展方向,江苏化工,1996,24:4.
[4] 傅若农, 顾峻岭,近代色谱分析,北京:国防工业出版社,1998.
[5] 邹汉法,张玉奎,卢佩章,高效液相色谱法,北京:科学出版社,1998.
[6] Locascio GA, Tigier HA, Batlle AMDC, Estimation of molecular weights of proteins by agarose gel filtration. J. Chromatogr., 1969, 40: 453-457.
[7] Ansari AA, Mage RG, Molecular-weight estimation of proteins using Sepharose CL-6B in guanidine hydrochloride. J. Chromatogr., 1977, 140: 98-102.
[8] Nilsson G, Nilsson K. Molecular-weight distribution determination of clinical dextran by gel permeation chromatography. J. Chromatogr., 1974, 101: 137-153.
[9] Fischer L. Gel filtration chromatography. Amsterdam: Elsevier, 1980.
[10] Moore S and Sten W H. Chromatography of amino acid on sulfonated polytyrede resins. J. Biol.Chem, 1951, 192: 663-681.
[11] 秦启宗,毛家骏,金忠浩等, 化学分离法,北京:原子能出版社,1984.
[12] Cuatrecasas P, Wilchek M. Single-step purification of avidin from egg white by affinity chromatography on biocytin-Sepharose column. Biochem Biophys Res Commun, 1968, 33: 235-239.
[13] Axen R, Poarath J, Ernback S, Chemical coupling of peptides and proteins to polysaccharides by means of cyanogens halides. Nature, 1967, 214: 1302-1304.
[14] Hjerten S, Some general aspects of hydrophobic interaction chromatography. J. Chromatogr., 1973, 87: 352-331.
[15] Liu Z, Yin G, Feng S-H et al., Oscillatory electroosmosis-enhanced intra/inter-particle liquid transport and its primary applications in the preparative electrochromatography of proteins. J. Chromatogr. A, 2001, 921: 93–98.
[16] Xue B, Sun Y, Fabrication and characterization of a rigid magnetic matrix for protein adsorption. J. Chromatogr. A, 2002, 921, 109–119.
[17] Mould DL, Synge RL M, Separations of polysaccharides related to starch by electrokinetic ultrafiltration in collodion membranes. J. Biochem., 1954, 58: 571-585.
[18] Pretorious JW, Hopkins BJ, Schieke JD, A new concept for high-speed liquid chromatography. J. Chromatogr., 1974, 99: 23-30.
[19] Jorgenson JW, Lukacs KD, High-resolution separations based on electrophoresis and electroosmosis. J. Chromatogr., 1981, 218: 209-216.
[20] Kevin DA, Overview of capillary electrophoresis and capillary electrochromatography. J. Chromatogr. A, 1999, 856: 443-463.
[21] 谭国民,横向交变电场中的蛋白质电色谱研究[博士论文],天津:天津大学,2005.
[22] Robson MM, Cikalo M G, Myers P, et al., Capillary electrochromatography: A review. J.Microcol. Sep., 1997, 9: 357-372.
[23] Knox JH, Grant IH, Miniaturisation in pressure and electroendosmotically driven liquid chromatography: some theoretical conderations. Chromatographia, 1987, 24: 135-143.
[24] Knox JH, Grant IH, Electrochromatography in packed tubes using 1.5 to 50 μm silica gels and ODS bonded silica gels. Chromatographia, 1991, 32: 317.
[25] Keim.C, Ladisch MR, New system for preparative electrochromatography of proteins. Biotechnol. Bioeng., 2000, 70:72-81.
[26] Ivory CF, Gobie WA, Continuous counteracting chromatographic electrophoresis. Biotechnol. Prog. 1990, 6: 21-32.
[27] Rathore AS, Reynolds KJ, Colón LA, Joule heating in packed capillaries used in capillary electrochromatography. Electrophoresis, 2002, 23: 2918-2928.
[28] Rudge SR, Basak SK, Ladisch MR, Solute retention in electrochromatography by electrically induced adsorption. AIChE J., 1993, 39(5):797-808.
[29] Basak SK, Ladisch MR, Mechanistic description and experimental studies of electrochromatography of proteins. AIChE J., 1995, 41(11): 2499-2507.
[30] Kenneth JR, Cole D, Heriberto Cabezas. Improved preparative electrochromatography column design. J. Chromatogr. A, 1997, 760: 259～263
[31] Tan GM, Shi QH, Sun Y, Retention behavior of proteins in size-exclusion electrachromatography with a low-voltage electric field perpendicular to the liquid phase streamline. Electrophoresis, 2005, 26: 3084-3093.
[32] Tan GM, Shi QH, Sun Y, Oscillatory transvers electric field enhances mass transger and protein capacity in ion-exchange electrochromatography. J. Chromatogr. A, 2005, 1098: 131-137.
[33] Tan GM, Dong XY, Sun Y, Oscillatory transvers electric field enhances protein resolution and capacity of size-exclusion chromatography. J. Sep. Sci., 2006,29: 684-690.
[34] Swingle SM, Tiselius A, Tricacium phosphate as an adsorbent in the chromatography of protein. J. Biochem., 1951, 48: 171-174.
[35] Sober HA, Peterson EA, Chromatography of proteins on cellulose ion-exchangers. J. Am. Chem. Soc., 1954, 76: 1711-1712.
[36] Lathe GH, Ruthven CRJ, The separation of substances and estimation of their relative molecular sizes by the use of columns of starch in water. J. Biochem., 1956, 62: 665-674.
[37] Porath J, Flodin P, Gel filtration: A method for desalting and group separation. Nature, 1959, 183: 1657-1659.
[38] Hjerten S, Chromatographic separation according to size of macromolecular and cell particles on columns of agarose suspensions. A. Biochem. Biophys., 1962, 99: 466-475.
[39] Unger KK, Kern R, Minou MC, High performance gel permeation chromatography with a new type of silica packing material. J. chromatogr., 1974, 99: 435-442.
[40] Regnier FE, Noel RJ, Glycerpropylsilane bonded phases in the steric exclusion chromatography of biological macromolecules. J. Chromatogr. Sci., 1976, 14: 316-320.
[41] Hashimoto T, Sasaki M, Aiura M, Kato Y, Molecular confirmation and crystal structure of the form of poly(ethylene oxybenzoate). J. Sci. Polym, Phys. Ed., 1978, 16: 19-89.
[42] Hjerten S, The preparation of agarose spheres for chromatography of molecules and particles. Biochem. Biophys. Acta, 1964, 79: 393-398.
[43] Bengtsson S, Philipson L, Chromatography of animal viruse on pearl-condensed agar. Biochem. Biophys. Acta, 1975, 79: 399-406.
[44] 杨之礼,苏茂尧,纤维素醚基础与应用,广州:华南理工大学出版社,1990.
[45] 唐爱民,粱文芷,纤维素的功能化,高分子通报,2000,4:1-9.
[46] Westman L, Lindstrom T, Swelling and mechanical properties of cellulose hydrogels I. preparation, characterization, and swelling behavior. J. Appl. Polym. Sci., 1981, 26: 2519-2532.
[47] Peska J, Stamberg J, Pelzbauer Z, Regenerated cellulose in the bead form after treatments and their effects on the porous structure of cellulose. Cell. Chem. Technol., 1978, 21:419-428.
[48] Stamberg J, Bead cellulose. Separation and Purification Methods, 1988, 17: 155-183.
[49] 朱伯儒,史作清,何炳林,球形纤维素吸附剂的制备和应用研究进展,精细化工,1996,13:42-46
[50] O’Neill JJ, Reichardt EP, Cellulose pellets. U.S.Pat., 2,543,928, 1951.
[51] 印寿根,李朝兴等,球状纤维素的制备及其应用,高分子通报,1996,2:100-104.
[52] Fujita M, Watanabe N, Satota N. Cellulose-based porous and spherical particles. Jan Kokai jokkyo koho Jp 03, 170, 501, 1991.
[53] Laszleiewicz B, Struszczyk H, Skorecka-Kubicka H, et al. Manufacture of micro crystalline cellulose. Pol. PL, 139, 735, 1987.
[54] Budschuh L, Farago J, Gimesi I. Method and apparatus for manufturing microporous and adsorbent. PCT Int. Appl. WO 9100, 14, 1989.
[55] Scarpa I, Beavins A, Stipanoic B, Cellulose chromatographic supports and method. US Pat. Appl. 818, 937, 1992.
[56] Lenfeld J, Peska J, Stamberg J. Preparation of porous bead cellulose with technical grain size angew. Makromol. Chem., 1992, 197: 201-206.
[57] OoKuma S, Igarashi K, Hara M, et al. Fine particles of porous ion-exchange cellulose process for their production and affinity capacity. Int. Appl. WO 89, 09, 651, 1989.
[58] Gemeiner P, Polakovic M, Mislovicova D, Stefuca V, Cellulose as a (bio)afftnity carrier: properties, design and applications. J. Chromatogr. B, 1998, 715: 245-271.
[59] Boeden HF, Pommerening K, Becker M, Rupprich C, Holtzhauer M, Loth F, Muller R, Bertram D, Bead cellulose derivatives as supports for immobilization and chromatographic purification of proteins. J. Chromatogr., 1991, 552: 389-414.
[60] Desgmukh, NR, Lali AM, Adsorptive purification of pDNA on superporous rigid cross-linked cellulose matrix. J. Chromatogr. B, 2005, 818: 5-10.
[61] 傅若农,高效液相色谱近年进展(上),国外分析仪器技术与应用,1997,2:10-20.
[62] Al?rio E. Rodrigues, Permeable packings and perfusion chromatography in protein separation. J. Chromatogr. B, 1997, 699: 47-61.
[63] Afeyan NB, Regnier FE, Dean RC. Perfusion chromatography. US Pat 5019270, 1991.
[64] Weeten KA, Tweeten TN. Reversed-phase chromatography of proteins on resin-based wide-pore packings. J. Chromatogr., 1986, 359: 111-119.
[65] Lehman ED, Joyce JG, Freymeyer DK et al, A novel process for the large-scale purification of recombinant tick anticoagulant peptide using perfusion chromatography. Bio. Technol., 1993, 11: 207-212.
[66] Lundell N, Markides K, Two-dimensional liquid chromatography of peptides: An optimization strategy. Chromatographia, 1992, 34(5-8): 369-375.
[67] Gadam SD, Cramer SM, Salt effects in anion exchange displacement chromatography: Comparison of pentosan polysulfate and dextran, sulfate displacers. Chromatographica, 1994, 39(7/8): 409-418.
[68] Gustavsson PE, Axelsson A, Larsson PO, Superporous agarose beads as a hydrophobic interaction chromatography support. J. Chromatogr. A, 1999, 830: 275-284.
[69] Hahn R, Panzer M, Hansen E, Mollerup J, Jungbauer A, Mass transfer properties of monoliths Sep. Sci. Technol. 2002, 37: 1545-1565.
[70] Hahn R, Jungbauer A, Peak broadening in protein chromatography with monoliths at very Fast separations. Anal. Chem. 2000, 72: 4853-4858.
[71] Cabrera K, Lubda D, A new monolithic - type HPLC column for fast separations, J. High Rcsol. J. High Resol. Chromatogr. 2000, 23, 93-99.
[72] Umemura T, Ueki Y, Tsunoda K, Katakai A, Tamada M, Haraguchi H, Preparation and characterization of methacrylate-based semi-micro monoliths for high-throughput bioanalysis. Anal Bioanal. Chem., 2006, 14, 566-571.
[73] Zhang ML, Sun Y., Poly (glycidyl methacrylate-divinylbenzene- triallylsocyanurate) continuous-bed protein chromatography. J. Chromatogr. A, 2001, 912, 31-38.
[74] Savina I, Galaev IY, Mattiasson B, Anion- exchange polymer brushes of N, N-dimethylaminoethyl-methacrylate. J. Chromatogr. A, 2005, 1092: 199-205.
[75] Podgornik R, Barut M, Strabcar A, Josic D, Koloini T, Construction of large-volume monolithic columns. Anal. Chem., 2000, 15: 5693-5699.
[76] Pflegerl K, Podgornik A, BergerE,Jungbauer A, Screening for peptide affinity ligands on CIM monoliths. Biotechnol. Bioeng., 2002, 79: 733-740
[77] Pflegerl K, Podgornik A ,Schallaun E, Jungbauer A, Direct Synthesis of Peptides on Convective Interaction Media Monolithic Columns for Affinity Chromatography J. Comb. Chem., 2002, 4(1): 33-37
[78] Du KF, Yang D, Sun Y, Fabrication of high-permeability and high-capacity monoliths for protein chromatography. J. Chromatogr. A, 2007, in press.
[79] Torres R, Pessela BCC, Fuentes M, Mateo C, Munilla AR, Fernandez-Lafuente R, Guisán JM, Supports coated with PEI as a new tool in chromatography. Enzyme and Microbial Technology, 2006, 39: 711-716.
[80] Mahdavinia GR, Zohuriaan-Mehr MJ, Pourjavadi A, Modified chitosan III, superabsorbency, salt- and pH-sensitivity of smart ampholytic hydrogels from chitosan-g-PAN. Polym. Adv. Technol. 2004, 15: 173-180.
[81] Kaneko Y, Nakamura S, Sakai K, Aoyagi T, Kikuchi A, Sakurai Y, Okano T, Rapid Deswelling Response of Poly(N-isopropylacrylamide) Hydrogels by the Formation of Water Release Channels Using Poly(ethylene oxide) Graft Chains. Macromolecules, 1998, 31: 6099-6105.
[82] Annaka M, Matsuura T, Kasai M, Nakahira T, Hara Y, Okano T, Preparation of comb-type N-isopropylacrylamide hydrogel beads and their application for size-selective separation media. Biomacromolecules, 2003, 4: 395-403.
[83] Kim SY, Cho SM, Lee YM, Kim SJ, Thermo- and pH-responsive behaviors of graft copolymer and blend based on chitosan and N-isopropylacrylamide. J Appl. Polym. SCi., 2000, 78: 1381-1391.
[84] 张兴英,程钰,赵京波,高分子化学,北京:化学工业出版社,2006.
[85] Choi SH, Kim MS, Ryoo JJ, Lee KP, Shin HD, Kim SH, Lee YH, Graft copolymer-metal complexes obtained by radiation grafting on polyethylene film. J. Appl. Polym. SCi., 2000, 77: 500-508.
[86] Choi SH, Park SY, Nho YC, Electrochemical properties of polyethylene membrane modified with carboxylic acid group.Radiat. Phys. Chem. 2000, 57: 179-186.
[87] Choi SH, Hwang YM, Ryoo JJ, Lee KP, Ohta K, Takeuchi T, Jin JY, Fujimoto C, Surfacegrafting of glycidyl methactylate on silica gel and polyethylene beads. Electrophoresis, 2003, 24: 3181-3186.
[88] Gupta KC, Sahoo S, Khandekar K, Graft copolymerization of ethyl acrylate onto cellulose using ceric ammonium nitrate as initiator in aqueous medium. Biomacromolecules, 2002, 3: 1087-1094.
[89] Cao YM, Qing XS, Sun J, Zhou FM, Lin SA, Graft copolymeization of acrylamide onto carboxymethyl starch. European Polymer J., 2002, 38: 1921-1924.
[90] Chowdhury P, Banerjrr M, Graft polymerization of methyl methacrylate onto polyvinyl alcohol using Ce4+ initiator. J. Appl. Polym. Sci., 1998, 70: 523-527.
[91] Müller E, Ionenaustauscher, EP 0722360 B1 1994.
[92] Müller E, Comparison between mass transfer properties of weak-anion- exchange resins with graft-functionalized polymer layers and traditional ungrafted resins. J. Chromatogr. A, 2003, 1006: 229-240.
[93] Ma ZY, Guan YP, Liu XQ, Liu HZ, Synthesis of magnetic chelator for high-capacity immobilized metal affinity adsorption of protein by cerium initiated graft polymerization. Langmuir, 2005, 21: 6987-6994.
[94] Rühe J, Surface-attached polyfunctional polymer networks for sensor chips. European Patent, 2002, No. 1176423.
[95] Ulbricht M, Yang H, Porous Polypropylene membranes with different carboxyl polymer brush layers for reversible protein binding via surface-initiated graft copolymerization. Chem. Mater., 2005, 17: 2622-2631.
[96] Kawai T, Saito K, Lee W, Protein binding to polymer brush, based on ion-exchange, hydrophobic, and affinity interactions. J. Chromatogr. B, 2003, 790: 131-142.
[97] Zhang ML, Sun Y. Cooperation of solid granule and solvent as porogenic agents - a novel porogenic mode of biporous media for protein chromatography. J. Chromatogr. A, 2001, 922: 77-81.
[98] Shi Y, Sun Y, Fabrication and characterization of a novel biporous spherical adsorbent for protein chromatography. Chromatographia 2003, 57: 29-35.
[99] Wu L, Bai S, Sun Y, Development of rigid bidisperse porous microspheres for high-speed protein chromatography. Biotechnol. Prog., 2003, 19: 1300-1306.
[100] Sun G.Y, Yang Z, Dong XY, Sun Y, Biporous polymeric beads fabricated by double emulsification for high-speed protein chromatography. J. Appl. Polym. Sci., 2007, 103: 17-23.
[101] Shi QH, Zhou X, Sun Y, A novel superorous agarose medium for high-speed protein chromatography. Biotechnol. Bioeng., 2005, 92: 643-651.
[102] Pharmacia AB, European Patent Application No.84850215.9, 1984
[103] Sundberg L, Porath J, Preparation of adsorbents for biospecific affinity chromatography: ?. Attachment of group-containing ligands to insoluble polymers by means of bifunctional oxiranes. J. Chromatogr., 1974, 90: 87-93.
[104] Toufik J, Labarre D, Relationship between reduction of complement activation by polysaccharide surfaces bearing diethylaminoethyl groups and their degree of substitution. Biomater., 1995, 16: 1081-1088.
[105] Bceden HF, Pommerening K, Becker M, et al, Bead cellulose derivatives as supports for innobilization and chromatographic purification of proteins. J. Chromatogr., 1991, 552: 389-414.
[106] Wu G, Brown GR, Adsorption of bilirubin by amine-containing polyacrylamide resins. React. Polym., 1991, 14: 49-61.
[107] Liapis I, McCoy MA, Theory of perfusion chromatography. J. Chromatogr., 1992, 599: 87-104.
[108] Bird RB, Stewart WE, Lightfoot EN, Transport Phenomea. Wiley, New York, 1960.
[109] Rengnier M, Vega de la, Chenou C, Loureiro JM, Rodrigues AE, Mass transfer mechanisms in Hyper D media for chromatographic protein separation. Biochem. Eng. J., 1998, 1: 11-23.
[110] 周鑫,新型蛋白质色谱介质的研究:[博士论文],天津;天津大学,2002.
[111] Zhang SP, Sun Y, Ionic strength dependence of protein adsorption to dye-ligandadsorbents, AIChE J. 2002, 48: 178-186.
[112] Tong XD, Dong XY, Sun Y, Lysozyme adsorption and purification by expanded bed chromatography with a small-sized dense adsorbent. Biochem. Eng. J., 2002, 12: 117-124 (8).
[113] Chen WD, Dong XY, Sun Y, Analysis of diffusion models for protein adsorption to porous anion-exchange adsorbent. J. Chromatogr. A, 2002, 962: 29-40.
[114] Chen WD, Dong XY, Bai S, Sun Y, Dependence of pore diffusivity of protein on adsorption density in anion-exchange adsorbent. Biochem. Eng. J., 2003, 14: 45-50.
[115] Zhou X, Xue B, Bai S, Sun Y, Macroporous polymeric ion exchange of high capacity for protein adsorption. Biochem. Eng. J., 2002, 11: 13-17.
[116] Lawrence E, Weaver Jr, Carta G, Protein adsorption on cation exchangers: comparison of macroporous and gel-composite media. Biotechnol. Prog., 1996, 12: 342-355.
[117] Carta G, Ubiera A, Particle-size distribution effects in batch adsorption. AIChE J., 2003, 49: 3066-3073.
[118] Afeyan NB, Fulton SP, Perfusion chromatography packing materials for proteins and peptides. J. Chromatogr., 1991, 544: 267-279.
[119] Afeyan NB, Gordon NF, Mazsaroff I, Varady L, Fulton SP, Flow-through particles for the high-performance liquid chromatographic separation of biomolecules: perfusion chromatography. J. Chromatogr., 1990, 519: 1-29.
[120] Sun GY, Shi QH, Sun Y, Novel biporous polymeric stationary phase for high-speed protein chromatography. J. Chromatogr. A, 2004, 1061: 159-165.
[121] Savina IN, Mattiasson B, Galaev IY, Graft polymerization of vinyl monomers inside macroporous polyacrylamide gel, cryogel, in aqueous and aqueous-organic media initiated by diperiodatocuprate (Ш) complexes. J. Polym. Sci.: Part A: Polymer Chemistry, 2006, 44: 1952-1963.
[122] Peppas NA, Huang Y, Torres-Lugo M, Ward JH, Zhang J, Physicochemical foundations and structural design of hydrogels in medicine and biology. Annu. Rev. Biomed. Eng., 2000, 2: 9-29.
[123] Shukla SR, Athalye AR, Graft-copolymerization of glycidyl methacrylate onto cotton cellulose. J. Appl. Polym. Sci., 1994, 54: 279-288.
[124] Fexby S, Bülow L, Hydrophobic peptide tags as tools in bioseparation. Trends Biotechnol., 2004, 22: 511-516.
[125] Gupta KC, Sahoo S, Khandekar K, Graft copolymerization of ethyl acrylate onto cellulose using ceric ammonium nitrate as initiator in aqueous medium. Biomacromolecules, 2002, 3: 1087-1094.
[126] Chowdhury P, Pal CM, Graft copolymerization of methyl acrylate onto polyvinyl alcohol using Ce (IV) initator. Europ. Polym. J., 1999, 35: 2207-2213.
[127] Zhang SP, Sun Y, Study on protein adsorption kinetics to a dye-ligand adsorption by the pore diffusion model. J. Chromatogr. A, 2002, 964: 35-46.
[128] Boschetti E, Advance sorbents for preparative protein separation purpous. J. Chromatogr. A, 1994, 658: 207-236.
[129] Chen GF, Pa?es M, Marek M, Zhang YK, Morgenstern AS, Tallarek U, Dynamic of capillary electrochromatography: experimental study of flow and transport in particulate beds. Chem. Eng. Technol., 2004, 27: 417-428.
[130] Cikalo MG, Bartle KD, Myers P, Influence of the electrical double-layer on electroosmotic flow in capillary electrochromatography. J. Chromatogr. A, 1999, 836: 35-51.
[131] Keim C, Ladisch M, Model for temperature profiles in large diameter electrochromatography column, AIChE J., 2003, 49: 402-410.
[132] Jia GD, Sun Y, Dye-ligand affinity electrochromatography with transverse and/or longitudinal electric fields, 2007, Sep. Purif. Technol. In .
[133] Laskey RA, Honda BM, Mills AD, et al, Nucleosomes are assembled by an acidic protein which binds his-tones and transfers them to DNA. Nature, 1978, 275: 416-421.
[134] Ellis RJ, Hemmingsen SM. Molecular chaper ones: proteins essential for the biogenes is of some macromolecular structures. Sci, 1989, 14: 339-344.
[135] 李剑,分子伴侣、蛋白质聚集和大分子拥挤环境对蛋白质折叠的影响:[博士学位论文],北京;中国科学院生物物理研究所,2001.
[136] 张森,金属硫蛋白、CroEL 及 DsbC 研究:1.金属硫蛋白通过锌离子的结合和释放调节氧化还苑从?2.一种非 ATP 酶依赖的 GroEL 的分子伴侣活力;3.大肠杆菌二硫键异构酶DsbC的折叠与去折叠[博士论文]. 北京: 中国科学院生物物理研究所,2001.
[137] Sigler PB, Xu Z, Rye HS, Burston SG, Fenton W A and Horwich AL. Structure and function in GroEL-mediated protein folding. Annu. Rev. Biochem., 1998, 67: 581-608.
[138] Braig K, Otwinowski Z, Hegde R, Boisvert DC, Joachimiak A, Horwich AL and Sigler PB. The crystal structure of the bacterial chaperonin GroEL at 2.8 ?. Nature, 1994, 371: 578-586.
[139] Xu Z, Horwich A L and Sigler PB. The crystal structure of the asymmeitric GroEL-GroES-(ADP)7 chaperonin complex. Nature, 1997, 388:741-750.
[140] Sakikawa C, Taguchi H, Makino Y and Yoshida M. On the maximum size of proteins to stay and fold in the cavity of GroEL underneath GroES. J. Biol. Chem. 1999, 274:21251-21256.
[141] Buchner J, Schmidt M, Fuchs M, et at., GroEL facilitates refolding of citrate synthase by suppressing aggregation, Biochemistry, 1991, 30: 1586-1591.
[142] Dong XD, Yang H, Sun Y. Lysozyme reactivation using immobilized molecular chaperonin GroEL. Biotechnol. Tech. 1999, 13:637-641.
[143] Dong XD, Wang Y B, Liu X G. Kinetic model of lysozyme renaturation with the molecular chaperone GroEL. Biotechnol. Lett. 2001, 23:1165-1169.
[144] Blennow A, Surin BP, Ehring H. Isolation and biochemical characterization of highly purified Escherichia coli molecular chaperone Cpn60 (GroEL) by affinity chromatography and urea-induced monomerization. Biochimica et Biophysica Acta, 1995, 1252:69-78.
[145] 佟晓冬. 流化床吸附分离蛋白质的基础研究:[博士学位论文],天津; 天津大学,2002.
[146] 苑巍,离子交换电色谱纯化蛋白质的研究:[硕士论文],天津;天津大学2005.
[147] Laemmli UK. Cleavage of structure proteins during the assembly of the head of bacteriophage. Nature, 1970, 227: 22962-22968.
[2] 熊宗贵,生物技术制药,北京:高等教育出版社,1999.
[3] 徐炎华,欧阳平凯,韦萍,我国生物分离纯化技术现状及发展方向,江苏化工,1996,24:4.
[4] 傅若农, 顾峻岭,近代色谱分析,北京:国防工业出版社,1998.
[5] 邹汉法,张玉奎,卢佩章,高效液相色谱法,北京:科学出版社,1998.
[6] Locascio GA, Tigier HA, Batlle AMDC, Estimation of molecular weights of proteins by agarose gel filtration. J. Chromatogr., 1969, 40: 453-457.
[7] Ansari AA, Mage RG, Molecular-weight estimation of proteins using Sepharose CL-6B in guanidine hydrochloride. J. Chromatogr., 1977, 140: 98-102.
[8] Nilsson G, Nilsson K. Molecular-weight distribution determination of clinical dextran by gel permeation chromatography. J. Chromatogr., 1974, 101: 137-153.
[9] Fischer L. Gel filtration chromatography. Amsterdam: Elsevier, 1980.
[10] Moore S and Sten W H. Chromatography of amino acid on sulfonated polytyrede resins. J. Biol.Chem, 1951, 192: 663-681.
[11] 秦启宗,毛家骏,金忠浩等, 化学分离法,北京:原子能出版社,1984.
[12] Cuatrecasas P, Wilchek M. Single-step purification of avidin from egg white by affinity chromatography on biocytin-Sepharose column. Biochem Biophys Res Commun, 1968, 33: 235-239.
[13] Axen R, Poarath J, Ernback S, Chemical coupling of peptides and proteins to polysaccharides by means of cyanogens halides. Nature, 1967, 214: 1302-1304.
[14] Hjerten S, Some general aspects of hydrophobic interaction chromatography. J. Chromatogr., 1973, 87: 352-331.
[15] Liu Z, Yin G, Feng S-H et al., Oscillatory electroosmosis-enhanced intra/inter-particle liquid transport and its primary applications in the preparative electrochromatography of proteins. J. Chromatogr. A, 2001, 921: 93–98.
[16] Xue B, Sun Y, Fabrication and characterization of a rigid magnetic matrix for protein adsorption. J. Chromatogr. A, 2002, 921, 109–119.
[17] Mould DL, Synge RL M, Separations of polysaccharides related to starch by electrokinetic ultrafiltration in collodion membranes. J. Biochem., 1954, 58: 571-585.
[18] Pretorious JW, Hopkins BJ, Schieke JD, A new concept for high-speed liquid chromatography. J. Chromatogr., 1974, 99: 23-30.
[19] Jorgenson JW, Lukacs KD, High-resolution separations based on electrophoresis and electroosmosis. J. Chromatogr., 1981, 218: 209-216.
[20] Kevin DA, Overview of capillary electrophoresis and capillary electrochromatography. J. Chromatogr. A, 1999, 856: 443-463.
[21] 谭国民,横向交变电场中的蛋白质电色谱研究[博士论文],天津:天津大学,2005.
[22] Robson MM, Cikalo M G, Myers P, et al., Capillary electrochromatography: A review. J.Microcol. Sep., 1997, 9: 357-372.
[23] Knox JH, Grant IH, Miniaturisation in pressure and electroendosmotically driven liquid chromatography: some theoretical conderations. Chromatographia, 1987, 24: 135-143.
[24] Knox JH, Grant IH, Electrochromatography in packed tubes using 1.5 to 50 μm silica gels and ODS bonded silica gels. Chromatographia, 1991, 32: 317.
[25] Keim.C, Ladisch MR, New system for preparative electrochromatography of proteins. Biotechnol. Bioeng., 2000, 70:72-81.
[26] Ivory CF, Gobie WA, Continuous counteracting chromatographic electrophoresis. Biotechnol. Prog. 1990, 6: 21-32.
[27] Rathore AS, Reynolds KJ, Colón LA, Joule heating in packed capillaries used in capillary electrochromatography. Electrophoresis, 2002, 23: 2918-2928.
[28] Rudge SR, Basak SK, Ladisch MR, Solute retention in electrochromatography by electrically induced adsorption. AIChE J., 1993, 39(5):797-808.
[29] Basak SK, Ladisch MR, Mechanistic description and experimental studies of electrochromatography of proteins. AIChE J., 1995, 41(11): 2499-2507.
[30] Kenneth JR, Cole D, Heriberto Cabezas. Improved preparative electrochromatography column design. J. Chromatogr. A, 1997, 760: 259～263
[31] Tan GM, Shi QH, Sun Y, Retention behavior of proteins in size-exclusion electrachromatography with a low-voltage electric field perpendicular to the liquid phase streamline. Electrophoresis, 2005, 26: 3084-3093.
[32] Tan GM, Shi QH, Sun Y, Oscillatory transvers electric field enhances mass transger and protein capacity in ion-exchange electrochromatography. J. Chromatogr. A, 2005, 1098: 131-137.
[33] Tan GM, Dong XY, Sun Y, Oscillatory transvers electric field enhances protein resolution and capacity of size-exclusion chromatography. J. Sep. Sci., 2006,29: 684-690.
[34] Swingle SM, Tiselius A, Tricacium phosphate as an adsorbent in the chromatography of protein. J. Biochem., 1951, 48: 171-174.
[35] Sober HA, Peterson EA, Chromatography of proteins on cellulose ion-exchangers. J. Am. Chem. Soc., 1954, 76: 1711-1712.
[36] Lathe GH, Ruthven CRJ, The separation of substances and estimation of their relative molecular sizes by the use of columns of starch in water. J. Biochem., 1956, 62: 665-674.
[37] Porath J, Flodin P, Gel filtration: A method for desalting and group separation. Nature, 1959, 183: 1657-1659.
[38] Hjerten S, Chromatographic separation according to size of macromolecular and cell particles on columns of agarose suspensions. A. Biochem. Biophys., 1962, 99: 466-475.
[39] Unger KK, Kern R, Minou MC, High performance gel permeation chromatography with a new type of silica packing material. J. chromatogr., 1974, 99: 435-442.
[40] Regnier FE, Noel RJ, Glycerpropylsilane bonded phases in the steric exclusion chromatography of biological macromolecules. J. Chromatogr. Sci., 1976, 14: 316-320.
[41] Hashimoto T, Sasaki M, Aiura M, Kato Y, Molecular confirmation and crystal structure of the form of poly(ethylene oxybenzoate). J. Sci. Polym, Phys. Ed., 1978, 16: 19-89.
[42] Hjerten S, The preparation of agarose spheres for chromatography of molecules and particles. Biochem. Biophys. Acta, 1964, 79: 393-398.
[43] Bengtsson S, Philipson L, Chromatography of animal viruse on pearl-condensed agar. Biochem. Biophys. Acta, 1975, 79: 399-406.
[44] 杨之礼,苏茂尧,纤维素醚基础与应用,广州:华南理工大学出版社,1990.
[45] 唐爱民,粱文芷,纤维素的功能化,高分子通报,2000,4:1-9.
[46] Westman L, Lindstrom T, Swelling and mechanical properties of cellulose hydrogels I. preparation, characterization, and swelling behavior. J. Appl. Polym. Sci., 1981, 26: 2519-2532.
[47] Peska J, Stamberg J, Pelzbauer Z, Regenerated cellulose in the bead form after treatments and their effects on the porous structure of cellulose. Cell. Chem. Technol., 1978, 21:419-428.
[48] Stamberg J, Bead cellulose. Separation and Purification Methods, 1988, 17: 155-183.
[49] 朱伯儒,史作清,何炳林,球形纤维素吸附剂的制备和应用研究进展,精细化工,1996,13:42-46
[50] O’Neill JJ, Reichardt EP, Cellulose pellets. U.S.Pat., 2,543,928, 1951.
[51] 印寿根,李朝兴等,球状纤维素的制备及其应用,高分子通报,1996,2:100-104.
[52] Fujita M, Watanabe N, Satota N. Cellulose-based porous and spherical particles. Jan Kokai jokkyo koho Jp 03, 170, 501, 1991.
[53] Laszleiewicz B, Struszczyk H, Skorecka-Kubicka H, et al. Manufacture of micro crystalline cellulose. Pol. PL, 139, 735, 1987.
[54] Budschuh L, Farago J, Gimesi I. Method and apparatus for manufturing microporous and adsorbent. PCT Int. Appl. WO 9100, 14, 1989.
[55] Scarpa I, Beavins A, Stipanoic B, Cellulose chromatographic supports and method. US Pat. Appl. 818, 937, 1992.
[56] Lenfeld J, Peska J, Stamberg J. Preparation of porous bead cellulose with technical grain size angew. Makromol. Chem., 1992, 197: 201-206.
[57] OoKuma S, Igarashi K, Hara M, et al. Fine particles of porous ion-exchange cellulose process for their production and affinity capacity. Int. Appl. WO 89, 09, 651, 1989.
[58] Gemeiner P, Polakovic M, Mislovicova D, Stefuca V, Cellulose as a (bio)afftnity carrier: properties, design and applications. J. Chromatogr. B, 1998, 715: 245-271.
[59] Boeden HF, Pommerening K, Becker M, Rupprich C, Holtzhauer M, Loth F, Muller R, Bertram D, Bead cellulose derivatives as supports for immobilization and chromatographic purification of proteins. J. Chromatogr., 1991, 552: 389-414.
[60] Desgmukh, NR, Lali AM, Adsorptive purification of pDNA on superporous rigid cross-linked cellulose matrix. J. Chromatogr. B, 2005, 818: 5-10.
[61] 傅若农,高效液相色谱近年进展(上),国外分析仪器技术与应用,1997,2:10-20.
[62] Al?rio E. Rodrigues, Permeable packings and perfusion chromatography in protein separation. J. Chromatogr. B, 1997, 699: 47-61.
[63] Afeyan NB, Regnier FE, Dean RC. Perfusion chromatography. US Pat 5019270, 1991.
[64] Weeten KA, Tweeten TN. Reversed-phase chromatography of proteins on resin-based wide-pore packings. J. Chromatogr., 1986, 359: 111-119.
[65] Lehman ED, Joyce JG, Freymeyer DK et al, A novel process for the large-scale purification of recombinant tick anticoagulant peptide using perfusion chromatography. Bio. Technol., 1993, 11: 207-212.
[66] Lundell N, Markides K, Two-dimensional liquid chromatography of peptides: An optimization strategy. Chromatographia, 1992, 34(5-8): 369-375.
[67] Gadam SD, Cramer SM, Salt effects in anion exchange displacement chromatography: Comparison of pentosan polysulfate and dextran, sulfate displacers. Chromatographica, 1994, 39(7/8): 409-418.
[68] Gustavsson PE, Axelsson A, Larsson PO, Superporous agarose beads as a hydrophobic interaction chromatography support. J. Chromatogr. A, 1999, 830: 275-284.
[69] Hahn R, Panzer M, Hansen E, Mollerup J, Jungbauer A, Mass transfer properties of monoliths Sep. Sci. Technol. 2002, 37: 1545-1565.
[70] Hahn R, Jungbauer A, Peak broadening in protein chromatography with monoliths at very Fast separations. Anal. Chem. 2000, 72: 4853-4858.
[71] Cabrera K, Lubda D, A new monolithic - type HPLC column for fast separations, J. High Rcsol. J. High Resol. Chromatogr. 2000, 23, 93-99.
[72] Umemura T, Ueki Y, Tsunoda K, Katakai A, Tamada M, Haraguchi H, Preparation and characterization of methacrylate-based semi-micro monoliths for high-throughput bioanalysis. Anal Bioanal. Chem., 2006, 14, 566-571.
[73] Zhang ML, Sun Y., Poly (glycidyl methacrylate-divinylbenzene- triallylsocyanurate) continuous-bed protein chromatography. J. Chromatogr. A, 2001, 912, 31-38.
[74] Savina I, Galaev IY, Mattiasson B, Anion- exchange polymer brushes of N, N-dimethylaminoethyl-methacrylate. J. Chromatogr. A, 2005, 1092: 199-205.
[75] Podgornik R, Barut M, Strabcar A, Josic D, Koloini T, Construction of large-volume monolithic columns. Anal. Chem., 2000, 15: 5693-5699.
[76] Pflegerl K, Podgornik A, BergerE,Jungbauer A, Screening for peptide affinity ligands on CIM monoliths. Biotechnol. Bioeng., 2002, 79: 733-740
[77] Pflegerl K, Podgornik A ,Schallaun E, Jungbauer A, Direct Synthesis of Peptides on Convective Interaction Media Monolithic Columns for Affinity Chromatography J. Comb. Chem., 2002, 4(1): 33-37
[78] Du KF, Yang D, Sun Y, Fabrication of high-permeability and high-capacity monoliths for protein chromatography. J. Chromatogr. A, 2007, in press.
[79] Torres R, Pessela BCC, Fuentes M, Mateo C, Munilla AR, Fernandez-Lafuente R, Guisán JM, Supports coated with PEI as a new tool in chromatography. Enzyme and Microbial Technology, 2006, 39: 711-716.
[80] Mahdavinia GR, Zohuriaan-Mehr MJ, Pourjavadi A, Modified chitosan III, superabsorbency, salt- and pH-sensitivity of smart ampholytic hydrogels from chitosan-g-PAN. Polym. Adv. Technol. 2004, 15: 173-180.
[81] Kaneko Y, Nakamura S, Sakai K, Aoyagi T, Kikuchi A, Sakurai Y, Okano T, Rapid Deswelling Response of Poly(N-isopropylacrylamide) Hydrogels by the Formation of Water Release Channels Using Poly(ethylene oxide) Graft Chains. Macromolecules, 1998, 31: 6099-6105.
[82] Annaka M, Matsuura T, Kasai M, Nakahira T, Hara Y, Okano T, Preparation of comb-type N-isopropylacrylamide hydrogel beads and their application for size-selective separation media. Biomacromolecules, 2003, 4: 395-403.
[83] Kim SY, Cho SM, Lee YM, Kim SJ, Thermo- and pH-responsive behaviors of graft copolymer and blend based on chitosan and N-isopropylacrylamide. J Appl. Polym. SCi., 2000, 78: 1381-1391.
[84] 张兴英,程钰,赵京波,高分子化学,北京:化学工业出版社,2006.
[85] Choi SH, Kim MS, Ryoo JJ, Lee KP, Shin HD, Kim SH, Lee YH, Graft copolymer-metal complexes obtained by radiation grafting on polyethylene film. J. Appl. Polym. SCi., 2000, 77: 500-508.
[86] Choi SH, Park SY, Nho YC, Electrochemical properties of polyethylene membrane modified with carboxylic acid group.Radiat. Phys. Chem. 2000, 57: 179-186.
[87] Choi SH, Hwang YM, Ryoo JJ, Lee KP, Ohta K, Takeuchi T, Jin JY, Fujimoto C, Surfacegrafting of glycidyl methactylate on silica gel and polyethylene beads. Electrophoresis, 2003, 24: 3181-3186.
[88] Gupta KC, Sahoo S, Khandekar K, Graft copolymerization of ethyl acrylate onto cellulose using ceric ammonium nitrate as initiator in aqueous medium. Biomacromolecules, 2002, 3: 1087-1094.
[89] Cao YM, Qing XS, Sun J, Zhou FM, Lin SA, Graft copolymeization of acrylamide onto carboxymethyl starch. European Polymer J., 2002, 38: 1921-1924.
[90] Chowdhury P, Banerjrr M, Graft polymerization of methyl methacrylate onto polyvinyl alcohol using Ce4+ initiator. J. Appl. Polym. Sci., 1998, 70: 523-527.
[91] Müller E, Ionenaustauscher, EP 0722360 B1 1994.
[92] Müller E, Comparison between mass transfer properties of weak-anion- exchange resins with graft-functionalized polymer layers and traditional ungrafted resins. J. Chromatogr. A, 2003, 1006: 229-240.
[93] Ma ZY, Guan YP, Liu XQ, Liu HZ, Synthesis of magnetic chelator for high-capacity immobilized metal affinity adsorption of protein by cerium initiated graft polymerization. Langmuir, 2005, 21: 6987-6994.
[94] Rühe J, Surface-attached polyfunctional polymer networks for sensor chips. European Patent, 2002, No. 1176423.
[95] Ulbricht M, Yang H, Porous Polypropylene membranes with different carboxyl polymer brush layers for reversible protein binding via surface-initiated graft copolymerization. Chem. Mater., 2005, 17: 2622-2631.
[96] Kawai T, Saito K, Lee W, Protein binding to polymer brush, based on ion-exchange, hydrophobic, and affinity interactions. J. Chromatogr. B, 2003, 790: 131-142.
[97] Zhang ML, Sun Y. Cooperation of solid granule and solvent as porogenic agents - a novel porogenic mode of biporous media for protein chromatography. J. Chromatogr. A, 2001, 922: 77-81.
[98] Shi Y, Sun Y, Fabrication and characterization of a novel biporous spherical adsorbent for protein chromatography. Chromatographia 2003, 57: 29-35.
[99] Wu L, Bai S, Sun Y, Development of rigid bidisperse porous microspheres for high-speed protein chromatography. Biotechnol. Prog., 2003, 19: 1300-1306.
[100] Sun G.Y, Yang Z, Dong XY, Sun Y, Biporous polymeric beads fabricated by double emulsification for high-speed protein chromatography. J. Appl. Polym. Sci., 2007, 103: 17-23.
[101] Shi QH, Zhou X, Sun Y, A novel superorous agarose medium for high-speed protein chromatography. Biotechnol. Bioeng., 2005, 92: 643-651.
[102] Pharmacia AB, European Patent Application No.84850215.9, 1984
[103] Sundberg L, Porath J, Preparation of adsorbents for biospecific affinity chromatography: ?. Attachment of group-containing ligands to insoluble polymers by means of bifunctional oxiranes. J. Chromatogr., 1974, 90: 87-93.
[104] Toufik J, Labarre D, Relationship between reduction of complement activation by polysaccharide surfaces bearing diethylaminoethyl groups and their degree of substitution. Biomater., 1995, 16: 1081-1088.
[105] Bceden HF, Pommerening K, Becker M, et al, Bead cellulose derivatives as supports for innobilization and chromatographic purification of proteins. J. Chromatogr., 1991, 552: 389-414.
[106] Wu G, Brown GR, Adsorption of bilirubin by amine-containing polyacrylamide resins. React. Polym., 1991, 14: 49-61.
[107] Liapis I, McCoy MA, Theory of perfusion chromatography. J. Chromatogr., 1992, 599: 87-104.
[108] Bird RB, Stewart WE, Lightfoot EN, Transport Phenomea. Wiley, New York, 1960.
[109] Rengnier M, Vega de la, Chenou C, Loureiro JM, Rodrigues AE, Mass transfer mechanisms in Hyper D media for chromatographic protein separation. Biochem. Eng. J., 1998, 1: 11-23.
[110] 周鑫,新型蛋白质色谱介质的研究:[博士论文],天津;天津大学,2002.
[111] Zhang SP, Sun Y, Ionic strength dependence of protein adsorption to dye-ligandadsorbents, AIChE J. 2002, 48: 178-186.
[112] Tong XD, Dong XY, Sun Y, Lysozyme adsorption and purification by expanded bed chromatography with a small-sized dense adsorbent. Biochem. Eng. J., 2002, 12: 117-124 (8).
[113] Chen WD, Dong XY, Sun Y, Analysis of diffusion models for protein adsorption to porous anion-exchange adsorbent. J. Chromatogr. A, 2002, 962: 29-40.
[114] Chen WD, Dong XY, Bai S, Sun Y, Dependence of pore diffusivity of protein on adsorption density in anion-exchange adsorbent. Biochem. Eng. J., 2003, 14: 45-50.
[115] Zhou X, Xue B, Bai S, Sun Y, Macroporous polymeric ion exchange of high capacity for protein adsorption. Biochem. Eng. J., 2002, 11: 13-17.
[116] Lawrence E, Weaver Jr, Carta G, Protein adsorption on cation exchangers: comparison of macroporous and gel-composite media. Biotechnol. Prog., 1996, 12: 342-355.
[117] Carta G, Ubiera A, Particle-size distribution effects in batch adsorption. AIChE J., 2003, 49: 3066-3073.
[118] Afeyan NB, Fulton SP, Perfusion chromatography packing materials for proteins and peptides. J. Chromatogr., 1991, 544: 267-279.
[119] Afeyan NB, Gordon NF, Mazsaroff I, Varady L, Fulton SP, Flow-through particles for the high-performance liquid chromatographic separation of biomolecules: perfusion chromatography. J. Chromatogr., 1990, 519: 1-29.
[120] Sun GY, Shi QH, Sun Y, Novel biporous polymeric stationary phase for high-speed protein chromatography. J. Chromatogr. A, 2004, 1061: 159-165.
[121] Savina IN, Mattiasson B, Galaev IY, Graft polymerization of vinyl monomers inside macroporous polyacrylamide gel, cryogel, in aqueous and aqueous-organic media initiated by diperiodatocuprate (Ш) complexes. J. Polym. Sci.: Part A: Polymer Chemistry, 2006, 44: 1952-1963.
[122] Peppas NA, Huang Y, Torres-Lugo M, Ward JH, Zhang J, Physicochemical foundations and structural design of hydrogels in medicine and biology. Annu. Rev. Biomed. Eng., 2000, 2: 9-29.
[123] Shukla SR, Athalye AR, Graft-copolymerization of glycidyl methacrylate onto cotton cellulose. J. Appl. Polym. Sci., 1994, 54: 279-288.
[124] Fexby S, Bülow L, Hydrophobic peptide tags as tools in bioseparation. Trends Biotechnol., 2004, 22: 511-516.
[125] Gupta KC, Sahoo S, Khandekar K, Graft copolymerization of ethyl acrylate onto cellulose using ceric ammonium nitrate as initiator in aqueous medium. Biomacromolecules, 2002, 3: 1087-1094.
[126] Chowdhury P, Pal CM, Graft copolymerization of methyl acrylate onto polyvinyl alcohol using Ce (IV) initator. Europ. Polym. J., 1999, 35: 2207-2213.
[127] Zhang SP, Sun Y, Study on protein adsorption kinetics to a dye-ligand adsorption by the pore diffusion model. J. Chromatogr. A, 2002, 964: 35-46.
[128] Boschetti E, Advance sorbents for preparative protein separation purpous. J. Chromatogr. A, 1994, 658: 207-236.
[129] Chen GF, Pa?es M, Marek M, Zhang YK, Morgenstern AS, Tallarek U, Dynamic of capillary electrochromatography: experimental study of flow and transport in particulate beds. Chem. Eng. Technol., 2004, 27: 417-428.
[130] Cikalo MG, Bartle KD, Myers P, Influence of the electrical double-layer on electroosmotic flow in capillary electrochromatography. J. Chromatogr. A, 1999, 836: 35-51.
[131] Keim C, Ladisch M, Model for temperature profiles in large diameter electrochromatography column, AIChE J., 2003, 49: 402-410.
[132] Jia GD, Sun Y, Dye-ligand affinity electrochromatography with transverse and/or longitudinal electric fields, 2007, Sep. Purif. Technol. In .
[133] Laskey RA, Honda BM, Mills AD, et al, Nucleosomes are assembled by an acidic protein which binds his-tones and transfers them to DNA. Nature, 1978, 275: 416-421.
[134] Ellis RJ, Hemmingsen SM. Molecular chaper ones: proteins essential for the biogenes is of some macromolecular structures. Sci, 1989, 14: 339-344.
[135] 李剑,分子伴侣、蛋白质聚集和大分子拥挤环境对蛋白质折叠的影响:[博士学位论文],北京;中国科学院生物物理研究所,2001.
[136] 张森,金属硫蛋白、CroEL 及 DsbC 研究:1.金属硫蛋白通过锌离子的结合和释放调节氧化还苑从?2.一种非 ATP 酶依赖的 GroEL 的分子伴侣活力;3.大肠杆菌二硫键异构酶DsbC的折叠与去折叠[博士论文]. 北京: 中国科学院生物物理研究所,2001.
[137] Sigler PB, Xu Z, Rye HS, Burston SG, Fenton W A and Horwich AL. Structure and function in GroEL-mediated protein folding. Annu. Rev. Biochem., 1998, 67: 581-608.
[138] Braig K, Otwinowski Z, Hegde R, Boisvert DC, Joachimiak A, Horwich AL and Sigler PB. The crystal structure of the bacterial chaperonin GroEL at 2.8 ?. Nature, 1994, 371: 578-586.
[139] Xu Z, Horwich A L and Sigler PB. The crystal structure of the asymmeitric GroEL-GroES-(ADP)7 chaperonin complex. Nature, 1997, 388:741-750.
[140] Sakikawa C, Taguchi H, Makino Y and Yoshida M. On the maximum size of proteins to stay and fold in the cavity of GroEL underneath GroES. J. Biol. Chem. 1999, 274:21251-21256.
[141] Buchner J, Schmidt M, Fuchs M, et at., GroEL facilitates refolding of citrate synthase by suppressing aggregation, Biochemistry, 1991, 30: 1586-1591.
[142] Dong XD, Yang H, Sun Y. Lysozyme reactivation using immobilized molecular chaperonin GroEL. Biotechnol. Tech. 1999, 13:637-641.
[143] Dong XD, Wang Y B, Liu X G. Kinetic model of lysozyme renaturation with the molecular chaperone GroEL. Biotechnol. Lett. 2001, 23:1165-1169.
[144] Blennow A, Surin BP, Ehring H. Isolation and biochemical characterization of highly purified Escherichia coli molecular chaperone Cpn60 (GroEL) by affinity chromatography and urea-induced monomerization. Biochimica et Biophysica Acta, 1995, 1252:69-78.
[145] 佟晓冬. 流化床吸附分离蛋白质的基础研究:[博士学位论文],天津; 天津大学,2002.
[146] 苑巍,离子交换电色谱纯化蛋白质的研究:[硕士论文],天津;天津大学2005.
[147] Laemmli UK. Cleavage of structure proteins during the assembly of the head of bacteriophage. Nature, 1970, 227: 22962-22968.