CMPAs

Chiral resolution by HPLC can by divided into three categories (1) a direct resolution using a chiral stationary phase (CSP) (2) addition of a chiral agent to the mobile phase, which reacts with the enantiomeric analytes (chiral mobile phase additive method (CMPA)) (3) an indirect method that utilizes a precolumn diastereomer formation with a chiral derivatization reagent (Misl anova and Hutta, 2003). 

Analysis using a CMPA is usually resolved on a nonchiral column. A transient diastereomeric complex is formed between the enantiomer and the chiral component in the mobile phase, similar to the complexes formed with chiral stationary phases. A review by Liu and Liu (2002) cites several papers where addition of CPMAs has been used in analyzing amphetamine-related compounds. Some CPMAs include amino acid enantiomers, metal ions, proteins, and cyclodextrins. Advantages of this method of analysis include the use of less expensive columns and more flexibility in the optimization of chiral separation (Misl anova and Hutta, 2003). 

In recent years, for analytical purposes the direct approach has become the most popular. Therefore, only this approach will be discussed in the next sections. With the direct approach, the enantiomers are placed in a chiral environment, since only chiral molecules can distinguish between enantiomers. The separation of the enantiomers is based on the complex formation of labile diastereoisomers between the enantiomers and a chiral auxiliary, the so-called chiral selector. The separation can only be accomplished if the complexes possess different stability constants. The chiral selectors can be either chiral molecules that are bound to the chromatographic sorbent and thus form a CSP, or chiral molecules that are added to the mobile phase, called chiral mobile phase additives (CMPA). The combination of several chiral selectors in the mobile phase, and of chiral mobile and stationary phases is also feasible. 

The development of a plethora of HPLC CSPs in the 1980s and 1990s has, to a large extent, made the use of chiral mobile-phase additives (CMPAs) redundant in most modem pharmaceutical analytical laboratories [23]. Before this period, chiral selectors were used routinely as additives in HPLC, but are now only used for a small number of specific applications [23]. CMPAs are used to form... 

Since one or more of the interactions in these systems might originate from the stationary phase, only a two- or a one-point interaction between the solute and the selector is necessary for mechanisms (2) and (3) to occur [50]. However, some of the CMPAs used in HPLC [37,40,51,52] have also been used as chiral selectors in CE [53-56], which indicates that at least one of the separation mechanisms between the selector and enantiomers is selective complex formation in the mobile phase in these cases, since there is no stationary phase present in CE. A recent example by Yuan et al. [57] is presented in Eigure 17.1. The authors introduced the use of (R)-A,A,A-trimethyl-2-aminobutanol-bis(trifluoromethane-sulfon)imidate as the chiral selector for enantioseparation in HPLC, CE, and GC. This chiral liquid serves simultaneously as a chiral selector and a co-solvent. 

The use of CMPA is flexible and is convenient for exploring new chiral selectors. The stationary phases used for the CMPAs are less expensive than CSPs, whereas the additives are often quite expensive. Furthermore, the complex mobile phase often limits the choice of detection method (e.g., mass spectrometry [MS]) that could be used, which makes the CMPAs less commonly used than CSPs. Only a few applications have been published during the last 10 years [39,58-60]. A recent example with a chiral selector used as both the CSP and the CMPA is shown in Figure 17.2 [43]. For further reviews on the use of CMPA, see Refs. [35,40,49]. 

A majority of the chiral purity assays made with HPLC published during the last decade are based on separation on CSPs and subsequent UV detection (Table 17.3). The polysaccharide phases seem to be the dominating CSPs, but there is an even distribution in the methods that uses normal-and reversed-phase modes. A few of the methods utilize CMPA [39,59,60] or indirect separation by chemical derivatization [16,18]. However, it seems that the majority of the published papers... 

Table 17.3 contains only few examples of where CMPA have been applied for pharmaceutical analysis. The majority of selectors that have been used are different types of cyclodextrines [39,60], but there is also a macrocyclic antibioticum in the list [59]. Guo et al. [59] applied norvancomycin as a CMPA for an assay of (S)-ketoprofen in capsules, the results are displayed in Eigure 17.6B. They... 

FIGURE 17.6 Norvancomycin used as the CMPA. Column Hypersil BDS, mobile phase AcN 20mM TEA buffer (pH 5.2) containing 2.0mM norvancomycin (35 65), (A) racemic solution of ketoprofen and (B) (5)-ketoprofen formulation. (Reprinted from Guo, Z.S. et al., J. Pharm. Biomed. Anal., 41, 310, 2006. Copyright Elsevier, 2006. With permission.)... 

CMPAs have rarely been used for chiral analysis in biological matrixes, and have, to the authors knowledge, not been applied during the last decade. One of the reasons is probably the incompatibility between involatile CMPAs and MS detection. For recent reviews on chiral pharmaceutical analysis by LC/MS, see Refs. [114,116,119,120]. 

CSP. CMPA proteins, polypeptides Synthetic Polymers based on analytical and/or bead material... 

CMPA, CE MEKC CSP e.g. cyclodextrins, amino acids cholic acids, tartaric acids, alkaloids etc. and Derivatives Thereof analytical to preparative excellent, fair to moderate... 

CSP, CMPA Newly Designed Small Molecules analytical to preparative excellent to moderate... 

Beads = pure polymeric particles with similar chiral information to the corresponding sorbent (CSp) coated on silica gel CE = capillary electrophoresis CSP = chiral stationary phase CMPA — chiral mobile phase additive MEKC = micellar electrokinetie capillary chromatography. 

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. 

In contrast to the various CSPs mentioned so far, but still based on covalently or at least very strongly adsorbed chiral selectors (from macromolecules to small molecules) to, usually, a silica surface, the principle of dynamically coating an achiral premodified silica to CSPs via chiral mobile phase additives (CMPA) has successfully been adapted for enantioseparation. The so-called reverse phase LC systems have predominantly been used, however, ion-pairing methods using nonaqueous mobile phases are also possible. 

In the CMPA mode, additional equilibria must be considered (1) the reversible mixed complex formation of SO and SAs (2) the adsorption/desorption processes of the SO added to the mobile phase with the stationary phase (3) the adsorption/desorption processes of the [SO-SA] complexes with the stationary phase and also the adsorption/desorption of the noncomplexed SA analytes (see Scheme 2). 

The overall observed retention of the enantiomers, and thus the elution order, is based on several kinetically and thermodynamically controlled parameters concerned with stereorecognition nonstereoselective interactions of all partners SO(R), SA(R S), and particularly of the [SO(RI-SA(KI] and [SO(K)-SA(Si] complexes with the achiral stationary phase, also play a role (Figure 21). Therefore the retention order may be reversed for a specific pair of enantiomers depending on whether a covalently bound CSP or a CMPA is applied, but using the same chiral molecule (part) as chiral selector. These general principles, shown schematically for a CLEC system, are further complicated by the complexity of the entire system, hence they are difficult to anticipate and each case must be studied individually. 

Case II dynamically coated CSP via a chiral mobile phase additive = (S)-CMPA ... 

However, the CMPA concept for CLEC. introduced by Lindner and co-workers139 for the resolution of dansyl amino acids using a chiral triamine and Zn(II) as transient metal ions parallel to the work of Gil-Av146, allowed the further development and successful use of amino acids and their derivatives together with Cu(II) as CMPAs in reverse-phase systems for the direct resolution of, for example, a-amino acids146. 

The logical extension of the ion-pair chromatographic technique, using chiral counterions as CMPAs has been impressively applied, for the first time, to the resolution of chiral amino alcohols (betablockers) by Pettersson and co-workers142. 

CMPA as SO many other amino acid derivatives have been used as CMP As... 

On the other hand, the direct chromatographic approach involves the use of the chiral selector either in the mobile phase, a so-called chiral mobile phase additive (CMPA), or in the stationary phase [i.e., the chiral stationary phase (CSP)]. In the latter case, the chiral selector is chemically bonded or coated or allowed to absorb onto a suitable solid support. Of course chiral selectors still can be used as CMPAs, but the approach is a very expensive one owing to the high amount of chiral selector required for the preparation of the mobile phase, and the large amount of costly chiral selector that is wasted (since there is very little chance of recovering this compound). Moreover, this approach is not successftd in the preparative separation of the enantiomers. 

---