상 접촉법을 이용한 BSA의 AOT 역미셀으로 가용화: pH와 염의 영향

Solubilization of BSA into AOT Reverse Micelles Using the Phase-Transfer Method: Effects of pH and Salts

  • 노선균 (전남대학교 공과대학 화학공학과) ;
  • 강춘형 (고분자기술연구소)
  • 발행 : 2001.02.01

초록

상 접촉법을 이용하여 BSA를 음이온 계면활성제인 AOT와 isooctane으로 구성된 역미셀상으로 가용화하였다. 본 연구에서는 pH와 염의 종류 및 농도가 가용화 효율에 미치는 영향에 주목하였다. 0.1 M의 KCl, NaCl, $MgCl_2$, 그리고 $CaCl_2$와 같이 1:1 염 혹은 1:2 염을 수용액에 첨가하면 첨가한 염의 종류에 따라 pH=5~7사이에서 최대의 가용화 효율을 보였다. $CaCl_2$와 NaCl의 경우, 1 M 까지는 첨가한 염의 농도가 증가함에 따라 효율도 증가하였으나 $MgCl_2$의 경우에는 특이한 경향을 보였다. 염의 농도가 낮거나 pH가 낮은 경우에는 단백질이 광범위하게 수용액상과 유기상의 계면에서 침전되는 것은 주목할 부분이다. 역미셀의 water-pool 크기는 물에 대한 계면활성제의 몰비인 $W_0$를 측정하여 추산하였다. $MgCl_2$의 경우에는 pH변화에 관계없이 $W_0$=20의 거의 일정한 값을 보인 반면 $CaCl_2$와 KCl의 경우 염 농도가 증가함에 따라 $W_0$는 감소하였다.

Bovine serum albumin(BSA) was solubilized into the reverse micellar phase consisting of sodium bis(2-ethylhexyl) sulfosuccinate(AOT) and isooctane using the phase transfer method. Of particular interest in this study were the effects of pH and the added salt type and concentration on the solubilization efficiency. When univalent or divalent salts such as KCl, NaCl, $MgCl_2$, or $CaCl_2$ were added to the aqueous phase at a concentration of 0.1 M, maximum solubilization efficiency was attained at a pH ranging from 5 to 7, depending on the added salt type. Increased salt concentration up to 1 M resulted in an increased solubilization efficiency for $CaCl_2$ and NaCl, while the addition of $MgCl_2$ beyond 0.1 M showed an anomalous trend. Further, it was noteworthy that too a large extent the protein precipitated in the interface between the organic and aqueous phases at lower pHs and lower salt concentrations. The size of the reverse micelle water pool was estimated by measuring the molar ratio of the surfactant to the water, $W_0$. Irrespective of pH in the aqueous phase, the resulting value of $W_0$ was almost constant, eg., 20 for $MgCl_2$ . However, the value of $W_0$ decreased with increased salt concentration in the cases of KCl and $CaCl_2$.

키워드

참고문헌

  1. Science v.227 Separation techniques based on the opposition of two counteracting forces to produce a dynamic equilibrium O'Farrell P. H.
  2. Biotechnol. Prog. v.6 no.1 Continuous counteracting chromatographic electrophoresis Ivory C. F.;W. A. Gobie
  3. Chem. Eng. Sci. v.45 no.9 Mass Transfer Rate of Protein Extraction with Reversed Micelles Dekker M.;K. V. Piet;B. H. Bijsterbosch;P. Fijneman;R. Hilhorst
  4. Biotech. Bioeng. v.38 Protein Extraction by Reverse Micelles: A Study of the Factors Affection the Forward and Backward Transfer of alpha-Chymotrypsin and Its Activity Marcozz G.;N. Correa;P. L.Luisi;M. Casell
  5. Chem. Eng. Prog. Liquid-Liquid Extraction for Protein Separations Abbott N. L.;T. A. Hatton
  6. highly Selective Separations in Biotechnology Dekker M.;M. E. Leser
  7. Biotech. Bioeng. v.43 The Effect of pH and Ionic Strength on the Partitioning of Four Proteins in Reverse Micelles Systems Andrews B. A.;D. L. Pyle;J. A. Asenjo
  8. Enzyme Microb. Technol. v.8 Reverse micelles as a bioseparation tool Kadam K. L.
  9. J. Ferment. Bioeng. v.86 no.6 Extraction Characteristic of Bovine Serum Albumin Using Sodium Bis(2-ethyl-hexyl) Sulfosuccinate Reverse Micelles Shiomori K.;N. Ebuchi;Y. Kawano;R. Kuboi;I. Komasawa
  10. Biotech. Bioeng. v.39 Solubilizing Water Involved in Protein Extraction Using Reversed Micelles Ichikawa S.;M. Imai;M. Shimizu
  11. Biotech. Bioeng. v.47 Extraction of Lysozyne and Ribonucleasea Using Reverse Micelles: Limits to Protein Solubilization Lye G. J.;J. A. Asenjo;D. L. Pyle
  12. kor. J. Biotechnol. Bioeng. Spring Symposium Effect of pH and salt type for BSA Solubilization Using Reverse Micelles Rho S. G.;C. H. Kang;D. H. Park
  13. J. Chem. Eng. Japan. v.28 Effective purification method of large molecular weight proteins using conventional AOT reverse micelles Shiomori, K.;Kawano Y.;Kuboi, R.;Komasawa, I.
  14. J. Phys. Chem. v.92 Thermodynamics of the Uptake of Proteins by Reverse Micelles: A First Approximation Model Caselli M.;P. L. Luisi;M. Maestro;R. Roselli
  15. J. Phys. Chem. v.95 Structural Characterization of α-Chymotrypsin Containing AOT Reversed Micelles Rahaman. R. S.;T. A. Hatton
  16. Eur. J. Biochem. v.184 Protein transfer from an aqueous phase into reversed micelles : The effect of protein size and charge distribution Wolbert. R. B. G.;R. Hilhorst;G. Voskuilen;H. Nachtegaal;M. Dekker;K. Vantriet;B. H. Bijsterbosch
  17. J. Phys. Chem. v.92 Thermodynamic of the Uptake of Proteins by Reverse Micelles: Approximation Model M. Caselli M.;P. L. Luisi;M. Maestro;R. Roselli