Warfarin enantiomers

Chu, Y.Q., Wainer, I.W. (1988). The measurement of warfarin enantiomers in serum using coupled achiral/chiral high performance liquid chromatography. Pharm. Res. 5, 680-683. 

In reference 22, only qualitative responses were examined in the 12-experiment PB design, i.e., the selectivity factor and the resolution of the propranolol, praziquantel, and warfarin enantiomers. In Table 8, the resolution results of the three chiral substances are given. 

For example, in reference 22, nominal experiments were added to the experimental setup at the beginning, halfway, and at the end of the experimental design. For the response resolution of warfarin enantiomers, the results of these nominal experiments were 4.11, 4.08, and 4.05, respectively. The estimated %drift is —1.46% (Equation (1)). In this example, the drift is considered non-existing and corrected responses (Equation (2)) are not calculated. Thus, for each particular response, it is evaluated whether a time effect occurs. 

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. ... 

Zheng, J., and Shamsi, S. A. (2003). Combination of chiral capillary electrochromatography with electrospray ionization mass spectrometry method development and assay of warfarin enantiomers in human plasma. Anal. Chem. 75, 6295—6305. 

Chan K, Lo AC, Yeung JH, Woo KS. The effects of danshen Salvia miltior-rhiza) on pharmacodynamics and pharmacokinetics of warfarin enantiomers in rats. J Pharm Pharmacol 1995 47 402-406. 

In 1973, Stewart and Doherty [9] resolved enantiomers of tryptophan on a column packed with BSA-succinoylaminoethyl-agarose in a discontinuous elution procedure. The mobile phase used was 0.1 M borate buffer (pH 9.2). The chromatograms of this classical research are shown in Figure 2. Several years later, this technique was applied for the chiral resolution of warfarin enantiomers [10]. In 1981, the enantiomers of tryptophan and warfarin racemates were resolved on various serum albumin CSPs [11,21,22]. The same method was used for the resolution of other drugs [12-14]. Allenmark et al. [23] studied the resolution of a series of active racemic sulfoxides on a BSA column using 0.08 M phosphate buffer (pH 5.8) as the eluting solvent. 

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). 

Henne KR, Gaedigk A, Gupta G, et al. Chiral phase analysis of warfarin enantiomers in patient plasma in relation to CYP2C9 genotype. J Chromatogr B Biomed Sci Appl 1998 710 143-148. 

Takahashi H, Kashima T, Nomizo Y, et al. Metabolism of warfarin enantiomers in Japanese patients with heart disease having different CYP2C9 and CYP2C19 genotypes. Clin Pharmacol Ther 1998 63 519-528. 

Acenocoumarol, phenprocoumon, warfarin enantiomers/plasma HPLCMS MS Column Chira-Grom-2 (250 x 1 mm, 8 pm) Mobile phase ACN MeOH CH3COOH (gradient elution) Detection MS/MS, ionization ESI Extraction on-line SPE/column Poros R2/20 (2 X 30 mm) LOD 0.5 ng/mL LOQ 2 ng/mL [68]... 

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. 

Table 3 Mean Pharmacokinetic P rameters for Warfarin Enantiomers in the Rat and Human upon i.v. Administration of R(+)- or S -)- Warfarin...

Yacobi, A., Lai, Chii-Ming, and Levy, G, (1984). Pharmacokinetic and pharmacodynamic studies of acute interaction between Warfarin enantiomers and Metronidazol. /. Pharmacol Exp, Ther., 232 72-79,... 

A. Yacobi and G. Levy, Protein binding of warfarin enantiomers in serum of humans and rats, /. Pharmacokinet. Biojdtarm., 5 123 (1977). 

I. Fitos, C. Lagercrantz, T. Larsson, M. Simonyi, I. Sjoholm, and Z. Tegyey, Stereoselective binding of 3-acetoxy-l,4-benzodiazepin-2-ones and 3-hydroxy-l,4-benzodi epin-2-ones to human serum albumin Selective allosteric interaction with warfarin enantiomers, Biodiem. Pharmacol., 35 263 (1986). 

I. Fitos and M. Simonyi, Selective effect of clonazepam and (S)-uxepam on the binding of warfarin enantiomers to human serum albumin, /. Chroma-togr., 450 217 (1988). 

Takahashi H, Ishikawa S, Nomoto S. et al. Developmental changes in pharmacokinetics and pharmacodynamics of warfarin enantiomers in Japanese 59. 

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]... 

Naidong, W Lee, J.W Development and validation of a high-performance liquid chromatographic method for the quantitation of warfarin enantiomers in human plasma. J.Pharm.Biomed.Anal., 1993, 11, 785-792... 

Yamazaki, H. and T. Shimada (1997). Human liver cytochrome P450 enzymes involved in the 7-hydroxylation of R- and 5-warfarin enantiomers. Biochem. Pharmacol. 54, 1195-1203. 

Warfarin is a racemic mixture of S- andi -warfarin enantiomers, with the S-enantiomer possessing significantly (5-6 times) more anticoagulant properties. S-warfarin is metabolized primarily by CYP2C9 and i -warfarin by CYP1A2, CYP2C19 and CYP3A4. 

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

Figure 4 Pharmacokinetic and pharmacodynamic relationship of each warfarin enantiomer when 1 mg single warfarin enantiomer was administered to steady-state in five volunteers, followed by a short infusion of vitamin Kl. It is apparent that the S enantiomer is more efficient in preventing conversion of the inactive vitamin Kl epoxide to active vitamin Kl and increasing prothrombin time. (Plot constructed from data presented in Ref 290.)...

Warfarin enantiomers are extensively metabolized by liver, possess a low hepatic extraction ratio, and are extensively bound (> 99%) to plasma proteins (Table 3). Therefore any change in the protein binding of warfarin enantiomers may alter the clearance and plasma concentrations of R- and S-warfarin [54]. Yacobi and Levy [54] studied the plasma protein binding of racemic and individual enantiomers of warfarin in human blood. The free fraction of R-warfarin was significantly (32%) larger than that of S-warfarin (Table 3). The authors concluded that the difference in the potency of warfarin enantiomers could not be solely explained by the observed differences in the protein binding of the individual enantiomers but rather by the intrinsic ability of R- and S-warfarin for interactions with extravascular receptors. 

In general, despite lower free fraetion, S-warfarin is more rapidly eliminated than its antipode in humans, and hepatic metabolism acconnts for almost all of the elimination of the drug [60]. Warfarin is metabolized to 6-hydroxywarfarin, 7-hydroxywarfarin, and two rednced derivatives, the diastereomers of warfarin alcohol [60]. The metabolism of R-warfarin involves oxidation to 6-hydroxywarfarin and subseqnent reduction to (R,S)-warfarin alcohol. In contrast, S-warfarin is oxidized to 7-hydroxywarfarin and reduced to (S,S)-warfarin alcohol. The metabolic profiles of warfarin enantiomers in blood and urine were similar [53,60]. The ring-hydroxylated metabolites are inactive, and the activity of warfarin alcohols are snb-stantially less than the parent molecule [53,60]. S-warfarin may also be metabolized to 6-hydroxywarfarin [60]. 

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. 

A number of drugs that act as substrates of CYPIA and CYP2C may interact with the metabolism of warfarin enantiomers. For example, sulphaphenazole and tolbutamide are competitive inhibitors of S-warfarin hydroxylation with Ki values of 100 and 0.5 pM, respectively [62]. 

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