Chirobiotic CSPs

In recent years, the application of macrocyclic glycopeptide CSPs has been expanded to SFC, and teicoplanin aglycone phase (Chirobiotic TAG) was found to be the most effective closely followed by teicoplanin CSP (Chirobiotic T) [144, 184]. Because of a large number of polar groups available on the Chirobiotic CSPs, it is often required to work with considerable amount of polar additives in the mobile... 

The purpose of this study is only intended to illustrate and evaluate the decision tree approach for CSP prediction using as attributes the 166 molecular keys publicly available in ISIS. This assay was carried out a CHIRBASE file of 3000 molecular structures corresponding to a list of samples resolved with an a value superior to 1.8. For each solute, we have picked in CHIRBASE the traded CSP providing the highest enantioselectivity. This procedure leads to a total selection of 18 CSPs commercially available under the following names Chiralpak AD [28], Chiral-AGP [40], Chiralpak AS [28], Resolvosil BSA-7 [41], Chiral-CBH [40], CTA-I (microcrystalline cellulose triacetate) [42], Chirobiotic T [43], Crownpak CR(-i-) [28], Cyclobond I [43], DNB-Leucine covalent [29], DNB-Phenylglycine covalent [29], Chiralcel OB [28], Chiralcel OD [28], Chiralcel OJ [28], Chiralpak OT(-i-) [28], Ultron-ES-OVM [44], Whelk-0 1 [29], (/ ,/ )-(3-Gem 1 [29]. 

The macrocyclic antibiotic-based CSPs have not been used extensively in SFC. Two macrocyclic antibiotic CSPs, Chirobiotic T and Chirobiotic V, were included in a study of various CSPs in SFC. At least partial resolution of approximately half of the 44 test compounds could be obtained on these two CSPs in SFC [63]. A high concentration of modifier was necessary to elute some of the analytes. Enantioreso-lution of derivatized amino acids was also demonstrated in the same study. Flowever, a complex modifier comprised of methanol, water, and glycerol was required for separations performed on the Chirobiotic T CSP. The separation of coumachlor enantiomers on a vancomycin-based CSP (Chirobiotic V) in SFC is illustrated in Fig. 12-5 [32]. 

Fig. 12-5. Separation of coumachlor enantiomers on a Chirobiotic V CSP. Chromatographic conditions 15 % methanol in carbon dioxide, 2.0 mL min f 20 MPa, 30 °C.

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. 

Three types of macrocyclic antibiotic CSPs have been commercialized by Astec (Whippany, New York) and show complementary enantioselec-tivity Chirobiotic V (selector vancomycin), Chirobiotic T (teicoplanin)... 

The use of macrocyclic antibiotics as chiral selectors for HPLC was first proposed by Armstrong et al. [50] in 1994. The most successful of the CSPs are based on the glycopeptide antibiotics vancomycin, teicoplanin and ristocetin A and are commercially available through Advanced Separation Technologies Inc. (Astec Inc.) as Chirobiotic V , Chirobiotic 1 and Chirobiotic R , respectively. More recently, a number of other derivatives of these antibiotics have also been developed offering different stereoselectivities. A comprehensive handbook is now available from Astec Inc. [51 ] alongside a number of recent review articles... 

FIGURE 6 Effect of mobile phase composition on the resolution of enantiomers of different racemates in normal phase HPLC on antibiotic CSPs. (a) First ( ) and second ( ) eluted enantiomers of y-phenyl-y-butyrolactone and first ( ) and second (O) eluted enantiomers of 4-phenyl-2-methoxy-6-oxo-2,4,5,6-tetrahydropyridine-3-carbonitrile on Chirobiotic T column and (b) first ( ) and second (O) eluted enantiomers of mephenytoin on Chirobiotic V column. (From Refs. 1 and 21.)... 

TABLE 3 Effect of pH on the Chiral Resolution of Several Racemates on Chirobiotic T CSP Using Methanol-1% Triethylammoniurn Acetate Buffer (20 80, v/v) as the Mobile Phase... 

FIGURE 9 Effect of mobile phase pH on the chiral resolution of mandelic acid on Chirobiotic T CSP using ethanol-water-TEA (17.5 70.2 12.3, v/v/v) as the mobile phase. (From Ref. 25.)... 

TABLE 5 Effect of Flow Rate on Enantiomeric Resolution on Chirobiotic V and Chirobiotic T CSPs... 

FIGURE 10 Effect of temperature on enantiomeric resolution on antibiotic CSPs. (a) k, a, and Rs for proglumide (O), 5-methyl-5-phenylhydantoin ( ) and iV-corbyl-DL-pheny-lalanine (x) on Chirobiotic V column using acetonitrile-1 % triethylammonium acetate buffer (10 90, v/v) as the mobile phase and (b) separation of enantiomers of /1-methyl phenylalanine on the Chirobiotic T column using water-methanol (10 90, v/v) as the mobile phase at (A) 1°C, (B) 20°C, (C) 50°C. 1 = erythro-L 2 = erythro-D 3 = threo-L 4 = threo-D. (From Refs. 1 and 22.)... 

The teicoplanin CSP (Chirobiotic T) exhibits enantioselectivity for underivatized and Mderivatized (FMOC or Z) amino acids, hydroxy carboxylic acids and other chiral acids including chiral phenols, small peptides, neutral aromatic analytes and cyclic aromatic and aliphatic amines [285] (see also Table 9.11). Selection of the mobile phase mcxle (reversed-phase, normal-phase, or polar-organic phase mode) follows the same criteria as described for vancomycin CSP. 

In addition to the vancomycin and teicoplanin CSPs, ristocetin A (Chirobiotic R) [289] and recently avoparcin [280] have been evaluated as novel chiral SOs and CSPs. It turned out that within the large family of macrocyclic antibiotics complementarity of enantioselectivity exists for different glycopeptides. As a consequence, very often it is possible to obtain a complete resolution by switching to a congeneric antibiotic CSP, if after optimization no baseline, but partial. separation can be achieved on a certain macrocyclic antibiotic type CSP (see Fig. 9.22). It can be expected that the enantioselectivity potential of closely related antibiotics will be further exploited in the future leading to an increase in the number of macrocyclic antibiotic type CSPs. 

Macrocyclic glycopeptide antibiotic based CSPs Chirobiotic R, Chirobiotic T, Chirobiotic V, Chirobiotic TAG and Chirobiotic modified V Advanced Separation Technologies, Inc., Whippany, NJ, USA... 

Compatible CSPs Daicel, Whelk Daicel, Whelk and others Daicel cydodextrin Chirobiotic... 

More immobilized cellulose and amylose phases are currently tmder development at Daicel. Daicel is promoting the concept of a CSP library of more than 100 stationary phases that are not currently commercially available, which can be screened for enantiomer separations. Astec has recently presented an improved linking of antibiotic phases (Chirobiotic). As yet, no chiral monolithic stationary phases are commercially available, although one can assume that various companies are currently endeavoring to change the surface of monolithic material with chiral modifiers. 

Now that the e, s, a, b, and v system parameters are known for the four Chirobiotic columns and the A, B, E, S, and V solute descriptors are obtained for five enantiomeric compounds, it is possible to calculate the retention factor k of the five enantiomeric compounds using Eq. (5) and compare it with the experimental two retention factors obtained for the two enantiomers on the CSP. Figure 7 illustrates the results three situations were observed, (i) The retention factors predicted by LSER corresponded to the first eluting enantiomer (e.g., dihydrofurocoumarin. Fig. 7a) (ii) the LSER-predicted retention factors corresponded to the last eluting enantiomer (e.g., 5-methyl-5-phenyl hydantoin in the normal-phase mode. Fig. 7b) and (iii) the LSER-predicted retention factors did not correspond to a particular enantiomer for all mobile phase compositions (e.g., 5-methyl-5-phenyl hydantoin in the RPLC mode and bromacil. Fig. 7c and d). From a mechanistic point of view, it can be speculated that in case i, the chiral selector has overall attractive enan-tioselective interactions with the second enantiomer more retained than the LSER prediction in case ii, the chiral selector has overall repulsive enantioselective interactions with the first enantiomer less retained than the LSER prediction and in case iii, the chiral selector has enantioselective interactions with both enantiomers. 

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