특성화된 poly(hydroxyethyl-methacrylate) 비드를 이용한 human immunoglobulin G의 흡착분리에 관한 연구(I) - Thiophilic poly(hydroxyethyl-methacryate) 비드의 제조 -

임재홍; 장우진; 장순호; 조영일

HWAHAK KONGHAK, Vol.37, No.2, 330-335, April, 1999

A Study on Adsorptive Separation of Human Immunoglobulin G with the Characterized Poly (hydroxyethyl-methacrylate) Beads (I) - Preparation of Thiophilic Poly (hydroxyethyl-methacrylate) Beads -

임재홍, 장우진, 장순호, 조영일

초록

Divinylsulfone(DVS)과 β-mercaptoethanol을 이용한 일련의 반응을 통해, 흡착담체 조건에 맞게 제조된 poly(hydroxyethyl-methacrylate)(PHEMA) 비드를 thiophilic-특성화하고자 하였다. 실험변수들은 Na₂CO₃ 농도, pH, 반응시간 그리고 각 반응물의 농도로 하였으며, 반응물의 결합량이 최대값을 얻을 때 변수를 고정하였다. DVS를 통한 PHEMA의 활성화 반응에 있어서, PHEMA비드들의 가교현상없이 얻을 수 있는 최대 결합랑은 Na₂CO₃ 0.5 mol/L, pH 12.5, 반응 시간 8 시간 그리고 DVS 2 mol/L에서 약 16.2mmol DVS/g PHEMA이었다. 또한 활성화된 PHEMA(APHEMA)와 β-mercaptoethanol을 반응시킨 결합반응에서는 Na₂CO₃ 0.25 mol/L, pH 9, 반응 시간 36시간 그리고 β-mercaptoethanol 4 mol/L에서 최대값인 약 514.4 μmol β-mercaptoethanol/g APHEMA를 얻었다.

PoIy(hydroxyethyl-methacrylate)(PHEMA) beads, prepared in accordance with the standards of adsorptive matrix, were tried to have the thiophilic characters by a series of reactions with divinylsulfone(DVS) and -mercaptoethanol. The experiments parameters were Na₂CO₃ concentration, pH, reaction time and the concentration of the reactants. When the bound mass was maximum, the parameter fixed. In the activation step of PHEMA with DVS, the maximum bound quantity of DVS was 16.2mmol DVS/g PHEMA at Na₂CO₃ 0.5 mol/L pH 12.5, reaction time 8hr and DVS 2mol/L. And in the coupling reaction with the activated PHEMA(APHEMA) and -mercaptoethanol, the fixed parameters were Na₂CO₃ 0.25 mol/L, pH 9, reaction time 36 hr and -mercaptoethanol 4 mol/L for the upmost mass of about 514.4 mol -mercaptoethanol/g APHEMA.

Keywords:Thiophilic Adsorbents;polyHEMA Beads;Divinylsulfone;9-Mercaptoethanol

[References]

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Lihme A, Heegaard MH, Anal. Biochem., 192, 64 (1991)
Konecny P, Brown RJ, Scouten WH, J. Chromatogr. A, 673, 45 (1994)
Huchens TW, Magnuson JS, Yip TT, J. Immunol. Methods, 128, 89 (1990)
Sulk B, Birkenmeier G, Kopperschla G, J. Immunol. Methods, 149, 165 (1992)
Hutchens TW, Porath J, Anal. Biochem., 159, 217 (1986)
Porath J, Belew M, Trends Biotechnol., 5, 225 (1987)
Finger UB, Brummer W, Kneips E, Thomames J, Kula MR, J. Chromatogr. B, 675, 197 (1996)
Hermanson GT, "Immobilized Affinity Ligand Techniques," Academic Press Inc., San Diego (1992)
Turkova J, "Bioaffinity Chromatography," Elsevier, Amsterdam (1993)
Skely PJ, Tighe BJ, Polym. Commun., 20, 1051 (1979)
Piskin E, Int. J. Artif. Org., 9, 401 (1986)
Horak D, Svec F, Kalal J. Adamyan A, Skuba N, Titova M, Dan V, Varava B, Trostenyuk N, Vorankova O, Gumargalieava K, Timochina V, Biomaterials, 9, 367 (1988)
Im JH, Park JH, Jang SH, Han HS, Joe YI, HWAHAK KONGHAK, 36(5), 669 (1998)
Im JH, Jang WJ, Kim HS, Chun KY, Han HS, Joe YI, HWAHAK KONGHAK, 36(6), 887 (1998)
Porath J, Laas T, Janson J, J. Chromatogr., 103, 49 (1975)
Oscarsson S, Porath J, Anal. Biochem., 176, 330 (1989)
Marcus SL, Balbinder E, Anal. Biochem., 48, 448 (1972)
Knudsen KL, Hansen MB, Henriksen LR, Andersen BK, Lihme A, Anal. Biochem., 201, 170 (1992)
Noel RJ, O'Hare WT, Street G, J. Chromatogr. A, 734, 241 (1996)
Lowe CR, Dean PDG, "Affinity Chromatography," John Wiley & Sons, London (1974)
Porath J, Sunbdberg L, Nature New Biol., 238, 261 (1972)
Solomons TWG, "Organic Chemistry 6th edit," John Wiley & Sons, Inc., New York (1996)

[Referenced By]

[1] Porath J, Maisano F, Belew M, FEBS, 185(2), 306 (1985)

[2] Lihme A, Heegaard MH, Anal. Biochem., 192, 64 (1991)

[3] Konecny P, Brown RJ, Scouten WH, J. Chromatogr. A, 673, 45 (1994)

[4] Huchens TW, Magnuson JS, Yip TT, J. Immunol. Methods, 128, 89 (1990)

[5] Sulk B, Birkenmeier G, Kopperschla G, J. Immunol. Methods, 149, 165 (1992)

[6] Hutchens TW, Porath J, Anal. Biochem., 159, 217 (1986)

[7] Porath J, Belew M, Trends Biotechnol., 5, 225 (1987)

[8] Finger UB, Brummer W, Kneips E, Thomames J, Kula MR, J. Chromatogr. B, 675, 197 (1996)

[9] Hermanson GT, "Immobilized Affinity Ligand Techniques," Academic Press Inc., San Diego (1992)

[10] Turkova J, "Bioaffinity Chromatography," Elsevier, Amsterdam (1993)

[11] Skely PJ, Tighe BJ, Polym. Commun., 20, 1051 (1979)

[12] Piskin E, Int. J. Artif. Org., 9, 401 (1986)

[13] Horak D, Svec F, Kalal J. Adamyan A, Skuba N, Titova M, Dan V, Varava B, Trostenyuk N, Vorankova O, Gumargalieava K, Timochina V, Biomaterials, 9, 367 (1988)

[14] Im JH, Park JH, Jang SH, Han HS, Joe YI, HWAHAK KONGHAK, 36(5), 669 (1998)

[15] Im JH, Jang WJ, Kim HS, Chun KY, Han HS, Joe YI, HWAHAK KONGHAK, 36(6), 887 (1998)

[16] Porath J, Laas T, Janson J, J. Chromatogr., 103, 49 (1975)

[17] Oscarsson S, Porath J, Anal. Biochem., 176, 330 (1989)

[18] Marcus SL, Balbinder E, Anal. Biochem., 48, 448 (1972)

[19] Knudsen KL, Hansen MB, Henriksen LR, Andersen BK, Lihme A, Anal. Biochem., 201, 170 (1992)

[20] Noel RJ, O'Hare WT, Street G, J. Chromatogr. A, 734, 241 (1996)

[21] Lowe CR, Dean PDG, "Affinity Chromatography," John Wiley & Sons, London (1974)

[22] Porath J, Sunbdberg L, Nature New Biol., 238, 261 (1972)

[23] Solomons TWG, "Organic Chemistry 6th edit," John Wiley & Sons, Inc., New York (1996)