Proteins as chiral selectors

TABLE 9 Examples of the Chiral Separation of Drugs Using Proteins as Chiral Selectors... 

J Haginaka. Enantiomer separation of drugs by capillary electrophoresis using proteins as chiral selectors. J Chromatogr A 875 235-254, 2000. 

E DeLorenzi, G Massolini. Riboflavin-binding proteins as chiral selectors in HPLC and CE. Pharm Sci Technol Today 2 352-364, 1999. 

Although all proteins are complex in structure and chiral in nature, some of them could achieve the status of a chiral selector in liquid chromatography. The complex structures of proteins are the result of the different intramolecular hydrogen-bonding, disulfide bridges, and other types of bonding. All of the proteins used for chiral resolution in liquid chromatography are obtained from animals except for cellobiohydrolase-I. The structures and properties of some of the most commonly used proteins as chiral selectors are discussed herein. 

Millot M.C., Separation of drug enantiomers by hquid chromatography and capillary electrophoresis, using immobilized proteins as chiral selectors. J. Chromatogr. B, 797,131-159 (2003). 

Armstrong et al. ° first introduced chiral stationary phases based on macrocyclic antibiotics. Vancomycin, ristocetin A, teicoplanin, avoparcin, rifamycin B and thiostrepton are used as chiral selectors. They posses a broad enantiorecognition range, similar to protein based CSPs. However, CSPs based on macrocyclic antibiotics show higher stability and capacities.Underivatized amino acids, N-derivatized amino-acids, acidic compounds, neutrals, amides, esters and amines can be separated.The first four of the above-mentioned chiral selectors appear to have the largest enantiorecognition range.The selectors can also be derivatized to obtain different enantioselectivities. 

An extremely important aspect in pharmaceutical research is the determination of drug optical purity. The most frequently applied technique for chiral separations in CZE remains the so-called dynamic mode where resolution of enantiomers is carried out by adding a chiral selector directly into the BGE for in situ formation of diastereomeric derivatives. Various additives, such as cyclodextrins (CD), chiral crown ethers, proteins, antibiotics, bile salts, chiral micelles, and ergot alkaloids, are reported as chiral selectors in the literature, but CDs are by far the selectors most widely used in chiral CE. 

Brush-type, proteins, CDs, natural molecular imprint-based polymers (MIP), and macrocyclic antibiotics have been immobilized as chiral selectors on packed-CEC columns. Zheng and Shamsi demonstrated the possibility of using chiral CEC—ESI/MS with a commercially packed column for the determination of warfarin enantiomers in human plasma using coumachlor as an internal standard (IS). Robustness of this chiral CEC capillary was recently improved by a novel procedure and applied for the simultaneous enantiosepara-tion of height /1-blockers with multimodal CSP using different combinations of vancomycin and teicoplanin, as presented in Figure 5. ... 

HSA and BSA were used as chiral selectors to separate ofloxazin, propranolol, and verapamil enantiomers. With ketoprofen and warfarin as displacers, preferred binding sites at both proteins were discussed. 

This protein is extracted from pancreatic tissues. This protein occurs in a-form and /1-form, but the a-form is used as chiral selector in liquid chromatography. The molecular mass is 25,000, with an isoelectric point of 8.1-8.6. It is inhibited by metal ions. The protein is useful for chiral resolution of amino acids and amino esters. 

As discussed earlier, the proteins used as chiral selectors in affinity chromatography cannot be used under the high-pressure HPLC with a variety of mobile phases therefore, these proteins were immobilized with some solid support such as hydroxyethylmethacrylate, polystyrene-divinylbenzene, polyethylene fibers, and silica gel [14,15]. Avariety of techniques have been used for the immobiliza-... 

Olsson and Blomberg [141] enantioseparated omeprazole and its metabolite 5-hydroxyomeprazole using open tubular capillary electrochromatography with immobilized avidin as chiral selector. The separation was performed with open tubular capillary electrochromatography. The protein avidin was used as the chiral selector. Avidin was immobilized by a Schiffs base type of reaction where the protein was via glutral-dehyde covalently bonded to the amino-modified wall of a fused-silica capillary, 50 /an i.d. Both racemates were baseline resolved. Resolution... 

A number of enantiomers have been resolved by forming diastereomeric complexes with a chiral selector which had been added to the mobile phase. These complexes can be resolved on conventional achiral sorbents. Metal chelates, ion-pairing agents, and proteins have been used as chiral selectors. 

Proteins, which are widely used as chiral selectors and the tradenames of the corresponding commercially available CSPs and HPLC columns, are summarized in Table 9.8, together with some characteristic properties of the proteins. The protein selectors are polymeric selectors made-up of a sequence of heterogeneous monomers, which provides the basis for a huge structural diversity. Specific folding (secondary and... 

The chiral SO is coated to the capillary wall. This technique is known as open-tubular electrochromatography (OT-EC) )5(X)1. Enantioselective OT-EC methods have been described for CDs 1277), proteins ).501), polysaccharides )5021 and terguride [503] as chiral selectors. The main disadvantage is the low loading capacity, and therefore this technique has not gained high popularity. 

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