Liquid chromatographic resolution of racemic rasagiline and its analogues on a chiral stationary phase based on (+)‐(18‐crown‐6)‐2,3,11,12‐tetracarboxylic acid

A liquid chromatographic chiral stationary phase based on (+)‐(18‐crown‐6)‐2,3,11,12‐tetracarboxylic acid was applied to the resolution of 15 analytes, including racemic rasagiline, a chiral drug for the treatment of Parkinson's disease, and its analogues. The composition of mobile phase was optimized to be ethanol/acetonitrile/acetic acid/triethylamine (80:20:0.2:0.3, v/v/v/v) by evaluating the chromatographic results for the resolution of five selected analytes under various mobile phase conditions. Under the optimized mobile phase conditions, racemic rasagiline was resolved quite well with a separation factor of 1.48 and resolution of 2.71 and its 14 analogues were also resolved reasonably well with separation factors of 1.06–1.54 and resolutions of 0.54–2.11. Among 15 analytes, racemic rasagiline was resolved best except for just one analyte. The analyte structure–enantioselectivity relationship indicated that racemic rasagiline has the most appropriate structural characteristics for resolution on the chiral stationary phase.     

正反相模式下键合纤维素手性固定相对手性化合物的拆分

在自行合成的键合型纤维素-(3,5-二甲基苯基氨基甲酸酯)(CDMPC-CSP)手性固定相上,在正相模式下对苯偶姻和甲霜灵外消旋体进行了拆分,考察了流动相中醇以及温度在手性识别中对手性分离的影响。同时考察了反相模式下外消旋体的拆分。     

High-performance liquid chromatographic resolution of 1-(1,4-benzodioxane-2-formyl)-piperazine enantiomers after chiral derivatization

Chiral separation of racemic mixtures is of the greatest importance to the pharmaceutical industry, as the isomers of a given racemate may exhibit substantially different pharmacological effects, not to mention possibly differing toxicity behaviour. A novel chiral separation method is developed for the determination of 1-(1,4-benzodioxane-2-formyl)piperazine (BFP) enantiomers. The indirect resolution is performed by applying precolumn derivatization with the chiral reagent 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (GITC). The resulting diastereoisomers are separated on a reversed-phase ODS column with methanol-potassium dihydrogen phosphate (0.02mol/L, 50:50) as mobile phase. UV detection is at 250 nm. The effect of mobile phase composition upon resolution and analysis time is investigated. Two diastereoisomers show nearly base-line separation under optimal chromatographic conditions. The presented study provides a simple and accurate method for the enantiomeric quality control and the optical purity assay of BFP.     

去甲万古霉素键合毛细管电色谱硅胶整体柱的制备及应用

采用溶胶-凝胶技术制备了具有高机械强度和良好通透性的毛细管硅胶整体柱。以国产大环抗生素去甲万古霉素为 手性选择试剂对所制备的整体柱进行化学衍生,成功地制备了去甲万古霉素键合手性硅胶整体柱。在反相和极性有机相模 式下考察了所制备柱的手性识别能力,并详细考察了流动相条件对分离的影响。研究结果表明,β-受体阻滞剂类药物在极 性有机流动相组成为甲醇-乙腈-乙酸-三乙胺(体积比为80∶20∶0.1∶0.1)时,可获得最佳分离。在反相色谱条件下,电 渗流仍主要由整体硅胶基质材料产生,而手性选择试剂的贡献甚小。在反相色谱条件下,多种不同结构类型的手性药物在 所制备的色谱柱上获得了分离。     

Liquid chromatographic enantiomer separation of racemic amine using chiral crown ether stationary phase

Direct enantioseparation of racemic amine, amino-thiophene-2-yl-acetonitrile (TAN), on chiral crown ether stationary phase [Crownpak CR (+)] is described in this study. The elution behavior and the effect of acid additives on resolution of racemic amine, TAN, is intensely investigated. Moreover, the chiral recognition mechanism in this specific system is proposed based on computational methods with the density functional theory. Diastereomeric complexation of the ammonium ion of racemic amine inside the cavity of chiral crown ether appears essential for the chiral discrimination. The pH of the mobile phase containing acid additives also acts as an important factor for the chiral recognition.     

Direct TLC Resolution of (±)-Ketamine and (±)-Lisinopril by Use of (+)-Tartaric Acid or (−)-Mandelic Acid as Impregnating Reagents or Mobile Phase Additives. Isolation of the Enantiomers

Direct resolution of the enantiomers of the racemic drugs ketamine and lisinopril has been achieved by TLC. Enantiomerically pure tartaric acid and mandelic acid were used as chiral impregnating reagents and as mobile phase additives. When (?)-mandelic acid was used as chiral impregnating reagent use of ethyl acetate–methanol–water 3:1:1 (v/v) as mobile phase enabled successful resolution of the enantiomers of both compounds. For lisinopril, the mobile phase acetonitrile–methanol–water–dichloromethane 7:1:1:0.5 (v/v) was successful when (+)-tartaric acid was used as impregnating agent. When (+)-tartaric acid was used as mobile phase additive the mobile phase acetonitrile–methanol–(+)-tartaric acid (0.5% in water, pH 5)–glacial acetic acid 7:1:1.1:0.7 (v/v) enabled successful resolution of the enantiomers of lisinopril. The effects on resolution of temperature, pH, and the amount of chiral selector were also studied. The separated enantiomers were isolated and identified. Spots were detected with iodine vapour. LODs were 0.25 and 0.27 μg for each enantiomer of ketamine with (+)-tartaric acid and (?)-mandelic acid, respectively, whereas for lisinopril LODs were 0.14 and 0.16 μg for each enantiomer with (+)-tartaric acid (both conditions) and (?)-mandelic acid, respectively.     

Reversed-phase chiral HPLC and LC/MS analysis with tris(chloromethylphenylcarbamate) derivatives of cellulose and amylose as chiral stationary phases

Three polysaccharide-derived chiral stationary phases (CSP) were evaluated for the resolution of more than 200 racemic compounds of pharmaceutical interest in the reversed-phase (RP) separation mode. The population of test probes was carefully evaluated in order to insure that it covers as completely as possible all structural diversity of chiral pharmaceuticals. RP showed the highest potential for successful chiral resolution in HPLC and LC/MS analysis when compared to normal phase and polar organic separation modes. Method development consisted of optimizing mobile phase eluting strength, nature of organic modifier, nature of additive and column temperature. The newer CSPs, cellulose tris(3-chloro-4-methylphenylcarbamate) and amylose tris(2-chloro-5-methylphenylcarbamate), were compared to the commonly used cellulose tris(3,5-dimethylphenylcarbamate) in regards to their ability to provide baseline resolution. Comparable success rates were observed for these three CSPs of quite complimentary chiral recognition ability. The same method development strategy was evaluated for LC/MS analysis. Diethylamine as additive had a negative effect on analyte response with positive ion mode electrospray (ESI⁺) MS(/MS) detection, even at very low concentration levels (e.g., 0.025%). Decreasing the organic modifier (acetonitrile or methanol) content in the mobile phase often improved enantioselectivity. The column temperature had only a limited effect on chiral resolution, and this effect was compound dependent. Ammonium hydrogencarbonate was the preferred buffer salt for chiral LC with ESI⁺ MS detection for the successful separation and detection of most basic pharmaceutical racemic compounds. Ammonium acetate is a viable alternative to ammonium hydrogencarbonate. Aqueous formic acid with acetonitrile or methanol can be successfully used in the separation of acidic and neutral racemates. Cellulose tris(3-chloro-4-methylphenylcarbamate) and amylose tris(2-chloro-5-methylphenylcarbamate) emerge as CSPs of wide applicability in either commonly used separation modes rivaling such well established CSPs as cellulose tris(3,5-dimethylphenylcarbamate). Screening protocols including these two new CSPs in the preferentially screened set of chiral columns have higher success rates in achieving baseline resolution in shorter screening time.     

Enantioseparations of basic and bifunctional pharmaceuticals by capillary electrochromatography using polysaccharide stationary phases

A fast screening strategy was developed in capillary electrochromatography (CEC) for the chiral separation of basic and bifunctional compounds. The screening conditions were determined on polysaccharide chiral stationary phases using 15 pharmaceutical compounds. The content and type of organic modifier, as well as the pH of the mobile phase appeared to have the largest influence on the chiral resolution. It was seen that for acidic compounds, our approach was not suitable. A generic mobile phase for basic and bifunctional compounds was determined. The testing on 20 additional compounds showed that the proposed mobile phase performed well since enantioselectivity was observed for 86% of the investigated compounds. A comparison of CEC and reversed-phase liquid chromatography (RPLC) results was attempted to demonstrate the potential of the used technique for chiral method development.     

酰胺型手性固定相反相拆分布洛芬药物对映体

利用酰胺型手性固定相反相拆分了布洛芬对映异构体,用有０．０１ｍｏｌ／Ｌ ＮＨ４Ａｃ的甲醇和水作流动相。在优化分离条件的同时,研究了不同的有机调节和对分离的影响,并探讨了分离的机理。     

Liquid chromatographic resolution of racemic amines,amino alcohols and related compounds on a chiral crown ether stationary phase

A chiral stationary phase (CSP) based on diphenyl-substituted 1,1'-binaphthyl crown ether was applied in resolving various racemic amines, amino alcohols and alpha-aminocarbonyl compounds including pharmaceutically important compounds such as amphetamine analogues, mexiletine, norepinephrine and norephedrine. The resolution was quite successful. In order to find out the effects of mobile phase additives on the chromatographic resolution behaviors, four selected racemic compounds were resolved on the CSP with the variation of the type and content of organic, acidic and cationic modifiers in aqueous mobile phase and with the variation of column temperature. The resolution behaviors were quite dependent on the type and the content of organic, acidic and cationic modifiers in aqueous mobile phase and on column temperature.     

Chiral resolution of flurbiprofen and ketoprofen enantiomers by HPLC on a glycopeptide-type column chiral stationary phase

Vancomycin is an amphoteric, glycopeptide, macrocyclic antibiotic. When attached to 5 microspherical silica gel, vancomycin proved to be an effective chromatographic chiral stationary phase that could be used in the reversed-phase mode. In this study, a bonded vancomycin chiral stationary phase (Chirobiotic Vtrade mark) was investigated for the chiral liquid chromatography analysis of ketoprofen and flurbiprofen. The selectivity factor (alpha) and the chiral resolution factor (RS) of Chirobiotic Vtrade mark were evaluated first as a function of the buffer pH and molarity, and second as a function of organic modifier type and composition of the mobile phase. Four organic modifiers (tetrahydrofuran, 2-propanol, 1,4-dioxane and methanol) have been tested for their selectivity. Optimized conditions using 20% of tetrahydrofuran in ammonium nitrate (100 mM, pH 5) were selected for the enantioseparation of flurbiprofen and ketoprofen from their racemic forms. At pH 5, these acidic compounds are almost negatively charged, while the chiral selector possesses a positive charge allowing it to interact electrostatistically with the analytes. Using these chromatographic conditions, the column stability was excellent over several months of experiments.     

利用酰胺型手性固定相正相拆分β-受体阻滞剂对映体

利用酰胺型手性固定相正相直接拆分了β－受体阻滞剂阿替洛尔,讨论了三元流动相中１,２－二氯乙烷和甲醇含量的改变以及不同极性调节剂的使用对分离的影响。优化的流动相组成为Ｖ（正己烷）∶Ｖ（１,２－二氯乙烷）∶Ｖ（甲醇）＝６８∶２６∶６,并在探讨分离机理的同时比较了阿替洛尔、噻利洛尔和普萘洛尔３种β－受体阻滞剂在酰胺型手性固定相上拆分的结果     

Liquid Chromatographic Resolution of Racemic 2-Phenyl-1,1-cyclo- propanedicarbonitrile and its Analogues on Chiral Stationary Phase

IntroductionLiquid chromatographic resolution of enantiomers on chiral stationary phases is nowemployed as the most convenient and accurate means of determining enantiomericcomposition of chiral compounds"'. There are reports on the resolution of chiralcyclopro-pane compounds"', but the resolution of the title compounds has not beenreported. We have synthesized the racemic and optically active enantiomers of 2-(Xphenyl)-l,I-cyclopropanedicarbon and 2 Q -, and 2 D -naphthyl-l,lcyclopropanedic…     

Reviewing mobile phases used on Chiralcel OD through an application of data mining tools to CHIRBASE database

During the past decade, thousands of compounds have been resolved on Chiralcel OD (a cellulose-based chiral stationary phase) under diverse eluting conditions. Many researches have documented the effects of mobile phase on enantioselectivity for a given family of samples but today no comprehensive study aimed at identifying the associations between the structural features present on solute and appropriate mobile phase conditions has yet been proposed. In this review of mobile phases used on Chiralcel OD, we try to go far beyond a simple enumeration of eluting conditions and an effort is made to explore the utility of data mining tools for assessing the knowledge contained in CHIRBASE database. We have extracted from CHIRBASE the chemical features of 2363 chiral compounds separated on Chiralcel OD and their corresponding mobile phases. This data set was submitted to data mining programs for molecular pattern recognition and mobile phase predictions for new cases. Some substructural characteristics of solutes were related to the efficient use of some specific mobile phases. For example, the application of CH3CN/salt buffer at pH 6-7 was found convenient for reversed-phase separation of compounds bearing a tertiary amine functional group. Furthermore, a cluster analysis allowed the arrangement of the mobile phases according to similarity found in molecular patterns of solutes. A decision tree, which may lead to a more rational choice of the mobile phase under reversed-phase conditions, is also proposed.     

Direct TLC Resolution of (±)-Ketamine and (±)-Lisinopril by Use of (+)-Tartaric Acid or (−)-Mandelic Acid as Impregnating Reagents or Mobile Phase Additives. Isolation of the Enantiomers

Direct resolution of the enantiomers of the racemic drugs ketamine and lisinopril has been achieved by TLC. Enantiomerically pure tartaric acid and mandelic acid were used as chiral impregnating reagents and as mobile phase additives. When (−)-mandelic acid was used as chiral impregnating reagent use of ethyl acetate–methanol–water 3:1:1 (v/v) as mobile phase enabled successful resolution of the enantiomers of both compounds. For lisinopril, the mobile phase acetonitrile–methanol–water–dichloromethane 7:1:1:0.5 (v/v) was successful when (+)-tartaric acid was used as impregnating agent. When (+)-tartaric acid was used as mobile phase additive the mobile phase acetonitrile–methanol–(+)-tartaric acid (0.5% in water, pH 5)–glacial acetic acid 7:1:1.1:0.7 (v/v) enabled successful resolution of the enantiomers of lisinopril. The effects on resolution of temperature, pH, and the amount of chiral selector were also studied. The separated enantiomers were isolated and identified. Spots were detected with iodine vapour. LODs were 0.25 and 0.27 μg for each enantiomer of ketamine with (+)-tartaric acid and (−)-mandelic acid, respectively, whereas for lisinopril LODs were 0.14 and 0.16 μg for each enantiomer with (+)-tartaric acid (both conditions) and (−)-mandelic acid, respectively.

     

Evaluation of the chiral recognition properties as well as the column performance of four chiral stationary phases based on cellulose (3,5-dimethylphenylcarbamate) by parallel HPLC and SFC

The performance of four commercially available cellulose tris(3,5-dimethylphenylcarbamate) based chiral stationary phases (CSPs) was evaluated with parallel high performance liquid chromatography (HPLC) and super critical fluid chromatography (SFC). Retention, enantioselectivity, resolution and efficiency were compared for a set of neutral, basic and acidic compounds having different physico-chemical properties by using different mobile phase conditions. Although the chiral selector is the same in all the four CSPs, a large difference in the ability to retain and resolve enantiomers was observed under the same chromatographic conditions. We believe that this is mainly due to differences in the silica matrix and immobilization techniques used by the different vendors. An extended study of metoprolol and structure analogues gave a deeper understanding of the accessibility of the chiral discriminating interactions and its impact on the resolution of the racemic compounds on the four CSPs studied. Also, a clear difference in enantioselectivity is observed between SFC and LC mode, hydrogen bonding was found to play an important role in the differential binding of the enantiomers to the CSPs.     

纤维素衍生物涂敷型高效液相色谱手性固定相的制备及手性拆分研究

合成了纤维素-三(苯甲酯)(CTB)和纤维素-三(氨基苯甲酸酯)(CTPC)衍生物,并涂敷于自制的球形硅胶上(5μm,13nm),制备了用于高效液相色谱手性拆分的固定相.考察了涂敷量、流动相等因素对手性拆分的影响,测定了一个不对称氢化酯化反应的对映体过剩值(e.e.值).     

Transforming chiral liquid chromatography methodologies into more sensitive liquid chromatography-electrospray ionization mass spectrometry without losing enantioselectivity

LC-electrospray ionization (ESI) MS conditions were optimized for the individual chiral separation of 19 compounds of pharmaceutical interest using the macrocyclic glycopeptide-based chiral stationary phases in both polar organic and reversed-phase modes (RPM). The influence of mobile phase composition and MS additive type on sensitivity was investigated for all classes of compounds tested. Compounds with amine or amide groups were efficiently separated, ionized, and detected with the addition of 0.1% (w/w) ammonium trifluoroacetate to the solvent system in either the reversed-phase or polar organic mode (POM). Macrocyclic glycopeptide coupled column technology was initially used to screen all chiral compounds analyzed. Baseline resolution of enantiomers was then achieved with relatively short retention times and high efficiencies on Chirobiotic T, Chirobiotic V or Chirobiotic R narrow bore chiral stationary phases. The polar organic mode offered better limits of detection (as low as 100 pg/ml) and sensitivity over reversed-phase methods. An optimum flow-rate range of 200-400 microl/min was necessary for sensitive chiral LC-ESI-MS analysis.     

Unusual resolution of N-(3,5-dinitrobenzoyl)-alpha-amino acids on a chiral stationary phase based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid

While HPLC chiral stationary phases (CSPs) based on chiral crown ethers have been known useful for the resolution of only racemic primary amino compounds or some secondary amino compounds, in this study, we first demonstrated that the CSP based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid is also useful for the resolution of N-benzoyl-alpha-amino acids, which do not contain a primary or secondary amino group. Especially, N-(3,5-dinitrobenzoyl)-alpha-amino acids were resolved better than corresponding N-(3-nitrobenzoyl)- or N-benzoyl-alpha-amino acids, the separation (alpha) and the resolution factors (R(S)) for the resolution of eight N-(3,5-dinitrobenzoyl)-alpha-amino acids being in the range of 1.06-1.81 and 0.54-2.81, respectively. The optimum mobile phase condition was the mixture of acetic acid-triethylamine-acetonitrile with the ratio of 0.05/0.25/100 (v/v/v).     

高效液相色谱手性流动相添加法拆分阿卓乳酸对映体

采用C₁₈反相色谱柱,以磺丁基醚-β-环糊精（SBE-β-CD）作为手性流动相添加剂,建立了阿卓乳酸对映体的高效液相色谱拆分方法。考察了环糊精衍生物类型、手性添加剂浓度、流动相pH、流速和柱温对手性分离的影响,同时探讨了高效液相色谱采用磺丁基醚-β-环糊精分离阿卓乳酸对映体的分离机制及包结常数,确定了色谱条件为YMC-Pack ODS-A C₁₈色谱柱（250 mm×4.6 mm,5 μm）,流动相为含25 mmol/LSBE-β-CD的0.1 mol/L磷酸盐缓冲液（pH 2.68）-乙腈（90：10,v/v）,流速为0.6 mL/min,柱温为30 ℃,紫外检测波长为220 nm。对映体的保留时间分别为26.65和28.28 min,分离度为1.68。本方法分离度好,简便易行,且比使用手性固定相分离更加经济。