Liquid chromatography optical isomers

One application in liquid chromatography which does alter the separation process is the use of a specific series of derivatives to enable the separation of chiral (optical isomers) forms of alcohols, amines and amino acids using reverse-phase separation. FLEC is available in the two chiral forms (+)-l-(9-fluorenyl) ethyl chloroformate and (—)-l-(9-fluorenyl) ethyl chlorofor-mate (Figure 3.12). Reaction of two stereoisomers of a test compound (e.g. T+ and T—) with a single isomer of the derivatizing reagent (e.g. R+) will result in the formation of two types of product, T+R+ and T—R+. It is possible to separate these two compounds by reverse-phase chromatography. 

Normal-phase liquid chromatography is thus a steric-selective separation method. The molecular properties of steric isomers are not easily obtained and the molecular properties of optical isomers estimated by computational chemical calculation are the same. Therefore, the development of prediction methods for retention times in normal-phase liquid chromatography is difficult compared with reversed-phase liquid chromatography, where the hydrophobicity of the molecule is the predominant determinant of retention differences. When the molecular structure is known, the separation conditions in normal-phase LC can be estimated from Table 1.1, and from the solvent selectivity. A small-scale thin-layer liquid chromatographic separation is often a good tool to find a suitable eluent. When a silica gel column is used, the formation of a monolayer of water on the surface of the silica gel is an important technique. A water-saturated very non-polar solvent should be used as the base solvent, such as water-saturated w-hexane or isooctane. 

Jandera, R et al.. Effect of the mobile phase on the retention behaviour of optical isomers of carboxylic acids and amino acids in liquid chromatography on bonded teicoplanin columns, J. Chromatogr. A, 917, 123, 2001. 

Immobilization. The ability of cyclodextrins to form inclusion complexes selectively with a wide variety of guest molecules or ions is well known (1,2) (see Inclusion COMPOUNDS). Cyclodextrins immobilized on appropriate supports are used in high performance liquid chromatography (hplc) to separate optical isomers. Immobilization of cyclodextrin on a solid support offers several advantages over use as a mobile-phase modifier. For example, as a mobile-phase additive, p-cyclodextrin has a relatively low solubility. The cost of y- or a-cyclodextrin is high. Furthermore, when employed in thin-layer chromatography (dc) and hplc, cyclodextrin mobile phases usually produce relatively poor efficiencies. 

Traditionally, chiral separations have been considered among the most difficult of all separations. Conventional separation techniques, such as distillation, liquid—liquid extraction, or even some forms of chromatography, are usually based on differences in analyte solubilities or vapor pressures. However, in an achiral environment, enantiomers or optical isomers have identical physical and chemical properties. The general approach, then, is to create a "chiral environment" to achieve the desired chiral separation and requires chiral analyte—chiral selector interactions with more specificity than is obtainable with conventional techniques. 

Deacetamidocolchicine (18) was prepared by the classic syntheses of Eschenmoser (38) and van Tamelen (39). The racemate was resolved by medium-pressure liquid chromatography on swollen, microcrystalline cellulose triacetate prepared at 5°C, and the enantiomers were collected at -70°C (32). The (-)-enantiomer with the same biaryl configuration as natural (aS,7S)-colchicine (1) was eluted first and found to be essentially optically pure. Thermal racemization of the optical isomers gave the ther-... 

Armstrong, D.W., Optical isomer separation by liquid chromatography, Anal. Chem., 59, 84A, 1987. 

A series of 1,3,2-benzodithiazole J-oxides have been prepared by oxidation of 2-benzyl- and 2-phenylethyl-l,3,2-benzodithiazoles 48 <1996ENA13>. One equivalent of MCPBA leads to mono-3 -oxide 49, whereas oxidation with an excess of MCPBA yielded a mixture of dioxide 50 and trioxide 51, which were isolated as optically pure isomers by chiral liquid chromatography (Scheme 2). 

K. Shimizu, T. Kakimoto, K. Ishi, Y. Fujimoto, H. Nishi, and N. Tsumagari, New derivatization reagent for the resolution of optical isomers in diltiazem hydrochloride by high-performance liquid chromatography, J. Chromotogr., 357 119 (1986). 

Nagai, T., Kamiyama, S., Nagai, T., Forensic Toxicologic Analysis of Methamphetamine Optical Isomers by High Performance Liquid Chromatography, Z. Rechtsmed., 101,151,1988. 

W. H. Pirckle, D. W. House, and J. M. Firm. Broad spectrum resolution of optical isomers using chiral high performance liquid chromatography bonded phases. J. Chromatogr. A 192 (1980) 143. 

LePage, J.N. Lindner, W. Davies, G. Seity, D.E. Karger, B.L. Resolution of the optical isomers of dansyl amino acids by reversed phase liquid chromatography with optically active metal chelate additives. Anal. Chem. 1979, 51 (3), 433-435. 

Weems, H. B. and Yang, S. K. (1982) Resolution of optical isomers by chiral high performance liquid chromatography separation of dihydrodiols and tetrahydrodiols ofbenzo[a]pyreneandbenz[a]anthracene. na/ Biochem. 125, 156-161. 

Ito, M. Kuriki, T. Goto, J. Nambara, T. Separation of ramipril optical isomers by high-performance liquid chromatography. J.Liq.Chromatogr., 1990, 13, 991-1000... 

The most widely used technique for the estimation of amino acid isomers is gas-liquid chromatography. Two basic strategies have been used to separate isomers by this technique. Both approaches appeared in the mid-1960 s, and both involve derivatiza-tion of the amino acids to suitably volatile acyl-esters (46,47). One method is similar in concept to the separation of disastereo-mers by ion exchange chromatography discussed above. In one step of the two step derivatization, the amino acids are esterified with an optically active agent. This procedure creates molecules with two centers of asymmetry which can be separated on a non-optically active liquid (stationary) phase (46). This method allows any laboratory equipped with a gas chromatograph to perform Isomer analyses. 

---