Shibusawa Yoichi, Yamakawa Yutaka, Noji Ryoko, Yanagida Akio, Shindo Heisaburo, Ito Yoichiro
Tokyo University of Pharmacy and Life Science, Division of Structural Biology and Analytical Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan.
J Chromatogr A. 2006 Nov 10;1133(1-2):119-25. doi: 10.1016/j.chroma.2006.08.004. Epub 2006 Aug 22.
Three-phase solvent systems were efficiently utilized for high-speed counter-current chromatography (HSCCC) to separate multiple components with a wide range of hydrophobicity. The compositions of three-phase systems were optimized according to their physical parameters such as volume ratio, viscosity and specific gravity of upper (UP), middle (MP) and lower (LP) phases. The three-phase systems composed of n-hexane-methyl acetate-acetonitrile-water (4:4:3:4, v/v/v/v) was selected for HSCCC separation of a mixture of 15 standard compounds with a wide range in hydrophobicity from beta-carotene to tryptophan. The separation was initiated by filling the column with a mixture of MP and LP both as a stationary phase followed by elution with UP to separate the hydrophobic compounds. Then the mobile phase was switched to MP to elute the moderately hydrophobic compounds, and finally the polar compounds still retained in the column were fractionated by eluting the column with LP. The system successfully resolved all 15 compounds in one-step operation in 70 min.
三相溶剂系统被有效地用于高速逆流色谱法（HSCCC），以分离多种具有广泛疏水性的成分。根据三相系统的物理参数，如上层（UP）、中层（MP）和下层（LP）相的体积比、粘度和比重，对三相系统的组成进行了优化。由正己烷 - 乙酸甲酯 - 乙腈 - 水（4:4:3:4，v/v/v/v）组成的三相系统被选用于HSCCC分离15种标准化合物的混合物，这些化合物的疏水性范围从β - 胡萝卜素到色氨酸。分离开始时，用MP和LP的混合物作为固定相填充柱子，然后用UP洗脱以分离疏水性化合物。然后将流动相切换到MP以洗脱中等疏水性化合物，最后，通过用LP洗脱柱子来分离仍保留在柱中的极性化合物。该系统在70分钟内通过一步操作成功分离了所有15种化合物。
