Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, China.
J Chromatogr A. 2010 Apr 30;1217(18):3044-52. doi: 10.1016/j.chroma.2010.02.077. Epub 2010 Mar 4.
This work concentrates on a chiral separation technology named biphasic recognition applied to resolution of alpha-cyclohexylmandelic acid enantiomers by high-speed counter-current chromatography (HSCCC). The biphasic chiral recognition HSCCC was performed by adding lipophilic (-)-2-ethylhexyl tartrate in the organic stationary phase and hydrophilic hydroxypropyl-beta-cyclodextrin in the aqueous mobile phase, which preferentially recognized the (-)-enantiomer and (+)-enantiomer, respectively. The two-phase solvent system composed of n-hexane-methyl tert-butyl ether-water (9:1:10, v/v/v) with the above chiral selectors was selected according to the partition coefficient and separation factor of the target enantiomers. Important parameters involved in the chiral separation were investigated, namely the types of the chiral selectors (CS); the concentration of each chiral selector; pH of the mobile phase and the separation temperature. The mechanism involved in this biphasic recognition chiral separation by HSCCC was discussed. Langmuirian isotherm was employed to estimate the loading limits for a given value of chiral selectors. Under optimum separation conditions, 3.5-22.0 mg of alpha-cyclohexylmandelic acid racemate were separated using the analytical apparatus and 440 mg of racemate was separated using the preparative one. The purities of both of the fractions including (+)-enantiomer and (-)-enantiomer from the preparative CCC separation were over 99.5% determined by HPLC and enantiomeric excess reached 100% for the (+/-)-enantiomers. Recovery for the target compounds from the CCC fractions reached 85-88% yielding 186 mg of (+)-enantiomer and 190 mg of (-)-enantiomer. The overall experimental results show that the HSCCC separation of enantiomer based on biphasic recognition, in which only if the CSs involved will show affinity for opposite enantiomers of the analyte, is much more efficient than the traditional monophasic recognition chiral separation, since it utilizes the cooperation of both of lipophilic and hydrophilic chiral selectors.
这项工作集中于一种手性分离技术，名为双相识别，应用于通过高速逆流色谱（HSCCC）拆分α-环己基扁桃酸对映异构体。通过在有机固定相中添加亲脂性（-）-2-乙基己基酒石酸和在水相流动相中添加亲水性羟丙基-β-环糊精，可以进行双相手性识别 HSCCC，分别优先识别（-）-对映体和（+）-对映体。根据目标对映异构体的分配系数和分离因子，选择由正己烷-叔丁基醚-水（9:1:10，v/v/v）组成的两相溶剂系统，并加入上述手性选择剂。研究了手性分离中涉及的重要参数，即手性选择剂（CS）的类型；每种手性选择剂的浓度；流动相的 pH 值和分离温度。讨论了 HSCCC 中这种双相识别手性分离涉及的机制。采用 Langmuir 等温线估计给定手性选择剂值的负载极限。在最佳分离条件下，使用分析仪器分离 3.5-22.0 毫克 α-环己基扁桃酸外消旋体，使用制备仪器分离 440 毫克外消旋体。通过 HPLC 确定制备型 CCC 分离的两个馏分（包括（+）-对映体和（-）-对映体）的纯度均超过 99.5％，（+/-）-对映体的对映体过量均达到 100％。从 CCC 馏分中回收目标化合物的收率达到 85-88％，得到 186 毫克（+）-对映体和 190 毫克（-）-对映体。总体实验结果表明，基于双相识别的 HSCCC 对映体分离比传统的单相识别手性分离更有效，因为它利用了亲脂性和亲水性手性选择剂的协同作用，只有涉及 CS 才会表现出对分析物的相反对映体的亲和力。
