The Factors Affecting the Backward-Transfer of Bovine Serum Albumin (BSA) from Sodium Bis(2-ethylhexyl) Sulfosuccinate (AOT) Reverse Micellar Solutions

Seon-Gyun Rho; Choon-Hyoung Kang

Korean Journal of Chemical Engineering, Vol.20, No.3, 517-521, May, 2003

Seon-Gyun Rho, and Choon-Hyoung Kang^†

Faculty of Chemical Engineering, Chonnam National University, Gwangju 500-757, Korea

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The factors affecting the back-extraction efficiency of Bovine Serum Albumin (BSA, 65kDa, pI 4.9) solubilized in an AOT reverse micellar solution, prepared by the injection method, to an excess aqueous phase were investigated. In particular, effects of pH, the type of salt and its concentration in the excess aqueous phase were examined. Furthermore, by comparing CD spectra of the back-extracted BSA with the feed BSA, the structural changes of the protein during the extraction process were determined. The addition of 1 : 1 salt such as KCl or NaCl to the aqueous phase resulted in a 100% recovery of the protein to the aqueous phase at a pH higher than its isoelectric point pI. This high efficiency of the back-extraction might be due to the change in the interactions between the protein and micellar aggregates driven by the added salt. For 1 : 2 salts like MgCl2 or CaCl2, BSA was back extracted with lower than 20% extraction efficiency. Maximum efficiencies were achieved at about pH=7 and pH=8 for monovalent and divalent salts, respectively. From the CD spectra of back-extracted BSA, it was observed that denaturation of BSA was not significant during the extraction process.

Keywords:Reverse Micelle;AOT;Injection Method;Back-Extraction;BSA

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[Related Articles]

[1] De TK, Maitra A, Adv. Colloid Interface Sci., 59, 95 (1995)

[2] Dekker M, Hilhorst R, Leane C, Anal. Biochem., 178, 217 (1989)

[3] Dekker M, Leser ME, "Highly Selective Separations in Biotechnology," Chapman & Hall, London (1994)

[4] Goklen KE, Hatton TA, Sep. Sci. Technol., 22, 831 (1987)

[5] Gupta RB, Han CJ, Johnston KP, Biotechnol. Bioeng., 44(7), 830 (1994)

[6] Hilhorst R, Sergeeva M, Heering D, Rietveld P, Fijneman P, Wolbert RB, Dekker M, Bijsterbosch BH, Biotechnol. Bioeng., 46(4), 375 (1995)

[7] Hong DP, Kuboi R, Komasawa I, Korean J. Chem. Eng., 14(5), 334 (1997)

[8] Hong DP, Lee SK, Kuboi R, J. Chromatogr. B, 743, 203 (2000)

[9] Jolivalt C, Miner M, Renon H, "Downstream Processing and Bioprocessing; Recovery and Purification of Biological Products," ACS Symposium series 419, Hamel, J.P., Hunter, J.B. and Sikdar, S.K. eds., American Chemical Society, Washington D.C. (1990)

[10] Jolivalt C, Miner M, Renon H, J. Colloid Interface Sci., 135, 85 (1990)

[11] Kadam KL, Enzyme Microb. Technol., 8, 266 (1986)

[12] Krei GA, Hustedt H, Chem. Eng. Sci., 47, 99 (1992)

[13] Leser ME, Mrkoci K, Biotechnol. Bioeng., 41, 489 (1993)

[14] Marcozz G, Correa N, Luisi PL, Caselli M, Biotechnol. Bioeng., 38, 1239 (1991)

[15] Mitchell DJ, Ninham BW, J. Chem. Soc.-Faraday Trans., 77, 601 (1981)

[16] Nagahama K, Noritomi H, Koyama A, Fluid Phase Equilib., 116(1-2), 126 (1996)

[17] Rabie HR, Helou D, Weber ME, Vera JH, J. Colloid Interface Sci., 189(2), 208 (1997)

[18] Rabie HR, Vera JH, Fluid Phase Equilib., 122(1-2), 169 (1996)

[19] Shiomori K, Ebuchi N, Kawano Y, Kuboi R, Komasawa I, J. Ferment. Bioeng., 86(6), 581 (1998)

[20] Shiomori K, Kawano Y, Kuboi R, Komasawa I, J. Chem. Eng. Jpn., 28(6), 803 (1995)

[21] Wolbert RBG, Hilhorst R, Voskuilen G, Nachtegaal H, Dekker M, Vant Riet K, Bijsterbosch BH, Eur. J. Biochem., 184, 627 (1989)