Separation Of Isomers Of Drugs By High-Performance Liquid Chromatography On Chiral Stationary Phase

This thesis focuses on the enantioseparation using chiral stationary phases (CSPs) by high performance liquid chromatography (HPLC), including enantioseparation of a group of herbicides and a group of chiral antifungal drugs with similar structure under normal phase mode and discussion of the recognition mechanism. Besides, the cis and trans isomers separation of a novel anti-ulcer drug lafutidine and its intermediate 2-Butene-l,4-diol were studied.In this thesis, the chiral columns included small molecule Pirkle type (S,S)-Whelk-Ol, high molecule CSP ChiraSpher and four kinds of high molecule cellulose derivative CSPs, cellulose tribenzoate (CTB), cellulose tris(4-methyl benzoate) (CTMB), cellulose triphenylcarbamate (CTPC) and cellulose tris(3,5-dimethyl phenyl carbamate) (CDMPC).In chapter 1 and chapter 2, the literatures of the stereochemistry and its separation, the kinds of Pirkle type CSP and cellulose derivative CSP, the factor influencing chiral selectivity, the mechanism of chiral recognition were briefly reviewed.In chapter 3, the enantioseparation of a group of herbicides with similar structure was studied: (1) the enantioseparation of diclofop was investigated on four kinds of coated cellulose derivative chiral columns (CTB, CTMB, CTPC and CDMPC) for the first time. It was the result that the retention of diclofop was strongest on CTMB, but diclofop obtained better enantioseparation on CDMPC than on CTMB. By the investigation of the influence of different polar alcohol additive in the mobile phase on the chiral separation and the compared experiment on different column, it was found that the alcohol additive could change the solid environment of chiral cave by interacting with the chiral and non-chiral dot in or near the chiral cave of the CSP. Different alcohol has different change ability, which correspondingly leads to different chiral separation efficiency. Diclofop gained better enantioseparation on cellulose derivative CSP when large alcohol, isopropanol and n-butanol were used as the additive in the mobile phase; (2) The Enantioseparation of four aryloxyphenoxypropionic acid herbicides, diclofop, fluazifop, quizalofop-P and fenoxaprop by HPLC on self-prepared cellulose derivative CDMPC and (S, S)-Whelk-Ol.The mobile phase composition including the type and concentration ofthe alcohol modifier in hexane was varied, and the influence on the retention factor(k'), the separation factor( ) and the resolution (Rs) for each enantiomer was studied. The influence of the structure of the analytes on the chiral separation was investigated. And then their chiral recognition mechanism was discussed. The experiment results showed that fenoxaprop obtained best resolution on CDMPC, while the other three solutes obtained best separation on (S,S)-WheIk-Ol. It was concluded that the four aryloxyphenoxypropionic acid herbicides obtained excellent separation and the chiral recognition mechanisms of CDMPC and (S,S)-Whelk-Ol differs greatly. And the structure of solutes and CSP play a key role in the chiral recognition; (3) On (S, S)-Whelk-Ol and four kinds of self-prepared cellulose derivative columns (CTB, CTMB, CTPC, CDMPC), naproanilide and its intermediate -chloro-propionanilide were both experimented for enantioseparation. Naproanilide obtained best separation on CDMPC. On CTPC naproanilide just obtained part enantioseparation when isopropanol was used as the polar additive. As to -chloro-propionanilide, it obtained baseline enantioseparation on (S, S)-Whelk-Ol and CDMPC, but just obtained part enantioseparation on CTPC when ethanol was used as the polar additive. It is indicated that the difference of structure of naproanilide and -chloro-propionanilide led to different separation efficiency. The influence of volume and concentration of polar alcohol modifier in mobile phase on the enantioseparation of naproanilide and its intermediate on CDMPC was studied. Following are the results: (a) As to a determined alcohol additive, changing volume and concentration led to identical change trend of the separation efficiency; (b) But the order of the chromatographic parameters of the solutes under different alcohol changed. It is because that the density and molecular weight of the different alcohol is different.In chapter 4, the enantioseparation of a group of chiral antifungal drugs were investigated on (S,S)-Whelk-Ol and four different cellulose derivative columns (CTB, CTMB, CTPC, CDMPC). The drugs included three kinds of imidazole (tetramisole, miconazole, imazalil) and seven kinds of triazole (tebuconazole, hexaconazole, myclobutanil, diniconazole, uniconazole, paclobutrazol, triadimenol). For tetramisole, imazalil and uniconazole, it was the first time for enantioseparation. The enantioseparation of each solute on a CSP differed from its structural characteristic and the different CSP showed different chiral discrimination capability for each solute. Following are theconclusions: (1) The retention of most of the solutes was stronger on (S,S)-Whelk-Ol than on the cellulose derivative columns, but the enantioseparation capability was better on the cellulose derivative columns except CTB; (2) Tetramisole was not eluted except on CDMPC. As to other solutes: (a) On (S,S)-Whelk-Ol, just miconazole, hexaconazole and triadimenol obtained enantioseparation; (b) On CTB, the retention of the solutes was weakest and just miconazole, imazalil obtained part enantioseparation; (c) On CTMB, CTPC and CDMPC, the solutes obtained enantioseparation of different degree by changing the mobile phase. Imazalil and tebuconazole obtained best enantioseparation on CTMB. Uniconazole, paclobutrazol, and triadimenol obtained best enantioseparation on CTPC. And the others obtained best enantioseparation on CDMPC. Besides, the influence of mobile phase composition was studied: (a) The retention and resolution factors of the solutes increased with decreasing the alcohol concentration in mobile phase, while separation factors kept steady; (b) On (S,S)-Whelk-Ol, acid modifier in mobile phase had no obvious effect on the enantioseparation; (c) On the four cellulose derivative columns, the most favorable alcohol modifier of each solutes was different. It may be due to that while competing with hydrogen bond interaction between solute and CSP, the different polar alcoholic modifier will have different effect on the steric structure of solute and CSP, especially the chiral cavity of CSP. Thus, for different solute, the inclusion and the steric fitness of solute shape in chiral cavity of CSP may differ greatly, which resulted in different enantioseparation.In chapter 5, the cis and trans isomers separation of 2-butene-l,4-diol and lafutidine were studied by HPLC on two kinds of chiral columns: (S,S)-Whelk-0 1 and ChiraSpher. The isomers of 2-butene-l,4-diol can be separated on both chiral columns while the isomers of lafutidine can only be resolved on ChiraSpher column. The influence of different type and amount of mobile phase modifier on the isomers separation was extensively studied. The resolution of cis and trans isomers of 2-butene-l,4-diol was 2.61 on (S,S)-Whelk-0 1 column with hexane-ethanol (97:3, v/v) as the mobile phase. The resolution of lafutidine was 1.89 on ChiraSpher column with hexane-ethanol-THF-diethylamine (92:3:5:0.1, v/v/v/v) as the mobile phase. LC-MS methods were developed to identify the isomer peaks.