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데옥시리보뉴클레오사이드를 분리하기 위한 최적 조건
Optimum Condition for Resolution of Deoxyribonucleosides
HWAHAK KONGHAK, April 1998, 36(2), 343-346(4), NONE
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Abstract
역상 고성능 액체크로마토그래피를 이용하여 다섯 가지 데옥시리보뉴클레오사이드를 분리하였다. 본 연구에서는 최적분리조건을 얻기 위해서 계단함수조건에서 새로운 이동상으로 바뀌는 경우 이동상의 소요부피를 새로이 설정하고 체류인자와 이동상의 조성의 관계식을 사용하였다. 이동상의 조성을 변하게 하는 구배용매조성법에서는 계단함수를 적용하여 조성과 구배시간을 결정하였다. 물을 기본으로 유기용매인 methanol을 첨가하는 이성분계 이동상을 사용하였다. 용출곡선은 이차선형식인 lnk’=AF2+BF+C을 사용하여 단이론으로부터 계산하였다. F는 이동상에서 유기용매의 부피분율이고, A, B, C는 실험상수이다. 최적이동상은 다섯 가지 성분간의 분리도와 가장 늦게 용출되는 성분, dAdo,의 체류시간으로부터 정하였다. 계산된 최적 분리조건은 초기조성에서 순수한 물을 사용하고 7분 후에 조성을 methanol을 19%로 계단함수로 첨가하는 경우이고 실험값과 이론값은 잘 일치하였다.
The separation condition for five deoxyribonucleosides(dCyd, dUrd, dGuo, dThd, and dAdo) was determined by the optimization developed in this work with RP-HPLC(Reversed-Phase High Performance Liquid Chromatography). The method utilized the newly defined parameter, Vm,1 and the relationship of retention factor and mobile phase composition. In a gradient-elution mode by changing mobile phase compositions, the composition and gradient time were determined in the stepwise-gradient mode. In this work, the binary systems of mobile phase(water/methanol) were applied. The elution profiles were calculated by the plate theory based on the quadratic equation of retention factor, lnk’=AF2+BF+C. F denotes the volume fraction of organic modifier in the mobile phase, while A, B, and C the empirical constants. The optimal mobile phase was obtained by comparing the calculated results of the resolutions of the five components and the retention times of last-eluting component(dAdo). The final result showed that the initial mobile phase was pure water, then after 7 min the second composition of water/methanol, 81/19(v/v,%), was step-changed. In this case, the agreement between the experimental data and the calculated values was good.
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