Achiral separations

FIGURE 14.3 (a) Achiral separation of NBD-alanine in the rat cerebrum, (b) The chiral... 

With the increased popularity of LC-MS, the problem of overlapping enantiomer peaks from other amino acids has largely been resolved. The mass spectrometer can act as an additional dimension of separation (based on mass to charge ratio). Thus, only amino acids having the same mass-to-charge ratio must be separated achirally (see Desai and Armstrong, 2004). This additional dimension of separation also has implications for the applications in the matrices discussed previously. With the ability of the mass spectrometer to discriminate on the basis of mass, this lessens the need for complete achiral separation. For example, an LC-MS method was recently developed to study the pharmacokinetics of theanine enantiomers in rat plasma and urine without an achiral separation before the enantiomeric separation (Desai et al., 2005). In such matrices, proteins must still be removed by appropriate sample preparation. 

FIGURE 14.4 Achiral separation (a) of rye grass extract containing 2-(2,4-diclorophenoxy) propionoic acid (2,4-DP) on a C18 column and subsequent chiral separation (b) of the heart-cut portion on a Chirobiotic T CSP. Reprinted from Schneiderheinze et al. (1999) with permission John Wiley Sons. 

FIGURE 14.5 Separations involving voriconazole (1), its mirror image (2), related diaster-eomers (3), chlorinated impurities (4), and an achiral impurity 5. (a) Achiral separation of compounds 1-5 on an amino column with hexane/ethanol mobile phase (b) Chiral separation of compounds 1-5 on Chiralpak As column with hexane/ethanol mobile phase (c) Achiral-chiral multidimensional separation with the amino and chiral column coupled in series. Reprinted from Ferretti et al. (1998) with permission from Vieweg Verlag. 

G. N. Okafo and J. K. Roberts, Development of achiral separation methods in pharmaceutical analysis. Pharmaceutical Analysis, 2003, 31-73. 

Achiral (separation of diastereomeric derivatives) and chiral (sepa-ration of enantiomers) chromatography gas chromatography high-performance liquid chromatography super- and sub-critical fluid chromatography thin-layer chromatography... 

Sanger-van de Griend et al. (29) determined the binding constants of several local anaesthetics with DM-/3-CD. These data showed that the achiral separation of analogues is a result of their mobility difference, whereas the resolution of enantiomers results from the difference in their binding constants with CDs. 

Enantiomeric pnrity assays have also been performed without chromatographic separation being conducted prior to detection, for example, with circular dichroism (CD) and MS. Bertncci et al. [110] developed a chiral assay for pulegone, oxazepam, and warfarin by combining simnltaneons UV, CD, and g factor detection on an achiral separation system with a Hypersil CN colnmn and a mobile phase of hexane 2-PrOH (90 10). The precision (RSD%) of the method ranged from 0.6% to 2.6%, and the LOQs were between 0.1% and 1% (0.2-2.2 j,g). For fnrther information concerning the application of CD and polarometric detection for chiral detection, see the review by Bobbitt and Linder [111]. 

Many of the modes of CEC illustrated in Table 3.1 are applicable to both gradient and isocratic elution, aqueous and non-aqueous conditions, as well as to chiral and achiral separations and these will be discussed within the appropriate sections. The complex mechanisms responsible for selectivity will not be discussed, rather this chapter will be limited to describing the scope for application of the different CEC modes. 

Wallenborg SR, Lurie IS, Arnold DW, Bailey CG. On-chip chiral and achiral separation of amphetamine and related compounds labeled with 4-fluoro-7-nitro-benzofurazane. Electrophoresis 2000 21 3257-3263. 

Temperature. Especially the latter parameter should not be underestimated. Although, as a rule of thumb, achiral separations are often performed at elevated temperatures, it is generally believed that separations on chiral stationary phases should best be performed at lower temperatures. Nevertheless, sometimes it turns out that chiral separations are entropy controlled and better selectivities are obtained at higher temperatures [13-16]. 

Whereas the separation of racemates on these two CSP are obvious, recent applications demonstrate that achiral isomers, especially aromatic compounds with substituents in different positions, are extraordinarily well separated on Chiralcel-OD and Chiralpak-AD as well (Figure 21-9). It is to be expected that further examples will follow and more and more achiral separation problems will be solved in the future on CSP. 

Capillary approaches have been shown to be useful for many chiral separations as well as achiral separations. For chiral separations, separation buffer additives containing chirogenic centers (tecoplainin, erythromycin, vancomycin, or cyclodextrans) have facilitated the resolution of enantiomers [26,30,31]. Chiral capillary separations could readily be combined with mass spectrometry because the volume of effluent moving from the separation capillary to the ion source is small and makeup solvent is commonly added by means of an union to stabilize the ion beam. Chiral capillary separations provide an attractive alternative to analytical-scale normal-phase separations when using atmospheric pressure ion-ization mass spectrometry. 

FITC-labeled amino acids as model compounds were separated in sodium dodecyl sulfate/borate buffer at pH 9.4 with y-cyclodextrin as chiral discriminator. Separation speed achieved with the microchip was one order of magnitude faster than that with conventional CE. Bailey and co-workers demonstrated chiral and achiral separation of amphetamine and analogous compounds labeled with 4-fluoro-7-nitrobenzofurazane on an S-folded separation channel. Phosphate buffer with sulfated y-cyclodextrin and sodium dodecyl sulfate as additives was used. It is expected that chiral separations on microfabricated electrophoresis devices with high speed and high throughput will find its way into pharmaceutical and biomedical sciences as well as into other areas. 

SKC is also very useful for achiral separations. Fig ures 2 -7.29-8, and 29-9 illustrate three lypical and tli-versc applications of SFC. Figure 29-7 shows the separation of a scries of dimcihylpolysiloxanc oligomers... 

Development of achiral separation methods in pharmaceutical analysis... 

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