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HWAHAK KONGHAK,
Vol.39, No.6, 705-708, 2001
Vol.39, No.6, 705-708, 2001
HPMC와 GFC에 의한 유청 단백질의 분리
Separation of Whey Proteins by High Performance Membrane Chromatography and Gel Filtration Chromatography
본 연구에서는 유청 단백질 중에서 α-lactalbumin와 β-lactoglobulin을 HPMC(High Performance Membrane Chromatography)와 GFC(Gel Filtration Chromatography)를 이용하여 분리하였다. 이동상 조성과 구배용매 조성법에 따른 각기 유청 단백질의 분리도를 고찰하여 최적 이동상을 구하였다. HPMC의 분리 메카니즘은 음이온 교환작용이며 고정상은 CIM DEAE, QA disk(16 x 3 mm), 이동상은 buffer A(20 mM Tris-HCI, pH 7.3)와 buffer B(buffer A+1 M NaCl)을 사용하였다. 최적 이동상은 Buffer A/Buffer B=100/0-30/70 vol%, gradient time 1 min, 30/70-10/90 vol%, gradient time 2 min을 실험적으로 얻었고, 4 ml/min의 이동상 유속에서 5분내에 α-lactalbumin와 β-lactoglobulin를 분리할 수 있었다. 분자량의 차이가 분리 메카니즘인 GFC에 의해서는 이동상으로 물(100%)인 일정용매 조성법과 buffer A(water+0.1%v/v TFA)와 buffer B(ACN+0.1%v/v TFA)를 사용하여 구배용매 조성법으로 분리하였다. 음이온교환 메카니즘에 의한 HPMC가 크기배제의 원리에 의한 GFC보다 α-lactalbumin와 β-lactoglobulin를 더 잘 분리하였다.
α-lactalbumin and β-lactoglobulin in whey proteins were separated by HPMC(High Performance Membrane Chromatography) and GFC(Gel Filtration Chromatography). The separation mechanism of HPMC was anion-exchange, and the stationary phases were anion CIM(Convective Interaction Media) DEAE and QA disks(16 x 3 mm). Two types of mobile phase were used, buffer A(20 mM Tris-HCI, pH 7.3) and buffer B(buffer A+1 M NaCl). The optimum mobile phase and operating condition were buffer A/Buffer B=100/0-30/70 vol%, gradient time 1 min, 30/70-10/90 vol%, gradient time 2 min, In this experimental condition, α-lactalbumin and β-lactoglobulin were separated within 5 min at a mobile phase flow rate of 4 ml/min. In GFC, characterized by size exclusion, water in isocratic mode and water with TFA in gradient mode were utilized as mobile phases, Separation of the two whey proteins was performed better by HPMC than by GFC.
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