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.     

Liquid chromatographic enantioseparation of aryl alpha-amino ketones on a crown ether-based chiral stationary phase

A liquid chromatographic chiral stationary phase based on (3,3'-diphenyl-1,1'-binaphthyl)-20-crown-6 covalently bonded to silica gel was applied in the resolution of aryl alpha-amino ketones including cathinone, the main psychoactive alkaloid found in the leaves of the khat plant. The resolution was excellent, the separation factors ranging between 1.72 and 8.58 and the resolution factors (R(S)) ranging between 2.60 and 11.10. The chromatographic resolution behaviour was dependent on the type and the content of organic and acidic modifiers and the ammonium acetate concentration in aqueous mobile phase and the column temperature.     

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.     

Liquid chromatographic resolution of gemifloxacin mesylate on a chiral stationary phase derived from crown ether

A new racemic fluoroquinolone antibacterial agent, gemifloxacin mesylate, has been successfully resolved on a chiral stationary phase (CSP) derived from (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid. Compared to the Crownpak CR(+) column, the CSP used in this study was more effective for the resolution of racemic gemifloxacin mesylate, especially in terms of analytical time. The resolution of gemifloxacin mesylate enantiomers on the CSP was found to be dependent on the type and content of organic and acidic modifiers in the aqueous mobile phase and the column temperature.     

The preparative chromatographic enantioseparation of a racemic morphanthridine analog on a chiral stationary phase

Summary The preparative chromatographic enantioseparation of a chiral morphoanthridine analog has been performed on an analytical column using amylose-tris(3,5-dimethylphenylcarbamate) as chiral stationary phase. The racemate (100 mg) was resolved to baseline within 15 min. This paper describes the development of the method, estimation of the capacity of the chiral stationary phase and discussed the potential of the chromatography if performed under preparative conditions. From the results and calculations presented it seems likely that the resolution of 70 tons year⁻¹ could easily be achieved on 30 kg of stationary phase with a mobile-phase consumption of only 720 L day⁻¹.     

Liquid chromatographic resolution of proton pump inhibitors including omeprazole on a ligand exchange chiral stationary phase

A ligand exchange chiral stationary phase (CSP) developed previously in this laboratory by bonding (R)-phenylglycinol derivative, sodium N-[(R)-2-hydroxy-1-phenylethyl]-N-undecylaminoacetate, to silica gel was successfully applied to the resolution of proton pump inhibitors (PPIs) including omeprazole, pantoprazole, lansoprazole and rabeprazole. For example, the separation factors (α) for the resolution of omeprazole, pantoprazole, lansoprazole and rabeprazole were 4.27, 5.28, 2.77 and 4.06, respectively, and the resolutions (R_S) were 2.53, 2.55, 1.93, and 2.01, respectively, when 65% acetonitrile aqueous solution containing 0.5 mM CuSO₄ and 0.05 mM triethylamine was used as a mobile phase. Based on the chromatographic behaviors for the resolution of PPI analogues on CSP 1, a chiral recognition mechanism utilizing the sulfoxide oxygen and the benzimidazole ring nitrogen of PPIs as bidentate coordination donors to form an enantioselective ternary complex with the central Cu(II) ion and the chiral stationary bidentate ligand was proposed.     

Liquid chromatographic separation of phosphoramidate diastereomers on a polysaccharide-type chiral stationary phase

To improve the therapeutic potential of anti-HIV nucleoside analogues (d4T, AZT, 3TC and ddl), the delivery of the corresponding monophosphate from neutral, membrane-permeable prodrugs has been realised by the synthesis of lipophilic phosphoramidate triester prodrugs, such as the simple phenyl-L-alaninephosphate derivatives. However, the present non-stereoselective synthesis results in a mixture of 1:1 diastereomers, which differ from the configuration of the phosphorus atom asymmetric center. Since each diastereomer may have different biological activity and pharmacokinetic profile, analytical methods have to be developed for their separation. This work aims at showing the ability of a polysaccharide-type chiral stationary phase to resolve such diastereomers in reversed-phase high-performance liquid chromatography. The influence of operating parameters has been studied to optimise the separation; a thermodynamic approach has also been investigated to gain an insight in the retention mechanism of the prodrugs. Preliminary validation study (linearity, accuracy, repeatability) has yielded good results; in addition, the feasibility of HPLC-electrospray-mass spectrometry (HPLC-ESI-MS) coupling has been demonstrated and it is expected that this will lead to lower detection limits.     

Liquid chromatographic resolution of racemic biotin

Summary The separation of the enantiomeric forms of biotin (vitamin H) using preparative chiral HPLC is described.     

Liquid chromatographic resolution of 2-hydroxycarboxylic acids on a new chiral stationary phase derived from (S)-leucine

Enantiomers of racemic 2-hydroxycarboxylic acids have been resolved as their O-ethoxycarbonyl pi-basic anilide derivatives on a new chiral stationary phase (CSP) derived from N-(3,5-dinitrobenzoyl)leucine N-phenyl N-alkylamide and the resolution results have been compared with those on various commercial pi-acidic CSPs. The resolution results demonstrate that the new CSP derived from N-(3,5-dinitrobenzoyl)leucine N-phenyl N-alkylamide is most effective among the five CSPs tested for the resolution of 2-hydroxycarboxylic acid derivatives. In order to elucidate the chiral recognition mechanism exerted by the new CSP, the resolution of slightly differently modified derivatives of 2-hydroxycarboxylic acids on the new CSP has been investigated. Based on the resolution results, a chiral recognition mechanism utilizing three simultaneous interactions such as the face to face pi-pi interaction and the two hydrogen bonding interactions between the CSP and the more retained enantiomer of the analyte has been proposed.     

Liquid chromatographic enantiomer resolution of N-fluorenylmethoxycarbonyl alpha-amino acids and their ester derivatives on polysaccharide-derived chiral stationary phases

The liquid chromatographic enantiomer separation of N-fluorenylmethoxycarbonyl (FMOC) protected alpha-amino acids and their ethyl ester derivatives was performed on polysaccharide-derived chiral stationary phases, Chiralcel OD, Chiralpak AD, and Chiralpak AS. In general, Chiralcel OD and Chiralpak AD showed good performance for resolution of N-FMOC alpha-amino acids and their ethyl esters, respectively. All investigated N-FMOC alpha-amino acid enantiomers were baseline separated on Chiralcel OD or Chiralpak AD, whereas N-FMOC alpha-amino acid ethyl ester enantiomers were baseline resolved (alpha = 1.15-3.03) on Chiralpak AD, except for two analytes. The L-enantiomers of all examined FMOC alpha-amino acid ethyl ester derivatives are preferentially retained on Chiralpak AD, while the elution orders of the other enantiomer separations are not consistent.     

Liquid chromatographic separation of enantiomers of beta-amino acids using a chiral stationary phase

The enantiomers of both alpha-substituted beta-alanines and beta-substituted beta-alanines may be chromatographically separated using silica-bonded chiral stationary phases derived from N-acetylated alpha-arylalkylamines. The amino acids are chromatographed as alkyl esters of N-3,5-dinitrobenzoyl derivatives; separability factors range from 1.11 to 1.65 for nine alpha-substituted beta-alanines and from 1.08 to 1.20 for nine beta-substituted beta-alanines. The enantiomers of beta-aminoisobutyrate and beta-leucine, chiral beta-amino acids occurring in animal tissues and physiological fluids, are among those resolved. The enantiomers of R,S-beta-aminoisobutyrate and several related alpha-alkyl-beta-alanines were prepared by chromatographic resolution of diastereomeric dipeptides.     

Direct chromatographic resolution of carnitine and O-acylcarnitine enantiomers on a teicoplanin-bonded chiral stationary phase.

R-(-)-Carnitine (vitamin B(T)) plays an important role in human energy metabolism, by facilitating the transport of long-chained fatty acids across the mitochondrial membranes. Its (S)-enantiomer acts as a competitive inhibitor of carnitine acetyltransferase, causing depletion of the body R-(-)-carnitine stock. Consequently, the separation of carnitine enantiomers is very important both to study their biological activities and to control the enantiomeric purity of pharmaceutical formulations. In the present paper we describe an easy, fast and convenient procedure for the separation of the enantiomers of carnitine and O-acylcarnitines by enantioselective HPLC on a laboratory-made chiral column containing covalently bonded teicoplanin as selector. High enantioselectivity factors (alpha values ranging from 1.31 to 3.02) and short-time analyses characterize the analytical procedure; in addition, analytes are easily detected by evaporative light scattering with no need for preliminary derivatization. The effects of pH and ionic strength of the mobile phase and of the nature of the organic modifier on the enantioselective separations were also investigated.     

Liquid chromatographic separation of the enantiomers of beta-amino acids on a ligand exchange chiral stationary phase

A liquid chromatographic ligand exchange chiral stationary phase (CSP) derived from (S)-leucinol was applied in the separation of the enantiomers of 12 beta-amino acids. The resolution was quite successful especially for the enantiomers of beta-amino acids containing aromatic functional group in the side chain. The chromatographic resolution behaviors were dependent on the organic modifier and Cu(II) concentration in aqueous mobile phase and the column temperature.     

Covalently bonded DNA aptamer chiral stationary phase for the chromatographic resolution of adenosine

In this work, a target-specific aptamer chiral stationary phase (CSP) based on the oligonucleotidic selector binding to silica particles through a covalent linkage was developed. An anti-d-adenosine aptamer was coupled, using an in-situ method, by way of an amide bond to macroporous carboxylic acid based silica. Frontal chromatography analysis was performed to evaluate the column properties, i.e., determination of the stationary phase binding capacity and the dissociation constant of the target-immobilized aptamer complex. It was found that such covalent immobilization was able to maintain the aptamer binding properties at a convenient level for an efficient enantioseparation. Subsequently, the separation of adenosine enantiomers was investigated under different operating conditions, including changes in the eluent’s ionic strength and the proportion of organic modifiers as well as column temperatures. It was demonstrated that, under various conditions of use and storage, the present CSP was stable over time.     

Direct chromatographic resolution of racemic aminoglutethimide and its acetylated metabolite using different cellulose based chiral stationary phases

Summary Two improved methods for the enantiomeric separation of racemic aminoglutethimide (±AG) and its acetylated metabolite (±AAG) have been developed. Direct liquid chromatographic resolution of the enantiomers of aminoglutethimide and its acetylated metabolite was accomplished using Chiralcel OD and Chiralcel OJ columns without any derivatization. Maximum resolution of 8.87 and 2.23 was obtained for the enantiomers of aminoglutethimide and its acetylated metabolite using a Chiralcel OD column, while maximum resolution of 10.34 and 7.01 was obtained for the enantiomers using a Chiralcel OJ column. Optimization of separation was obtained using different concentration of 2-propanol in hexane as a mobile phase.     

Preparation of chiral stationary phase via activated carbamate intermediate for liquid chromatographic optical resolution

Summary Chiral stationary phases (CSPs) for liquid chromatography were prepared by the way of an activated carbamate intermediate. The amino group of aminopropylsilyl silica gel was first activated by carbamylation with disuccinimido carbonate (DSC). The obtained activated carbamate silica gel (ACsil) proved useful as an intermediate for the preparation of urea-type CSPs. The reaction of ACsil with (S)- of (R)-1-(α-naphthyl)-ethylamine gave naphthylethylurea type CSPs. These CSPs were also obtained directly from aminopropylsilyl silica gel by its reaction with optically active (S)- or (R)-succinimido 1-(α-naphthyl)ethyl carbamate (SINEC). Several phenylthiohydantoin amino acid enantiomers and p-bromophenylcarbamyl amino acid enantiomers were resolved on the CSPs by elution with aqueous mobile phase.     

量子化学计算解析手性共价有机框架材料6色谱固定相的手性拆分机理

为探究手性共价有机框架材料6(Chiral Covalent Organic Frameworks 6, CCOF6)色谱固定相的手性拆分机理,首先运用ORCA程序对CCOF6及4对手性对映体进行结构优化,然后使用AutoDock程序对CCOF6及各对映体分子进行分子对接,获得CCOF6与对映体相互作用的初始构型;采用ORCA程序(B3LYP泛函,带DFT-D3校正,轨道基组为def2-TZVP, def2/J作为RI-J的辅助基组,RIJCOSX用来加速计算)对初始构型进行能量计算,以最终确定CCOF6与对映体的相互作用构型,并获得相应的结合自由能和结合自由能差;使用Multiwfn程序对ORCA结果进行独立梯度模型分析,并应用视觉分子动力学程序可视化展示CCOF6与对映体的弱相互作用。结果表明:(1)在计算结合自由能方面,考虑了溶剂效应的ORCA计算方法比不考虑溶剂效应的ORCA以及AutoDock计算方法更为精确;(2)CCOF6色谱固定相与对映体之间的结合自由能差绝对值越大,对映体的选择性因子也越大,然而对映体的分离度不一定会越大;(3)除S-1-苯基-1-丙醇是以羟基和CCO...     

Characterization of an unusual adsorption behavior of racemic methyl-mandelate on a tris-(3,5-dimethylphenyl) carbamoyl cellulose chiral stationary phase

An interesting adsorption behavior of racemic methyl mandelate on a tris-(3,5-dimethylphenyl)carbamoyl cellulose chiral stationary phase was theoretically and experimentally investigated. The overloaded band of the more retained enantiomer had a peculiar shape indicating a type V adsorption isotherm whereas the overloaded band of the less retained enantiomer had a normal shape indicating a type I adsorption behavior. For a closer characterization of this separation, adsorption isotherms were determined and analyzed using an approach were Scatchard plots and adsorption energy distribution (AED) calculations are combined for a deeper analysis. It was found that the less retained enantiomer was best described by a Tóth adsorption isotherm while the second one was best described with a bi-Moreau adsorption isotherm. The latter model comprises non-ideal adsorbate-adsorbate interactions, providing an explanation to the non-ideal adsorption of the more retained enantiomer. Furthermore, the possibility of using the Moreau model as a local model for adsorption in AED calculations was evaluated using synthetically generated raw adsorption slope data. It was found that the AED accurately could predict the number of adsorption sites for the generated data. The adsorption behavior of both enantiomers was also studied at several different temperatures and found to be exothermic; i.e. the adsorbate-adsorbate interaction strength decreases with increasing temperature. Stochastic analysis of the adsorption process revealed that the average amount of adsorption/desorption events increases and the sojourn time decreases with increasing temperature.     

手性流动相HPLC法拆分萘普生对映体的研究

将β-环糊精、甲基-β-环糊精、羟丙基-β-环糊精、L-脯氨酸作为手性流动相添加剂,系统地研究了D,L-萘普生在RP-HPLC系统中的拆分.分别考察了手性流动相的种类,手性试剂羟丙基-β-环糊精的浓度,流动相的pH值,修饰剂的种类及浓度,三乙胺浓度和柱温等对拆分效果的影响,以HP-β-CD为手性流动相添加剂,建立了HP-β-CD手性流动相分离萘普生对映体的方法.结果表明：当流动相为25 mol/L HP-β-CD、体积分数15%乙醇、体积分数0.5%三乙胺、pH3.5、柱温t25℃、流速V=1 mL/min时萘普生对映体得到了良好的基线分离,分离因子α可达1.29.