Separation selectivity with chiral mobile-phase additives

Two mechanisms foe chiral separations using chiral mobile-phase additives, analogous to models developed for ion-pair chromatography, have been proposed lo explain the chiral selectivity obtained using chiral mobile-phase additives. In one model, the chiral mobile-phase additive and the analyle enamiomers form diaslereomeric complexes in solution. As noted previously, diaslereomers may have slightly different physical properties such as mobile phase solubilities or slightly different affinities for the stationary phase. Thus, the chiral separation can be achieved with conventional columns. 

As mentioned above, enantioseparations in EKC rely on a chromatographic separation principle. Despite this fact, there are significant differences between these techniques. Responsible for all differences between chromatographic and electrophoretic enantioseparations is the property of the electrophoretic mobility to be selective for the analytes residing in the same physical phase [2]. Another important point is that in chromatographic techniques, except in the case of a chiral mobile phase additive (CMPA), the analyte is virtually immobile when associated with a chiral selector. In EKC the analyte selector complex is commonly mobile. 

The first chiral separation using pSFC was published by Caude and co-workers in 1985 [3]. pSFC resembles HPLC. Selectivity in a chromatographic system stems from different interactions of the components of a mixture with the mobile phase and the stationary phase. Characteristics and choice of the stationary phase are described in the method development section. In pSFC, the composition of the mobile phase, especially for chiral separations, is almost always more important than its density for controlling retention and selectivity. Chiral separations are often carried out at T < T-using liquid-modified carbon dioxide. However, a high linear velocity and a low pressure drop typically associated with supercritical fluids are retained with near-critical liquids. Adjusting pressure and temperature can control the density of the subcritical/supercritical mobile phase. Binary or ternary mobile phases are commonly used. Modifiers, such as alcohols, and additives, such as adds and bases, extend the polarity range available to the practitioner. 

ABSTRACT. The copper(n) complexes of p-cyclodextrins functionalized with aliphatic or pseudoaromatic amines were used for the chiral recognition of unmodified amino acids. Molecular recognition, assisted by non-covalent interactions, was proved by means of thermodynamic and spectroscopic (c.d., e.p.r. and fluorescence) measurements. A cis-disposition of amino groups seems to assist enantiomeric selectivity. The copper(II)-p-cyclodextrin complexes can be used as mobile phase additives in HPLC to separate enantiomeric mixtures of unmodified aromatic amino acids. 

Typical NP conditions involve mixtures of n-hexane or -heptane with alcohols (EtOH and 2-propanol). In many cases, the addition of small amounts (<0.1%) of acid and/or base is necessary to improve peak efficiency and selectivity. Usually, the concentration of alcohols tunes the retention and selectivity the highest values are reached when the mobile phase consists mainly of the nonpolar component (i.e., n-hexane). Consequently, optimization in NP mode simply consists of finding the ratio n-hexane/alcohol that gives an adequate separation with the shortest possible analysis time [30]. Normally, 20% EtOH gives a reasonable retention factor for most analytes on vancomycin and TE CSPs, while 40% is more appropriate for ristocetin A-based CSPs. Ethanol normally gives the best efficiency and resolution with reasonable backpressures. Other combinations of organic solvents (ACN, dioxane, methyl tert-butyl ether) have successfully been used in the separation of chiral sulfoxides on five differenf glycopepfide CSPs, namely, ristocetin A, teicoplanin, TAG, vancomycin, and VAG CSPs [46]. 

The first and most often encountered separation mechanism in CE is based on mobility differences of the analytes in an electric field these differences are dependent on the size and charge-to-mass ratio of the analyte ion. Analyte ions are separated into distinct zones when the mobility of one analyte differs sufficiently from the mobility of the next. This mechanism is exemplified by capillary zone electrophoresis (CZE) which is the simplest CE mode. A number of other recognized CE modes are variations of CZE. These are micellar electrokinetic capillary chromatography (MECC), capillary gel electrophoresis (CGE), capillary electrochromatography (CEC), and chiral CE. In MECC the separation is similar to CZE, but an additional mechanism is in effect that is based on differences in the partition coefficients of the solutes between the buffer and micelles present in the buffer. In CGE the additional mechanism is based on solute size, as the capillary is filled with a gel or a polymer network that inhibits the passage of larger molecules. In chiral CE the additional separation mechanism is based on chiral selectivity. Finally, in CEC the capillary is packed with a stationary phase that can retain solutes on basis of the same distribution equilibria found in chromatography. 

When compared to the batchwise preparative chromatography, Simulated moving bed (SMB) units exhibit a number of advantages. These advantages are primarily because of the continuous nature of the operation and the efficient use of the stationary and mobile phases, which allows a decrease in desorbent requirement and an improvement of the productivity per unit time and per unit mass of stationary phase. In addition, high performances can be achieved even at rather low values of selectivity and with a relatively small number of theoretical plates. Due to these positive features, SMB is particularly attractive in the case of enantiomer separations, since it is difTicult to separate enantiomers by conventional techniques. More recent applications related to chiral technology were reported [1-3]. 

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