Study On Enantioseparation Of Chiral Drugs By High Performance Liquid Chromatography

Enantioseparations of a series of chiral compounds, including chloramphenicol analogs and seven alkyl tropate and ketoprofen and nine ketoprofen esters were investigated using both chiral stationary phase (CSP) and chiral mobile phase additive (CMPA) by high performance liquid chromatography (HPLC). The effects of composition of mobile phase and the structure of solute on chiral separation were extensively studied. The mechanism of chiral selectors recognizing chiral compounds was discussed by altering the kind and the concentration of alcohol, the concentration of acetic acid and so on. On the other hand, enantioseparation of different solutes were compared on the same chiral stationary phase and enantioseparation of the same solute were compared on different chiral stationary phases. Thus the mechanism of chiral stationary phase recognized chiral compounds was achieved and it is favorable for the study of enantioseparartion of other chiral compounds.In the chapter 1 and chapter 2, the literatures on chiral separation were briefly reviewed. The methods of chiral separation were introduced, especially chiral stationary phase (CSP) and chiral mobile phase additive (CMPA) by high performance liquid chromatography (HPLC).In chapter 3, the Whelk-O1 chiral stationary phase (CSP) is a valuable column for the high-performance liquid chromatography (HPLC) resolution of the enantiomers of a wide variety of compounds. So the resolution of enantiomers of three compounds, which are the synthomycin and the thiamphenicol (THA) and the florfenicol (FLR). were studied on (S, S)-Whelk-O1 CSP using HPLC using n-hexane and alcohol additives as mobile phase. And the influence of the type and concentration of alcohol additives on chiral separation of solutes was investigated. Then the enantiomers of THA were confirmed by LC-MS. The results showed that three compounds were separated successfully. When iso-propanol and tert-butanol as mobile-phase additives, the separation factor (a) of synthomycin and THA was higher in contrast to ethanol. However, ethanoi was more favorable for chiral separation ofFLR. Furthermore, the best resolution of synthomycin was 1.89 using a mobile- phase of n-hexane/ethanol (97:3), and that of THA and FLR was separately 1.84 and 3.94 when a mobile-phase of n-hexane/ethanol (90:10) was used. So the method which was used to separate the enantiomers of this type of drugs was got and was also used to determinate the single isomer content which was about 0.5% in the drug as an impurity.In the chapter 4, optically active tartaric acid and its diester derivatives are well known as effective chiral selectors. In the current studies, a self-prepared diester of tartaric acid, (2R, 3R)-di-n-propyl tartrate (DPT) was used as a chiral mobile phase additive (CMPA) for the enantioseparation of seven kinds of alkyl tropate on a silica gel column by high performance liquid chromatography (HPLC). Four kinds of alkyl tropate were successfully enantioseparated using this chiral system, indicating that DPT is a novel useful chiral selector. The influences of mobile-phase composition and solute structure on enantioseparation were extensively studied. Although different types of alcoholic additive or various concentrations of alcoholic additive, DPT and dichloromethane in the mobile phase had great influence on the retention factor (k1) and resolution (Rs), the mobile phase composition essentially had no effect on the separation factor (a). In contrast, the structure of alkyl tropate greatly influenced the separation factor (a)). The results revealed that an increase in the bulkiness of the O-alkyl groups of tropic acid esters reduced the separation factors (a)), indicating that inhibition of the formation of effective hydrogen-bond interaction between DPT and solutes by the bulkiness of the O-alkyl group decreases the retentivity difference between the transient diasterieomeric complex pairs. Our observations suggested that the hydrogen-bond interactions between chiral additives and solutes might be the major chiral recognition mechanism for DPT as CMPA.In the chapter 5, enantiomers of ketoprofen and nine ketoprofen estetrs were extensively studied on five kinds of CSPs, namely ChiraSpher column, (S.S)-Whelk-Ol column. (R, R)-DNB-DPEDA column, cellulose tri(3.5-dimethylphenylcarbairate) column (CDMPC) and cellulose lris(4-methylbenzoate) (CTMB) in the nomial phase. In the current study, theinfluence of the kind and the concentration of alcoholic additives, the concentration of acetic acid on enantioseparation of racemates of ketoprofen, ketoprofen vinyl ester, and eight ketoprofen esters which were synthesized by enzyme as a kind of catalyzer was investigated extensively. On the other hand, the mechanism of chiral recognition of chial selector was discussed and the mechanisms of five kinds CSPs were compared. (1) The results showed that enantioseparation of seven kinds of ketoprofen esters was observed on Chiraspher CSP using normal mobile phase of the mixture of n-hexane and alcoholic additives. The solutes separated include racemic ketoprofen vinyl ester, glucose ester, galactose ester, mannose ester, fructose ester, lactose ester, sucrose ester and maltose ester. Influence of the chemical structure of solutes on chiral separation was studied extensively and the mechanism of chiral recognition was discussed by altering the composition of mobile phase. Besides, the method for separating and detecting ketoprofen ester by chiral stationary phase using high performance liquid chromatography was proposed. The results showed that seven ketoprofen esters achieved enantioseparations and three solutes obtained a baseline separation, namely ketoprofen glucose ester, mannose ester and maltose ester whose best resolution was respectively 1.83, 2.94 and 2.04. On the other hand, in the asymmetric cavities of ChiraSpher CSP the hydrogen bond interaction between solutes and CSP was the key to enantioseparation and retention of solutes. (2) And that enantioseparation of ketoperofen and six kinds of ketoprofen esters was also investigated on (S.S)-Whelk-Ol CSP. Ketoprofen and ketoprofen mannitol ester obtained a baseline separation, and the resolutions of two solutes were respectively 1.72 and 2.46. (3) On the (R, R)-DNB-DPEDA CSP enantiomers of ketoprofen and four ketoprofen esters were separated partly. (4) Besides, enantiomers of several ketoprofen esters were also separated partly on the CDMPC and CTMB CSPs. However, enantioseparation of ketoprofen vinyl ester was only observed on CDMPC CSP. In a word, the best chiral resolution of solutes was obtained on ChiraSpher CSP, enantioseparation of solues was better on the kind of Pirkle CSPs than on the kind of cellulose derivative CSP.