Warfarin enantiomers determination

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. ... 

J. P. Gramond and F. Guyon, Separation and determination of warfarin enantiomers in human plasma samples by capillary zone electrophoresis using a methylated /3-cyclodextrin-containing electrolyte, J. Chromatogr., 615 36 (1993). 

The measurement of warfarin enantiomers in serum using coupled achiral/chiral high-performance liquid chromatography" (110), An assay for the serum concentrations of (fi)-warfarin and (S)-warfarin was developed using the BSA CSF coupled to a Pinkerton internal-surface reverse-phase (ISRP) achiral column. The ISRP column was used to separate (R,S)-warfarin from the serum components and warfarin metabolites and to quantitate the total warfarin concentration. The eluent containing the (A,S)-warfarin was then selectively transferred to the BSA CSR where the enantiomers were enantioselectively resolved (a = 1.19) and the enantiomeric composition determined. 

Cai, W.M. Hatton, J. Pettigrew, L.C. Dempsey, R.J. Chandler, M.H.H. A simplified high-performance liquid chromatographic method for direct determination of warfarin enantiomers and their protein binding in stroke patients. Ther.DrugMon.it., 1994,16, 509-512 [chiral fluorescence detection phen-procoumon (IS) LOD 8 ng/mL post-column reaction detection derivatization]... 

Takahashi H, Kashima T, Kimura S, Muramoto N, Nakahata H, Kubo S, Shimoyama Y, Kajiwara M, Echizen H.Determination of unbound warfarin enantiomers in... 

Interestingly, microsomal 7-hydroxylation of racemic warfarin appears to be lower than that of S-warfarin, indicating the possibility of a metabolic interaction between warfarin enantiomers [61]. These in vitro studies showed that R-warfarin affected the catalytic activity of CYP2C9 by a noncompetitive mechanism [61]. However, in an earlier in vivo study a lack of enantiomeric interaction between R- and S-warfarin was suggested after single 1.5mg/kg doses of the individual enantiomers and racemic warfarin [62]. The enantiospecific results obtained were successfully used to predict the pharmacokinetics and pharmacodynamics of racemic warfarin. The apparently contradictory findings between the two studies may be due to a number of experimental factors. One study used a supraclinical dose of warfarin in vivo, whereas the other involved human microsomes in vitro, the relevant hepatic concentrations of which are difficult to determine. 

Boppana, V.K. Schaefer, W.H. Cyronak, M.J. High-performance liquid chromatographic determination of warfarin enantiomers in plasma with automated on-line sample enrichment. J. Biomed. Biophys. Meth. 2002,54, 315-326. 

Approximately 10% of patients require <1.5 mg/day of warfarin to achieve an INR of 2—3. These patients are more likely to possess one or two polymorphic alleles of CYP2C9, the major enzyme responsible for converting the S-enantiomer warfarin to its inactive metabolites. In comparison with the wild-type CYP2C9 1 allele, the variant alleles CYP2C9 2 and CYP2C9 3 have been shown to inactivate S-warfarin much less efficiently in vitro. The variant alleles are present in 10-20% of Caucasians, but in <5% of African Americans or Asians. Polymorphic variations in VKORCl, which encodes a component of the Vitamin K reductase complex (see Figure 54-6), also determine warfarin sensitivity. 

Whole body level parameters are determined by multiple organ parameters, which in turn are a reflection of multiple macromolecular parameters. Thus, in the case of a pair of enantiomers, the observed stereoselectivity for a particular parameter is dependent on the level of organization that the parameter represents and may be either amplified or attenuated [94]. For example, in the case of warfarin, the enantiomeric ratio (R/S 1.93) in half-life is determined primarily by differences in volume of distribution (R/S 1.83) rather than the modest difference in elearanee (R/S 1.05). In contrast, examination of the alternative metabolic routes generates marked differences in stereoselectivity, e.g., aromatic oxidation at the 6-, 7-, and 8-positions of the coumarin ring yields enantiomeric ratios of R/S 1.19, S/R 6.33, and R/S 5.2, respectively. These, together with the... 

Hermans, J.J. Thijssen, H.H. Human liver microsomal metabolism of the enantiomers of warfarin and acenocoumarol P450 isozyme diversity determines the differences in their pharmacokinetics. Br. J. Pharmacol. 1993, 110, 482-490. 

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