Chiral selectors in chromatography

GLYCOPEPTIDES AS CHIRAL SELECTORS IN CHROMATOGRAPHY AND CAPILLARY ELECTROPHORESIS... 

Thin-Layer Chromatography. Chiral stationary phases have been used less extensively in tic as in high performance Hquid chromatography (hplc). This may, in large part, be due to lack of avakabiHty. The cost of many chiral selectors, as well as the accessibiHty and success of chiral additives, may have inhibited widespread commerciali2ation. Usually, nondestmctive visuali2ation of the sample spots in tic is accompHshed using iodine vapor, uv or fluorescence. However, the presence of the chiral selector in the stationary phase can mask the analyte and interfere with detection (43). 

Capillary electrophoresis employing chiral selectors has been shown to be a useful analytical method to separate enantiomers. Conventionally, instrumental chiral separations have been achieved by gas chromatography and by high performance liquid chromatography.127 In recent years, there has been considerable activity in the separation and characterization of racemic pharmaceuticals by high performance capillary electrophoresis, with particular interest paid to using this technique in modem pharmaceutical analytical laboratories.128 130 The most frequently used chiral selectors in CE are cyclodextrins, crown ethers, chiral surfactants, bile acids, and protein-filled... 

When micelles are used, the CE technique becomes a micellar elec-trokinetic chromatography (MEKC) one. Natural surfactants, such as bile salts, digitonin and saponins, optically active synthetic surfactants, e.g., amino-acid derived ones, alkylglycoside-, tartaric acid- and steroidal glucoside-based surfactants, and high-molecular mass or polymerized surfactants, have been used as chiral selectors in In the lat-... 

Aiken, J.H., Huie, C.W. (1993). Use of microemulsion system to incorporate a lipophilic chiral selector in electrokinetic capillary chromatography. Chro-matographia 35 448-450. 

For obvious reasons CDs (and other dextrins) are potentially good chiral selectors for chromatography on the one hand they can be used as mobile phase additives (CMPA) in TLC45, HPLC46 and CE47 49 and on the other they can be covalently bonded onto solid supports50,51 and silica gei 52- 54 xhis approach can be extended to the preparative resolution of enantiomers41,55,56. 

Using amylose tris-3,5-dimethylphenylcarbamate as the chiral selector in enantioselective high-performance liquid chromatography, micropreparative resolution of the DHA racemate was achieved and the chromatographic behaviour in enantio-GC could be defined by coinjecting these references of definite chirality (Fig. 17.4) [13]. 

Although all proteins are complex in structure and chiral in nature, some of them could achieve the status of a chiral selector in liquid chromatography. The complex structures of proteins are the result of the different intramolecular hydrogen-bonding, disulfide bridges, and other types of bonding. All of the proteins used for chiral resolution in liquid chromatography are obtained from animals except for cellobiohydrolase-I. The structures and properties of some of the most commonly used proteins as chiral selectors are discussed herein. 

Ovotransferrin is also obtained from the white portion of a chicken egg and has been used as a chiral selector in liquid chromatography. This protein is also called conalbumin. It is a metal ion (iron, copper, manganese, and zinc) binding protein of molecular mass 70,000-78,000 and with an isoelectric point of 6.1-6.6. This protein is sensitive to acids and heat. 

This protein is extracted from pancreatic tissues. This protein occurs in a-form and /1-form, but the a-form is used as chiral selector in liquid chromatography. The molecular mass is 25,000, with an isoelectric point of 8.1-8.6. It is inhibited by metal ions. The protein is useful for chiral resolution of amino acids and amino esters. 

As discussed earlier, the proteins used as chiral selectors in affinity chromatography cannot be used under the high-pressure HPLC with a variety of mobile phases therefore, these proteins were immobilized with some solid support such as hydroxyethylmethacrylate, polystyrene-divinylbenzene, polyethylene fibers, and silica gel [14,15]. Avariety of techniques have been used for the immobiliza-... 

Clark BJ, Separations of enantiomeric compounds by chiral selectors in the mobile or solvent phase, in A Practical Approach to Chiral Separations by Liquid Chromatography, Subramanian G (Ed.), VCH, Weinheim, p. 311 (1994). 

There are three commonly used approaches to the separation of enantiomers by HPLC. Of these, two are based on operation with conventional reversed-phase column materials.72 One approach is to derivatize the sample prior to chromatography, leading to formation of diastereomeric products of the two enantiomers that can be separated by conventional HPLC,73,74 whereas the other uses chiral selectors in the mobile phase.75 76 The third approach involves the use of chiral stationary phases. 

As in the case of chromatography, a chiral selector is also required in CE for enantiomeric resolution. Generally, suitable chiral compounds are used in the background electrolyte (BGE) as additives and hence are called chiral selectors or chiral BGE additives. There are only a few publications available that deal with the chiral resolution on a capillary coated with the chiral selector in CE. The analysis of the chiral pollutants discussed in this chapter is restricted only to using chiral selectors in the BGE. The most commonly used chiral BGE additives are cyclo-dextrins, macrocyclic glycopeptide antibiotics, proteins, crown ethers, ligand exchangers, and alkaloids.A list of these chiral BGE additives is presented in Table 1. 

Thin layer chromatography is also used for direct enantiomeric resolution of D,L-arginine, D,L-histidine, d,l-lysine, D,L-valine, and D,L-leucine on silica gel plates impregnated with optically pure (IR, 3R, 5R)-2-azabicy-clo[3,3,0]octan-3-carboxylic acid, which serves as a chiral selector in the pharmaceutical industry. To successfully resolve D,L-amino acids, various combinations of aceto-nitrile-methanol-water were proposed. The spot was detected by ninhydrin (0.2% in acetone). 

The first applications of CDs as chiral selectors in CE were reported in capillary isotachophoresis (CITP) [2] and capillary gel electrophoresis (CGE) [3]. Soon thereafter, Fanali described the application of CDs as chiral selectors in free-solution CE [4] and Terabe used the charged CD derivative for enantioseparations in the capillary electrokinetic chromatography (CEKC) mode [5]. It seems important to note that although the experiment in the CITP, CGE, CE, and CEKC is different, the enantiomers in all of these techniques are resolved based on the same (chromatographic) principle, which is a stereoselective distribution of enantiomers between two (pseudo) phases with different mobilities. Thus, enantioseparations in CE are commonly based on an electrophoretic migration principle and on a chromatographic separation principle [6]. 

As shown in Eq. (2) together with the chiral recognition Kk + Kg), the other necessary requirement for enantioseparations in CE is a mobility difference between the free and the complexed analyte fXf - fjL, 0). Otherwise, it will be impossible to transfer a chiral recognition into a chiral separation. This requirement does not hold when neutral analytes are analyzed with neutral chiral selectors. In such a case, an additional buffer component is required that will assist in generating a difference between the mobilities of an analyte in its free and complexed forms with a chiral selector. This is achieved by an achiral micellar phase in cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) [9]. However, a charged CD or a chiral micellar phase can combine the... 

Capillary Electrophoresis (CE) and Micellar Electrokinetic Chromatography (MEKC). - Di(2-ethylhexyl) thiophosphoric acid (DEHTPA) has been earlier characterised by potentiometric titration, and quantified by capillary zone electrophoresis with carbonate buffer, operating at —20 kV, and using UV detection at 210 nm. ° Also, a comparison has been made of capillary electrophoresis (CE) and liquid chromatography (LC) for the enantiomeric separation of a-phosphonosulfonic acids, where CE used 3-cyclodextrin as chiral selector in a borate electrolyte. Alkylphosphonic acids, at trace levels in water, have been determined by CE coupled online with flame photomeric detection, and alkylphosphonic acid esters have been separated and determined by CE using indirect UV detection. 

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