Separator amide-bonded silica

Yoshida [37] has examined amide-bonded silica for peptide separations in HILIC mode. Amide-bonded silica phases such as the TSKgel Amide-80 (Tosoh Bioscience, Tokyo, Japan) are hydrophilic phases containing a carbamoyl group on a short aliphatic spacer. 

Several new phases applied to maltosaccharide analysis that seem to be promising have been described in the literature, for instance, a silica-phase covered with polymeric polyamine resin, and an entirely polymeric resin containing an amide function (35). In any case, partition chromatography is restricted mainly to the utilization of polar-bonded phases, as already described. Nevertheless, there have been a certain number of applications of reversed-phase chromatography that permit relatively simple separations to be achieved. Octadecyl-bonded silica phases are the most widely used, although few applications involve carbohydrate analysis. Their interest lies rather in the analysis of derivated sugars, where the selectivity increases (36). 

In 1998, Machida et al. [45] and Hyun et al. [46] developed a new CCE-based CSP (covalently bonded to silica gel see Sect. 8.2). This CSP was used successfully for the chiral resolution of certain racemic compounds using a variety of mobile phases. The most important applications of this CSP are for the resolution of amino acids, amino esters, amino alcohols, amines, amides, quinolone antibacterials, and other drugs having primary amino groups [46-51,64,65]. The typical chromatograms of the chiral resolution of amino acids on (+)-(18-crown-6)-2,3,ll,12-tetracarboxylic acid CSP are shown in Figure 4. The enantiomeric resolution of the racemic compound on CCE-based CSPs are listed in Table 2. There is no report available on the chiral separations at the preparative scale using these CSPs. 

Many of the chiral molecules containing amide groups were bonded to a solid support for the preparation of CSPs [16-19]. The racemic compounds resolved on these CSPs include a-hydroxycarbonyls, /i-hydroxycarbonyls, amino acids, amino alcohols, amine, and derivatized and underivatized diols. The preliminary chiral diamide phase [(/V-foriuyl-L-valyl)aminopropyl)silica gel] has sufficient separability for racemic /Y-acylatcd a-amino acid esters but not in other types of enantiomer [16]. Most of the eluents used with these CSPs are of normal phase mode, including w-hcxanc, 2-propanol, chlorinated organic solvents, and acetonitrile. 

For characterization and exploitation of the diamide-phase system, a chiral diamide, e,g., (Ill) was examined as a modifier in the mobile phase (solvent) in conjunction with a non-bonded (bare) silica. Such a chiral carrier separated enantiomeric N-acyl-d-amino acid esters and amides with separation factors comparable to those for bonded stationary phase systems. The resolution can be as cribed to diastereomeric complexation through amide-amide hydrogen bonding between the amide additive and enantiomeric solute molecules in the carrier solvent, followed by separation of the diastereomeric complexes by the (achiral) silica phase. This process should be applicable as widely as that involving chiral diamide-bonded stationary phase systems. 

In order to broaden the capabilities of the Pirkle concept, both polar and polarizable groups were introduced into the molecule. The most popular of this type of chiral stationary phase are the (R,R) Whelk-01 and the (S,S)Whelk-01 phases, the structures of which are shown below. These phases are more versatile and have a wider field of application than the phases previously described. The phases are covalently bonded to the silica and so they can be used with almost any type of solvent. However, they have been found to operate most effectively in the normal phase mode. It should be noted that the polarizable character of the aromatic ring is essential for the stationary phase to function well. As the Pirkle phases are generally available in both the (R) and (S) configurations, the reversal of the elution order of a pair of enantiomers is possible. This stationary phase was originally designed for the separation of the Naproxen enantiomers but has found a wide application to the separation of epoxides, alcohols, diols, amides, imides and carbamates. 

Diastereomeric amides are, in general, more readily separated than the corresjronding esters, particularly on normal phase silica or alumina chromatographic systems, where the amide can hydrogen bond with the stationary phase. Helmchen et al. [60] developed a series of concepts to predict the structural features of amides, which would improve separation factors, and also to construct models... 

More recently, several reports have appeared which describe the preparation of HPLC columns which contain CD chemically bonded to silica gel [15-18]. Of these, there are presently two types. The first consists of CD bonded to the silica via amide or amine bonds [15,16] while the second contains no nitrogen linkages [17,18]. This review article summarizes our chromatographic work to date with the latter type of CD bonded phases. In particular, we demonstrate the successful HPLC separation of enantiomers, epimers, cis-trans and other structural isomers as well as important classes of routine compounds by use of a P- or y-CD bonded phase. The obtained chromatographic separations are compared to those obtained on the more conventional normal or reversed phase packings. Additionally, the effect of changes of the pertinent chromatographic variables (such as flow rate, temperature, and solvent composition of the mobile phase) upon the separations are described. Lastly, a brief prospectus on the future of CD bonded phases in HPLC is presented. 

Polyamide as a stationary phase is used mainly for the separation of compounds that can form hydrogen bonds to the amide group. A typical example is the separation of phenolic compounds on polyamide, due to hydrogen bonds between the phenolic hydroxyl and the amide groups [80]. The drawback of these polyamide packings is that they are not as pressure stable as silica and that undesired swelling of the phase can occur with certain solvents. 

Before synthetic chiral stationary phases were developed, attempts were made to use naturally occurring chiral materials for the stationary phase. Quartz, wool, lactose and starch were inadequate but triacetylated cellulose has met with some success. The synthetic stationary phases introduced by Pirkle are able to interact with solute enantiomers in three ways, one of which is stereochemically dependent. Typically these interactions are based on hydrogen bonding, charge transfer (rc-donoi -acceptor based) and steric repulsive types. An independent chiral stationary phase therefore consists of chiral molecules each with three sites of interaction bound to a silica (or other) support. Early work in this area demonstrated that 5-arginine bound to Sephadex would resolve 3,4-dihydroxy-phenylalanine, and that direct resolution of chiral helicenes could be accomplished with columns packed with 2-(2,4,5,7-tetranitro-9-fluorenylideneaminoxy)-propionamide or tri-P-naphthol-diphosphate amide. Amino acid esters have also been resolved with a silica bound chiral binaphthyl crown ether, but better separations are achieved with A-acylated amino acid derivatives with amino-acid derived chiral stationary phases. 

Oi N, Kitahara H, Aoki F, Kisu N (1995) Direct separation of carboxylic acid enantiomers by high-performance liquid chromatography with amide and urea derivatives bonded to silica gel as chiral stationary phases. J Chromatogr A 689(2) 195-201 Oi N, Kitahara H, Matsushita Y, Kisu N (1996) Enantiomer separation by gas and high-performance liquid chromatography with tripeptide derivatives as chiral stationary phases. J Chromatogr A 722(l-2) 229-232... 

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