替考拉宁键合手性固定相的制备及其在反相条件下对手性化合物的拆分

以国产大环抗生素替考拉宁为手性选择剂制备了替考拉宁键合手性固定相(Tei-CSP),在反相条件下考察了键合相对华发令、西孟旦等手性药物、α-氨基酸(羟基酸)、衍生α-氨基酸的拆分效果。实验结果表明,在反相条件下,疏水(亲水)作用、静电作用对手性化合物在柱上的保留以及对映体的拆分起到了非常重要的作用;氨基酸在衍生前后,其在柱上的保留和手性识别机理发生了改变,衍生前,亲水作用参与保留机理,而衍生后,由于疏水作用增强,疏水作用参与保留,对映体在较低的有机改性剂条件下才能获得较好的分离。     

3，5-二甲基苯基异氰酸酯替考拉宁和苯基异氰酸酯替考拉宁手性固定相的评价与比较

用大环抗生素替考拉宁手性固定相(TE CSP)分别与3,5-二甲基苯基异氰酸酯和苯基异氰酸酯反应得到了两种新型的高效液相色谱手性固定相----3,5-二甲基苯基异氰酸酯替考拉宁手性固定相(DMP-TE CSP)和苯基异氰酸酯替考拉宁手性固定相(Ph-TE CSP)。用十八个手性化合物在反相及极性流动相模式对这两种CSP的对映体分离能力进行了评价和比较。在反相流动相中,十二个化合物（包括八个氨基酸和四个非氨基酸化合物----对羟基苯甘氨酸,拉米夫定,醇酸和去甲羟安定）的对映体在这两种手性固定相上都获得了分离,大部分的溶质在DMP-TE上获得了更强的保留和稍好的手性分离效果。在极性流动相中,六个氨基醇类化合物在DMP-TE上获得了更强的保留,但它们在两种CSP上的选择因子几乎没有区别。对自制的替考拉宁衍生物手性固定相进行评价和比较,将有助于大环糖肽类抗生素手性固定相手性识别机理的研究。     

利用“网包法”制备高效液相色谱大环抗生素类手性固定相

万古霉素和替考拉宁都属于糖肽类的大环抗生素，具有立体的环状结构和多个手性中心，是两种常见的手性识别材料，广泛应用于对映体的色谱手性分离分析。该文以万古霉素和替考拉宁为手性选择剂，哌嗪为单体，4，4'-二苯基甲烷二异氰酸酯（MDI）、1，6-己二异氰酸酯（HDI）和2，4-甲苯二异氰酸酯（TDI）为交联剂，通过界面聚合反应形成网状层包裹硅胶载体的方法制得6种高效液相色谱手性固定相，用于分离外消旋化合物，并与MDI直接交联万古霉素和替考拉宁在硅胶表面所得固定相进行了比较。结果表明，利用"网包法"和直接交联法制备的手性柱与商品万古霉素和替考拉宁柱之间具有互补性，均对不同的外消旋体有不同程度的拆分。     

替考拉宁及间甲基苯基异氰酸酯替考拉宁手性固定相的 对映体分离能力评价与比较

由替考拉宁手性固定相(TE CSP)制备出了一种新型的高效液相色谱手性固定相: 间甲基苯基异氰酸酯替考拉宁手性固定相(TI-TE CSP). 在反相流动相中用7种氨基酸和3种非氨基酸化合物对这两种手性固定相的手性分离能力进行了评价和比较. 考察了有机添加剂的种类和浓度, 缓冲液的pH值等条件对10个手性化合物在两种CSP上手性分离的影响, 计算得出了溶质在两种CSP上的手性选择性自由能差值, 同时初步探讨了这些溶质在两种CSP上的手性识别机理. 实验数据表明, 氨基酸在TE上保留更强, 但在TI-TE上得到了更好的手性分离效果. 结果显示, 经间甲基苯基异氰酸酯衍生化后的替考拉宁CSP在反相流动相中的分离能力有所提高.     

分子模型及其在手性识别机理研究上的应用

介绍了近十几年来在色谱手性识别机理研究中的分子模型。在这些模型中,采用量子力学、分子力学和分子动力学等方法计算了手性选择试剂与对映体之间的相互作用,并借助X射线晶体学、核磁共振技术和计算机模拟等技术建立了各种分子模型,研究在手性化合物分离过程中的手性识别机理。     

手性选择试剂环糊精与万古霉素对氨基酸对映异构体迁移顺序的影响

用毛细管电泳法分别在由两种手性选择试剂,β-环糊精(β-CD)和万古霉素(VMC)所组成的分离体系中,研究了19种氨基酸的衍生物的对映异构体的迁移顺序(EMO)。单一β-CD分离体系由含有不同浓度的β-CD的硼酸盐缓冲溶液组成;二元分离体系由不同浓度比例的β-CD和脱氧牛黄胆酸钠溶液所组成,各溶液的pH值为9.5。单一VMC分离体系含2mmol·L-1 VMC缓冲溶液(pH 7.2);二元VMC分离体系由浓度均为2mmol·L-1 VMC溶液和海美溴铵溶液组成(pH 7.2)。19种氨基酸须先与芴甲氧羰基氯反应使衍生化。结果表明:各氨基酸的对映体在相同或不同的分离体系中的EMO不一致,特别是在二元VMC体系中,氨基酸的EMO完全发生逆转。据此认为,对于目标氨基酸对映体的分离,可通过用不同的手性分离体系来实现。     

替考拉宁手性柱高效液相色谱法拆分萘哌地尔对映体及分离机制

在极性有机相条件下用替考拉宁手性柱(Chirobitotic T)直接分离萘哌地尔对映体。考察了三乙胺、醋酸的浓度、比例及柱温、流速对拆分的影响。应用Molecular operation Environment(MOE)软件模拟R/S-萘哌地尔与替考拉宁的相互作用,结合热力学参数探讨其分离机制。结果表明,随着甲醇中三乙胺和醋酸的浓度变小,分离度增大;三乙胺与醋酸的体积比为1∶1时,分离度最好。温度和流速的升高都会使分离度降低。优化后条件为:流动相为甲醇-三乙胺-醋酸(100∶0.002∶0.002,V/V),流速1 mL/min,柱温30℃。在此条件下,萘哌地尔对映体达到了最佳分离,分离度为2.0,理论塔板数达到5100。结合热力学参数和分子模拟结果表明,引起手性识别的主要作用力包括氢键、立体阻碍和π-π作用力。     

气相色谱手性固定相进展

1966年Gil-Av等人首次用手性固定相在毛细管柱上分离了氨基酸对映体。至今这种方法已发展为研究和分离对映体的一种重要技术,并愈来愈多地用于天然产物绝对构象(外激素、香料成分等)的测定,不对称选择合成中对映体纯度及过剩量测定,手性药物中对映体纯度的测定,手性化合物在化学转移机理等方面的研究。目前气相色谱中用于分离对映体的     

替考拉宁键合手性固定相高效液相色谱法直接拆分泮托拉唑钠对映体

以替考拉宁为手性选择剂制备了大环抗生素类手性固定相替考拉宁键合手性固定相(T-CSP),建立了T-CSP反相液相色谱直接拆分泮托拉唑钠对映体的方法。考察了流动相中有机改性剂的种类和比例、柱温以及流动相流速对拆分泮托拉唑钠对映体的影响。研究发现,用甲醇作有机改性剂比乙腈更有利于对映体的分离;在研究的温度范围内,随着柱温的升高,对映体的保留时间缩短,同时分离因子和分离度降低;在一定范围内降低流速有利于对映体的分离。采用T-CSP色谱柱(150 mm×4.6 mm i.d.,5 μm),以甲醇-水(体积比为35∶65)为流动相,在流速0.6 mL/min、检测波长290 nm、柱温20 ℃的条件下,泮托拉唑钠对映体获得了近于基线的分离,所建立的方法具有简便快速及重复性好等优点。     

微波辅助合成法快速制备替考拉宁开管毛细管电色谱柱

采用微波辅助合成技术,快速制备了以替考拉宁为固定相的开管毛细管电色谱柱。在pH 4.0~7.0的范围内比较了空管与替考拉宁修饰柱的电渗情况,表明替考拉宁开管毛细管电色谱柱有效地降低了电渗。用该色谱柱分离了多种手性对映体,均达到基线分离,体现了替考拉宁开管毛细管电色谱柱良好的分离性能。以DL-色氨酸考察了柱子的稳定性和重现性,结果显示采用微波辅助合成技术制得的替考拉宁开管毛细管电色谱柱具有良好的稳定性和重现性。     

Avoparcin, a new macrocyclic antibiotic chiral run buffer additive for capillary electrophoresis.

Avoparcin, like vancomycin, teicoplanin, and ristocetin A, belongs to the family of macrocyclic glycopeptide antibiotics. These antibiotics have all been used as effective chiral selectors for capillary electrophoresis (CE), thin-layer chromatography (TLC), and high performance liquid chromatography (HPLC). The present work focuses on avoparcin, which has been shown to be an excellent chiral selector for the CE enantioseparation of many N-blocked amino acids, as well as several anti-inflammatory drugs of pharmaceutical importance. The use of avoparcin as a chiral run buffer additive in CE is discussed, as well as the effects of changing experimental parameters, like avoparcin concentration, pH, organic modifiers, etc. Comparisons of enantioseparations of some N-3,5-dinitrobenzoyl-derivatized amino acids, using either avoparcin, ristocetin A, teicoplanin, or vancomycin in the run buffer, are also made. In general, vancomycin had the longest migration times, and ristocetin A the shortest, while avoparcin was intermediate. Generally, at least one of the four chiral selectors produced an excellent separation, while a different macrocyclic antibiotic produced a poor separation. Currently, we see no way to predict which chiral run buffer additive will be best or worst for an individual solute.     

HPLC separation of amino acid enantiomers and small peptides on macrocyclic antibiotic-based chiral stationary phases: a review

The search for new and effective chiral selectors capable of separating a wide variety of enantiomeric compounds is an ongoing process. In the past decade, macrocyclic antibiotics have proved to be an exceptionally useful class of chiral selectors for the separation of enantiomers of biological and pharmacological importance by means of HPLC, TLC and electrophoresis. More chiral analytes have been resolved through the use of glycopeptides than with all the other macrocyclic antibiotics combined (ansamycins, thiostrepton, aminoglycosides, etc.). The glycopeptides avoparcin, teicoplanin, ristocetin A and vancomycin have been extensively used as chiral selectors in the form of chiral bonded phases in HPLC, and HPLC stationary phases based on these glycopeptides have been commercialized. Teicoplanin, vancomycin, their analogs and ristocetin A seem to be the most useful glycopeptide HPLC bonded phases for the enantioseparation of proteins and unusal native and derivatized amino acids. In fact, the macrocyclic glycopeptides are to some extent complementary to one another: where partial enantioresolution is obtained with one glycopeptide, there is a high probability that baseline or better separation can be obtained with another. This review sets out to characterize the physicochemical properties of these antibiotics and their application in the enantioseparations of amino acids. The mechanism of separation, the sequence of elution of the stereoisomers and the relation to the absolute configuration are also discussed.     

High-performance liquid chromatographic enantioseparation of 2-aminomono- and dihydroxycyclopentanecarboxylic and 2-aminodihydroxycyclohexanecarboxylic acids on macrocyclic glycopeptide-based phases

The direct separation of the enantiomers of four 2-aminomono- or dihydroxycyclopentanecarboxylic acids and four 2-aminodihydroxycyclohexanecarboxylic acids was performed on chiral stationary phases containing macrocyclic glycopeptide antibiotics such as teicoplanin (Astec Chirobiotic T and T2), teicoplanin aglycone (Chirobiotic TAG) or ristocetin A (Chirobiotic R) as chiral selectors. The effects of the nature of organic modifiers, the pH, the mobile phase composition and the structures of the analytes on the separation were investigated. Chirobiotic TAG, and in some cases Chirobiotic T, proved to be the most useful of these columns. The elution sequence was determined in most cases.     

Chiral separations using the macrocyclic antibiotics: a review

The macrocyclic antibiotics have recently gained popularity as chiral selectors in CE, HPLC and TLC. The macrocyclic antibiotics used for chiral separations include the ansamycins, the glycopeptides, and the polypeptide antibiotic thiostrepton. Although not strictly considered macrocyclic antibiotics, the aminoglycosides are antibiotics that have been used for chiral separations in CE. More chiral analytes have been resolved using the glycopeptides than with the other macrocyclic antibiotics combined. The glycopeptides vancomycin, ristocetin A and teicoplanin have been used extensively as chiral selectors in CE, with ristocetin A appearing to be the most useful chiral selector followed by vancomycin and teicoplanin, respectively. The macrocyclic antibiotics have also been used as chiral bonded phases in HPLC, and HPLC stationary phases based on vancomycin, ristocetin A and teicoplanin have been commercialized. Ristocetin A seems to be the most useful glycopeptide HPLC bonded phase, but its greater expense can be a drawback. The macrocyclic antibiotics have been used with micelles to improve efficiency, provide unique selectivity, and extend the range of separations to neutral solutes. Changing the macrocyclic antibiotic used in CE or HPLC can significantly alter the enantioselectivity of the separations. In fact, the glycopeptide antibiotics are complementary to one another, where if a partial enantioresolution is obtained with one glycopeptide, there is a high probability that a baseline or better separation can be obtained with another.     

Comparative Enantioseparation of Seven Amino Alcohols on Teicoplanin‐Based Chiral Stationary Phases

Abstract

The macrocyclic antibiotics represent a relatively new class of chiral selectors in separation science and teicoplanin‐based chiral stationary phases (CSP) have been used successfully in a number of applications in high‐performance liquid chromatography. In the present studies, we self‐prepared two bonded CSPs–teicoplanin (TE) and teicoplanin phenyl isocyanate (TE‐Phe). Seven amino alcohols, propranolol, bisoprolol fumarate, atenolol, salbutamol, isoproterenol, metoprolol, and labetalol were enantioseparated on both self‐made CSPs using methanol as mobile phase and acetic acid (HOAc) and triethylamine (TEA) as mobile phase additives. On both CSPs, the different enantioseparation behavior of analytes with different structure was compared. The influence of the concentration of mobile phase additives (HOAc and TEA) on the enantioseparation was investigated. In all conditions, the retention factors (k′) of seven analytes on TE‐Phe CSP were larger than that on TE CSP. However, the separation factors (α) and resolutions (Rs) on TE‐Phe CSP were smaller than that on TE CSP. The results indicated that the derivatized TE‐Phe CSP is not efficient as original teicoplanin CSP. Our observations also suggested that, for teicoplanin‐based CSPs, π‐π interactions and dipole‐dipole between solutes and CSPs mainly contribute to the retention of solutes on CSPs while hydrogen bonding and steric interactions play important roles in the chiral recognition for teicoplanin‐based CSPs.     

Chiral analysis of UV nonabsorbing compounds by capillary electrophoresis using macrocyclic antibiotics: 1. Separation of aspartic and glutamic acid enantiomers

Glycopeptide antibiotics, namely vancomycin or teicoplanin, were evaluated in capillary electrophoresis for the analysis of UV nonabsorbing compounds such as aspartic and glutamic acid enantiomers. Electrophoretic runs were performed in laboratory-made polyacrylamide-coated capillaries using the partial filling-counter current method in order to avoid the presence on the detector path of the absorbing chiral selector. The background electrolyte consisted of an aqueous or aqueous-organic buffer in the pH range of 4.5-6.5 of sorbic acid/histidine and the appropriate concentration of chiral selector. Several experimental parameters such as antibiotic concentration and type, buffer pH, organic modifier, type and concentration of absorbing co-ion (for the indirect UV detection) were studied in order to find the optimum conditions for the chiral resolution of the two underivatized amino acids in their enantiomers. Among the two investigated chiral selectors, vancomycin resulted to be the most useful chiral selector allowing relatively high chiral resolution of the studied compounds even at low concentration. The optimized method (10 mM sorbic acid/histidine, pH 5, and 10 mM of vancomycin) was used for the analysis of real samples such as teeth dentine and beer.     

Chiral separation of amino acids and peptides by capillary electrophoresis

Chiral separation of amino acids and peptides by capillary electrophoresis (CE) is reviewed regarding the separation principles of different approaches, advantages and limitations, chiral recognition mechanisms and applications. The direct approach details various chiral selectors with an emphasis on cyclodextrins and their derivatives, antibiotics and chiral surfactants as the chiral selectors. The indirect approach deals with various chiral reagents applied for diastereomer formation and types of separation media such as micelles and polymeric pseudo-stationary phases. Many derivatization reagents used for high sensitivity detection of amino acids and peptides are also discussed and their characteristics are summarized in tables. A large number of relevant examples is presented illustrating the current status of enantiomeric and diastereomeric separation of amino acids and peptides. Strategies to enhance the selectivity and optimize separation parameters by the application of experimental designs are described. The reversal of enantiomeric elution order and the effects of organic modifiers on the selectivity are illustrated in both direct and indirect methods. Some applications of chiral amino acid and peptide analysis, in particular, regarding the determination of trace enantiomeric impurities, are given. This review selects more than 200 articles published between 1988 and 1999.     

Comparison of performance of Chirobiotic T, T2 and TAG columns in the separation of beta2- and beta3-homoamino acids

The enantiomers of eight unusual beta(2)- and beta(3)-homoamino acids were separated on chiral stationary phases containing the macrocyclic glycopeptide antibiotic teicoplanin (Chirobiotic T or T2) or teicoplanin aglycone (Chirobiotic TAG) as chiral selectors. The effects of the organic modifier, the mobile phase composition, and temperature on the separations were investigated. Linear van't Hoff plots were observed in the studied temperature range, 280-318 K, and the changes in enthalpy, Delta(DeltaH(o)), entropy, Delta(DeltaS(o)), and free energy, Delta(DeltaG(o)) were calculated. The values of the thermodynamic parameters depended on the nature of the selectors, the structures of the analytes, and especially the positions of the substituents on the analytes. A comparison of the separation performances of the macrocyclic glycopeptide stationary phases revealed that the Chirobiotic TAG column exhibited much better selectivity for beta(2)-homoamino acids, while the separation of beta(3)-homoamino acid enantiomers was better on Chirobiotic T or T2. The elution sequence was determined in some cases and was observed to be R < S.     

Retention mechanism of high-performance liquid chromatographic enantioseparation on macrocyclic glycopeptide-based chiral stationary phases

The development of methods for the separation of enantiomers has attracted great interest in the past 20 years, since it became evident that the potential biological or pharmacological applications are mostly restricted to one of the enantiomers. In the past decade, macrocyclic antibiotics have proved to be an exceptionally useful class of chiral selectors for the separation of enantiomers of biological and pharmacological importance by means of high-performance liquid chromatography (HPLC), thin-layer chromatography and electrophoresis. The glycopeptides avoparcin, teicoplanin, ristocetin A and vancomycin have been extensively used as chiral selectors in the form of chiral bonded phases in HPLC, and HPLC stationary phases based on these glycopeptides have been commercialized. In fact, the macrocyclic glycopeptides are to some extent complementary to one another: where partial enantioresolution is obtained with one glycopeptide, there is a high probability that baseline or better separation can be obtained with another. This review sets out to characterize the physicochemical properties of these macrocyclic glycopeptide antibiotics and, through their application, endeavors to demonstrate the mechanism of separation on macrocyclic glycopeptides. The sequence of elution of the stereoisomers and the relation to the absolute configuration are also discussed.