RESUMO
In this study, for the first time, nanofluid of magnetic-activated charcoal and hydrophobic deep eutectic solvent (AC@Fe3 O4 -DES) based dispersive magnetic solid-phase extraction was successfully applied for the determination and preconcentration of warfarin in plasma and urine samples. The hydrophobic DES was prepared by mixing tetramethylammonium chloride (as hydrogen bond acceptor) and thymol (as hydrogen bond donor) and acted simultaneously as both carrier and stabilizer for magnetic nanoparticles. In this method, the nanofluid as a new extraction solvent was rapidly injected into the aqueous sample, which led to improvement of the mass transfer of the analytes into the sorbent and reduction of the extraction time. In the screening step, the fractional factorial design was applied for selecting some important parameters which significantly affected the extraction procedure. The effective parameters were then optimized by Box-Behnken design. Under the optimal conditions, the limits of detection were in the range of 0.3-1.6 ng/ml. A good linear range was observed in the range of 1.0-500.0 ng/ml for water and 5.0-500.0 ng/ml for urine and plasma. The intra- and inter-day relative standard deviations were 2.7-3.2 and 1.9-4.5% for five replications, respectively. Based on the results, the proposed method was successfully applied for the determination of warfarin in biological samples, using high-performance liquid chromatography.
Assuntos
Carvão Vegetal/química , Cromatografia Líquida de Alta Pressão/métodos , Nanopartículas de Magnetita/química , Extração em Fase Sólida/métodos , Varfarina , Humanos , Interações Hidrofóbicas e Hidrofílicas , Limite de Detecção , Modelos Lineares , Compostos de Amônio Quaternário/química , Reprodutibilidade dos Testes , Solventes/química , Timol/química , Varfarina/sangue , Varfarina/isolamento & purificação , Varfarina/urinaRESUMO
In the present study, a new generation of water-immiscible natural deep eutectic solvents (DESs) was synthesized using borneol as a hydrogen-bonding acceptor and decanoic acid, oleic acid, and thymol as a hydrogen-bonding donor in different molar ratios. These green hydrophobic solvents which are chemically stable in aqueous solutions were used as extraction solvents for isolation and pre-concentration of warfarin in biological samples. In this method, fine droplets of DESs were dispersed into the sample solution by using the air-assisted liquid-liquid micro-extraction method to accelerate the cloudy emulsion system formation and increase the mass transfer of the analyte to the DES-rich phase. The borneol based deep eutectic solvent is a worthy generation of the extraction solvents in the ALLME method due to low-cost and less toxicity. A Plackett-Burman design was utilized for screening the experimental parameters. The effective parameters were then optimized by Box-Behnken design (BBD). Optimized extraction conditions were pH of sample solution of 3.9, number of aspiration/dispersion cycles of 15, the volume of DES of 60 µL, and rate and time of centrifuge of 6000 rpm and 10 min, respectively. Under the optimized conditions, the developed NADES-ALLME method exhibited a wide linear range of 5-500 µg L - 1 for plasma and urine samples with satisfactory recoveries above 88.80%. Limit of detections (LODs) and Limit of quantifications (LOQs) of warfarin were in the ranges of 0.5-2.7 and 1.65-8.91, respectively. The enrichment factors were obtained in the range of 148-164 and precisions were lower than 5.87%. Finally, the proposed method was successfully employed for the analysis of warfarin in human urine and plasma samples.
Assuntos
Canfanos/química , Microextração em Fase Líquida/métodos , Solventes/química , Varfarina/análise , Varfarina/isolamento & purificação , Adulto , Ar , Ácidos Decanoicos/química , Feminino , Química Verde , Humanos , Ligação de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Limite de Detecção , Ácido Oleico/química , Solventes/síntese química , Timol/química , Varfarina/sangue , Varfarina/urina , Água/químicaRESUMO
In this study, for the first time a rapid, selective and highly sensitive method was developed for simultaneous determination of warfarin and mycophenolic acid using carbon paste electrode modified by ß-cyclodextrin/multi-walled carbon nanotubes/cobalt oxide nanoparticles (ß-CD/MWCNTs/Co3O4NPs/CPE). The oxidation peaks of desired drugs was separated enough using the constructed electrode. Cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were utilized for study the electrochemical response of the fabricated electrode and its surface modification was investigated by electrochemical impedance spectroscopy (EIS). Under optimal conditions, the adsorptive stripping voltammetric responses were linear in the concentration ranges 0.05-150⯵M and 0.5-200⯵M for WAR and MPA, respectively. The correlation coefficients were greater than 0.99. The limits of detection for WAR and MPA were 0.02 and 0.03⯵M. The fabricated electrode was applied for the simultaneous determination of WAR and MPA in urine and human serum samples with satisfactory results.
Assuntos
Técnicas Eletroquímicas , Hidroxibenzoatos , Varfarina , Adulto , Carbono/química , Eletrodos , Humanos , Hidroxibenzoatos/sangue , Hidroxibenzoatos/urina , Masculino , Varfarina/sangue , Varfarina/urinaRESUMO
In the present work, a practical method that combines excitation-emission matrix fluorescence with a second-order calibration method based on an alternating trilinear decomposition (ATLD) algorithm was developed in order to simultaneously and directly determine the contents of warfarin (WAR) and aspirin (ASA) in human plasma and urine samples, even in the presence of unknown interferences. With the pre-estimated component number of 4, the obtained average spiked recoveries were 105.4 ± 7.8 and 104.2 ± 8.3% for WAR, 96.5 ± 2.8 and 91.2 ± 2.3% for ASA in human plasma and urine samples, respectively. Furthermore, the figures of merit were calculated and also inter- and intra-day experiments were performed that proved the proposed method is of great significance to the monitoring of clinical administration and also being a simple sample pretreatment at low-cost.
Assuntos
Aspirina/sangue , Aspirina/urina , Análise Química do Sangue/métodos , Urinálise/métodos , Varfarina/sangue , Varfarina/urina , Algoritmos , Calibragem , Humanos , Espectrometria de Fluorescência , Fatores de TempoRESUMO
Multiple factors can impact warfarin therapy, including genetic variations in the drug-metabolizing enzyme cytochrome P450 2C9 (CYP2C9). Compared with individuals with the wild-type allele, CYP2C9*1, carriers of the common *3 variant have significantly impaired CYP2C9 metabolism. Genetic variations in CYP2C9, the primary enzyme governing the metabolic clearance of the more potent S-enantiomer of the racemic anticoagulant warfarin, may impact warfarin-drug interactions. To establish a baseline for such studies, plasma and urine concentrations of R- and S-warfarin and 10 warfarin metabolites were monitored for up to 360 hours following a 10-mg warfarin dose in healthy subjects with 4 different CYP2C9 genotypes: CYP2C9*1/*1 (n = 8), CYP2C9*1/*3 (n = 9), CYP2C9*2/*3 (n = 3), and CYP2C9*3/*3 (n = 4). Plasma clearance of S-warfarin, but not R-warfarin, decreased multiexponentially and in a CYP2C9 gene-dependent manner: 56%, 70%, and 75% for CYP2C9*1/*3, CYP2C9*2/*3, and CYP2C9*3/*3 genotypes, respectively, compared with CYP2C9*1/*1, resulting in pronounced differences in the S:R ratio that identified warfarin-sensitive genotypes. CYP2C9 was the primary P450 enzyme contributing to S-warfarin metabolism and a minor contributor to R-warfarin metabolism. In the presence of a defective CYP2C9 allele, switching of warfarin metabolism to other oxidative pathways and P450 enzymes for the metabolic elimination of S-warfarin was not observed. The 10-hydroxywarfarin metabolites, whose detailed pharmacokinetics are reported for the first time, exhibited a prolonged half-life with no evidence of renal excretion and displayed elimination rate-limited kinetics. Understanding the impact of CYP2C9 genetics on warfarin pharmacokinetics lays the foundation for future genotype-dependent warfarin-drug interaction studies.
Assuntos
Anticoagulantes/química , Anticoagulantes/farmacocinética , Citocromo P-450 CYP2C9/genética , Varfarina/química , Varfarina/farmacocinética , Adolescente , Adulto , Anticoagulantes/sangue , Anticoagulantes/urina , Área Sob a Curva , Feminino , Genótipo , Meia-Vida , Humanos , Masculino , Taxa de Depuração Metabólica , Pessoa de Meia-Idade , Polimorfismo Genético , Varfarina/sangue , Varfarina/urina , Adulto JovemRESUMO
Roof rats (Rattus rattus) live mainly in human habitats. Heavy use of rodenticides, such as warfarin, has led to the development of drug resistance, making pest control difficult. There have been many reports regarding mutations of vitamin K epoxide reductase (VKOR), the target enzyme of warfarin, in resistant rats. However, it has been suggested there are other mechanisms of warfarin resistance. To confirm these possibilities, closed colonies of warfarin-susceptible roof rats (S) and resistant rats from Tokyo (R) were established, and the pharmacokinetics/pharmacodynamics of warfarin in rats from both colonies was investigated. R rats had low levels of warfarin in serum and high clearance activity. These rats can rapidly metabolize warfarin by hydroxylation. The levels of accumulation in the organs were lower than those of S rats. R rats administered warfarin showed high expression levels of CYP2B, 2C, and 3A, which play roles in warfarin hydroxylation, and may explain the high clearance ability of R rats. The mechanism of warfarin resistance in roof rats from Tokyo involved not only mutation of VKOR but also high clearance ability due to high levels of CYP2B, 2C and 3A expression possibly induced by warfarin.
Assuntos
Resistência a Medicamentos/fisiologia , Rodenticidas/farmacologia , Rodenticidas/farmacocinética , Varfarina/farmacologia , Varfarina/farmacocinética , Animais , Sistema Enzimático do Citocromo P-450/metabolismo , Resistência a Medicamentos/genética , Fezes/química , Hidroxilação , Rim/metabolismo , Fígado/metabolismo , Masculino , Microssomos Hepáticos/metabolismo , Mutação , Tempo de Protrombina , Ratos , Rodenticidas/sangue , Rodenticidas/urina , Vitamina K Epóxido Redutases/genética , Varfarina/sangue , Varfarina/urinaRESUMO
A simple and rapid liquid chromatographic-tandem mass spectrometric method has been developed and validated for the enantiospecific determination of R- and S-warfarin in human urine. Warfarin enantiomers were extracted from urine using methyl tert-butyl ether. Chromatographic separation of warfarin enantiomers and the internal standard d5-warfarin was achieved using a Astec Chirobiotic V column with gradient mobile phase at a flow rate of 400 µL/min over 10 min. Detection was performed on a TSQ Quantum Ultra triple quadrupole mass spectrometer equipped with a heated electrospray ionization source. Analytes were detected in negative ionization mode using selected reaction monitoring. Calibration curves were linear with a correlation coefficient of ≥0.996 for both enantiomers over a concentration range of 5-500 ng/mL. The intra- and interday accuracy and precision for both analytes were within ±9.0%. Excellent extraction efficiency and negligible matrix effects were observed. The applicability of the method was demonstrated by successful measurement of warfarin enantiomers in urine of patients with kidney disease. The method is simple, accurate and reproducible and is currently being used to support warfarin pharmacokinetic studies.
Assuntos
Anticoagulantes/urina , Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas em Tandem/métodos , Varfarina/urina , Calibragem , Humanos , Reprodutibilidade dos Testes , Espectrometria de Massas por Ionização por Electrospray , Estereoisomerismo , Varfarina/químicaRESUMO
The aim of this work was to develop a method for the clean-up and preconcentration of warfarin from biological sample employing a new molecularly imprinted polymer (MIP) as a selective adsorbent for solid-phase extraction (SPE). This MIP was synthesized using warfarin as a template, pyrrole as a functional monomer and vinyl triethoxysilane as a cross-linker. The molar ratio of 1:4:20 (template-functional monomer-cross-linker) showed the best results. Nonimprinted polymers (NIPs) were prepared and treated with the same method, but in the absence of warfarin. The prepared polymer was characterized by Fourier transmission infrared spectrometry and scanning electron microscopy. An adsorption process (SPE) for the removal of warfarin using the fabricated MIPs and NIPs was evaluated under various conditions. Effective parameters on warfarin extraction, for example, type and volume of elution solvent, pH of sample solution, breakthrough volume and maximum loading capacity, were studied. The limits of detection were in the range of 0.0035-0.0050 µg mL(-1). Linearity of the method was determined in the range of 0.0165-10.0000 µg mL(-1) for plasma and 0.0115-10.0000 µg mL(-1) for urine with coefficients of determination (R(2)) ranging from 0.9975 to 0.9985. The recoveries for plasma and urine samples were >95%.
Assuntos
Anticoagulantes/sangue , Anticoagulantes/urina , Impressão Molecular , Polímeros/química , Pirróis/química , Varfarina/sangue , Varfarina/urina , Adsorção , Cromatografia Líquida de Alta Pressão , Humanos , Limite de Detecção , Microscopia Eletrônica de Varredura , Extração em Fase SólidaRESUMO
OBJECTIVE: This study investigated clinical factors associated with negative urinary antigen tests (UAT) implemented for the diagnosis of pneumococcal community-acquired pneumonia (CAP) in adult patients. SUBJECTS AND METHODS: We reviewed the medical records of 755 adult patients who completed the UAT in our hospital between 2009 and 2012. Of these, we evaluated 63 patients with bacteriologically confirmed definite pneumococcal CAP (33 were UAT-positive, and 30 were UAT-negative). RESULTS: There was no significant difference between the UAT-positive and the UAT-negative patients regarding age, dehydration, respiratory failure, orientation, blood pressure (ADROP) score (the CAP severity score proposed by the Japanese Respiratory Society), gender, white blood cell counts, liver/kidney function tests, or urinalysis. However, serum C-reactive protein (CRP) concentrations were 31% lower in the UAT-negative patients than in the UAT-positive patients (p = 0.02). Furthermore, the prothrombin time-international normalized ratio was 50% higher in the UAT-negative patients than in the UAT-positive patients, although the difference did not reach statistical significance (p = 0.06). The prevalence of comorbidities was similar in both UAT-positive and UAT-negative patients. However, warfarin had been prescribed in 8 (27%) of the UAT-negative patients compared to only 1 (3%) of the UAT-positive patients (odds ratio = 11.6; p = 0.01). CONCLUSIONS: These results suggested that low serum CRP concentrations and the use of warfarin increased the possibility with which false-negative UAT results occurred in these patients with pneumococcal CAP.
Assuntos
Antígenos de Bactérias/urina , Pneumonia Pneumocócica/urina , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticoagulantes/urina , Proteína C-Reativa/análise , Infecções Comunitárias Adquiridas , Comorbidade , Feminino , Hospitais de Ensino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sensibilidade e Especificidade , Streptococcus pneumoniae , Tóquio , Varfarina/urinaRESUMO
In the present work, a simple and sensitive electrochemical sensor based on magnetic Fe3O4 nanoparticles modified carbon paste electrode (CPE) was introduced for detection of low level with 0.21µM of warfarin. Under the optimum experimental conditions the oxidation peak current of warfarin was used for its monitoring for the first time. The analytical curve was linear for warfarin concentrations from 0.5 to 1000µM with a limit of detection of 0.21µM by square wave anodic stripping voltammetry (SWASV). The proposed sensor showed excellent stability and was used for the determination of warfarin in tablet, human serum and urine with satisfactory results.
Assuntos
Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Compostos Férricos/química , Nanopartículas/química , Varfarina/análise , Carbono/química , Espectroscopia Dielétrica , Técnicas Eletroquímicas/economia , Eletrodos , Humanos , Concentração de Íons de Hidrogênio , Limite de Detecção , Microscopia Eletrônica de Varredura , Oxirredução , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Varfarina/sangue , Varfarina/química , Varfarina/urinaRESUMO
Here, we report on the use of human serum albumin (HSA)-modified Fe(3)O(4) nanoparticles (NPs) (HSA-Fe(3)O(4) NPs) for affinity-SALDI-MS of small drugs in human biological liquids. We demonstrated that HSA-Fe(3)O(4) NPs effectively captured small drugs from human urine and serum via the interactions between HSA and these drugs. The drugs adsorbed on HSA could then be identified by directly introducing the HSA-Fe(3)O(4) NPs into a mass spectrometer for SALDI-MS analysis. The ability of HSA to interact with multiple small drugs facilitated the simultaneous detection of a 4-drug-mixture in serum, viz., phenytoin, ibuprofen, camptothecin, and warfarin sodium, by affinity-SALDI-MS using HSA-Fe(3)O(4) NPs. In contrast, matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) with an organic matrix could detect only warfarin sodium. We also demonstrated the capacity of affinity-SALDI-MS to quantify warfarin sodium in urine samples across a range of 50 - 1000 µM (R(2) = 0.998) when using HSA-Fe(3)O(4) NPs. The detection sensitivity was further improved to a range of 5 - 100 µM (R(2) = 0.999) by using denatured HSA. The open structure of denatured HSA may enhance the effective extraction of small drugs from biological liquids, and increase the detection-sensitivity of affinity-SALDI-MS. Affinity-SALDI-MS using protein-modified Fe(3)O(4) NPs can open up new approaches to the analytical detection of small drugs in biological liquids by SALDI-MS.
Assuntos
Camptotecina/sangue , Ibuprofeno/sangue , Nanopartículas de Magnetita/química , Fenitoína/sangue , Albumina Sérica/química , Varfarina/sangue , Varfarina/urina , Óxido Ferroso-Férrico/química , Humanos , Peso Molecular , Sensibilidade e Especificidade , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Concurrent sample clean-up and enhancement in detection sensitivity for chiral capillary electrophoresis was demonstrated based on the coupling of salting-out extraction with acetonitrile stacking and the use of dimethyl-beta-cyclodextrin as the chiral selector for the sensitive and enantioselective separation of warfarin enantiomers in urine samples. By optimizing the pH of salting-out extraction, warfarin enantiomers can be efficiently extracted from the aqueous sample solution into a smaller volume organic solvent (acetonitrile) phase. The pressure injection of the enriched acetonitrile phase (containing ca. 1% NaCl) into the CE capillary at 10% capillary volume resulted in additional concentration of the warfarin enantiomers. The limit of detection for both warfarin enantiomers was as low as 1.5 ng/mL in urine sample. Our results show that the novel strategy offers improved sensitivity compared to conventional CE analysis, reaching a combined enrichment factor higher than 1000. Calibration curves of warfarin enantiomers in urine samples were found to be linear between 10 and 1000 ng/mL, and intra- and inter-day precision (N=9) for both warfarin enantiomers in terms of migration time and peak area were found to be within the range of 0.1-0.8% and 1.0-6.7%, respectively. The recovery of warfarin enantiomers from urine was ca. 90%.
Assuntos
Acetonitrilas/química , Fracionamento Químico/métodos , Eletroforese Capilar/métodos , Varfarina/urina , Feminino , Humanos , Proteínas/isolamento & purificação , Sensibilidade e Especificidade , Estereoisomerismo , Varfarina/químicaRESUMO
As a step toward exploring a targeted metabolomics approach to personalized warfarin (Coumadin) therapy, we developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method capable of quantifying specific enantiomeric (R and S) contributions of warfarin (WAR) and the corresponding hydroxywarfarins (OH-WAR) and glucuronides (-GLUC) generated by cytochrome P450s (CYP) and UDP-glucuronosyltransferases (UGTs), respectively. Evaluation of quality control samples and three commercially available human samples showed that our analytical approach has the ability to measure 24 unique WAR metabolites in human urine. Evaluation of the human data also provides new insights for evaluating WAR toxicity and begins characterizing important UGT metabolic pathways responsible for WAR detoxification. Data revealed the significance of specific metabolites among patients and the corresponding enzymatic capacity to generate these compounds, including the first report of direct WAR glucuronidation. On the basis of total OH-WAR levels, (S)-7-OH-WAR was the predominant metabolite indicating the significance of CYP2C9 in WAR metabolism, although other CYP2C enzymes also contributed to clearance of this isomer. (R)-WAR hydroxylation to OH-WARs was more diverse among the patients as reflected in varying contributions of CYP1A2 and multiple CYP2C enzymes. There was wide variation in the glucuronidation of WAR and the OH-WARs with respect to the compounds and patients. 6- and 7-OH-WAR were primarily (>70%) excreted as glucuronides unlike 4'-OH-WAR and 8-OH-WAR. For all patients, UGT1A1 is likely responsible for 6-O-GLUC production, although UGT1A10 may also contribute in one patient. 7-O-GLUC levels reflected contributions from potentially five different UGT1A enzymes. In all cases, WAR, 4'-OH-WAR, 8-OH-WAR, and the corresponding glucuronides were minor metabolites with respect to the others. Taken together, these data suggest that both P450 and UGT reactions contribute to the generation of excretable products in human urine, thereby generating complex metabolic networks.
Assuntos
Anticoagulantes/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Glucuronosiltransferase/metabolismo , Varfarina/metabolismo , Anticoagulantes/toxicidade , Anticoagulantes/urina , Cromatografia Líquida de Alta Pressão , Humanos , Metabolômica , Estereoisomerismo , Espectrometria de Massas em Tandem , Varfarina/toxicidade , Varfarina/urinaRESUMO
We report a patient who attempted suicide by taking warfarin as a coumarin rodenticide twice, and showed oral bleeding and an abnormally prolonged bleeding tendency after 10 days. He was conscious. His body temperature was 36.6 degrees C, the respiratory rate was 23/min, and blood pressure was 142/87 mmHg. A bite wound showing continuous bleeding was observed on the right side of the oral cavity. CT of the head and upper gastrointestinal endoscopy showed no intracranial and gastrointestinal bleeding,respectively. The PT and INR were extremely high, and complete blood count revealed anemia. Blood chemistry showed hypoproteinemia. Occult blood was strongly positive. Routine urine examination and electrocardiography showed neither abnormal nor characteristic findings. In drug/ poison screening tests, GC/MS analysis revealed warfarin artifacts in serum and urine samples. Warfarin was isolated from each sample, converted to a TMS derivative, and quantified by SIM analysis of GC/MS. During the clinical course, KaytwoN (vitamin K) was intravenously administered at a dose of 20 mg since the INR at the first consultation was 20.89. After admission to the ICU, 20 mg Kaywan (vitamin K) was orally administered, and the INR after 5 hours improved to 2.32. Kaywan administration (20 mg/day) was initiated, and he improved and was discharged 5 days after admission. The definite cause could be clarified by drug/poison analysis, and effective treatment could be performed.
Assuntos
Cumarínicos , Intoxicação/etiologia , Rodenticidas , Tentativa de Suicídio , Varfarina/intoxicação , Antifibrinolíticos/administração & dosagem , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Masculino , Pessoa de Meia-Idade , Intoxicação/diagnóstico , Intoxicação/tratamento farmacológico , Resultado do Tratamento , Vitamina K/administração & dosagem , Varfarina/sangue , Varfarina/urinaRESUMO
A high-performance liquid chromatography-mass spectrometry (HPLC-MS) method for the quantification of phenprocoumon, warfarin, and their known monohydroxylated metabolites in human plasma and urine was developed using a simple, selective solid-phase extraction scheme. Chromatographic separation was achieved on a reversed-phase Luna C18 column and step gradient elution resulted in a total run time of about 13 min. Limits of quantification (LOQ) were < or = 40 nM for the parent compounds and < or = 25 nM for the metabolites and the limit of detection (LOD) was < or = 2.5 nM for all analytes. Average recovery was 84% (+/- 3.7) and 74% (+/- 13.2) in plasma and urine, respectively. Intra- and inter-day coefficients of variation were < or = 8.6 and < or = 10.6% in plasma and urine, respectively. The method was successfully applied to the analysis of phenprocoumon samples from four healthy volunteers and should prove useful for future comparative studies of warfarin and phenprocoumon pharmacokinetics.
Assuntos
Anticoagulantes/farmacocinética , Cromatografia Líquida de Alta Pressão/métodos , Femprocumona/farmacocinética , Varfarina/farmacocinética , Anticoagulantes/sangue , Anticoagulantes/urina , Humanos , Femprocumona/sangue , Femprocumona/urina , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Varfarina/sangue , Varfarina/urinaRESUMO
UNLABELLED: M: The aim of this study was to investigate the effect of St John's wort and ginseng on the pharmacokinetics and pharmacodynamics of warfarin. METHODS: This was an open-label, three-way crossover randomized study in 12 healthy male subjects, who received a single 25-mg dose of warfarin alone or after 14 days' pretreatment with St John's wort, or 7 days' pretreatment with ginseng. Dosing with St John's wort or ginseng was continued for 7 days after administration of the warfarin dose. Platelet aggregation, international normalized ratio (INR) of prothrombin time, warfarin enantiomer protein binding, warfarin enantiomer concentrations in plasma and S-7-hydroxywarfarin concentration in urine were measured. Statistical comparisons were made using anova and 90% confidence intervals are reported. RESULTS: INR and platelet aggregation were not affected by treatment with St John's wort or ginseng. The apparent clearances of S-warfarin after warfarin alone or with St John's wort or ginseng were, respectively, 198 +/- 38 ml h(-1), 270 +/- 44 ml h(-1) and 220 +/- 29 ml h(-1). The respective apparent clearances of R-warfarin were 110 +/- 25 ml h(-1), 142 +/- 29 ml h(-1) and 119 +/- 20 ml h(-1) [corrected]. The mean ratio and 90% confidence interval (CI) of apparent clearance for S-warfarin was 1.29 (1.16, 1.46) and for R-warfarin it was 1.23 (1.11, 1.37) when St John's wort was coadministered. The mean ratio and 90% CI of AUC(0-168) of INR was 0.79 (0.70, 0.95) when St John's wort was coadministered. St John's wort and ginseng did not affect the apparent volumes of distribution or protein binding of warfarin enantiomers. CONCLUSIONS: St John's wort significantly induced the apparent clearance of both S-warfarin and R-warfarin, which in turn resulted in a significant reduction in the pharmacological effect of rac-warfarin. Coadministration of warfarin with ginseng did not affect the pharmacokinetics or pharmacodynamics of either S-warfarin or R-warfarin.
Assuntos
Anticoagulantes/farmacocinética , Hypericum , Panax , Extratos Vegetais/farmacologia , Varfarina/análogos & derivados , Varfarina/farmacocinética , Análise de Variância , Anticoagulantes/sangue , Anticoagulantes/farmacologia , Área Sob a Curva , Estudos Cross-Over , Interações Ervas-Drogas , Humanos , Coeficiente Internacional Normatizado , Masculino , Agregação Plaquetária/efeitos dos fármacos , Tempo de Protrombina , Varfarina/sangue , Varfarina/farmacologia , Varfarina/urinaRESUMO
The effect of the proton pump inhibitor omeprazole on the anticoagulation and the pharmacokinetics of warfarin enantiomers was studied in rats. Omeprazole given intraperitoneally in a daily dose of 0.67 mg/kg over 8 days had no significant effect on the absorption, distribution and the total serum clearance values of the S- and R-enantiomers of warfarin. Omeprazole did not affect the pre-treatment baseline blood coagulation and the in vitro rat serum protein binding of warfarin enantiomers. In vitro study with rat liver microsomes showed that omeprazole had an inhibitory effect on the hydroxylation of warfarin enantiomers. Results obtained from in vivo urinary excretion study revealed that omeprazole inhibited the formation clearance of both S- and R-form oxidative metabolites, but increased that of the overall reductive metabolites, and the renal clearance of S- and R-enantiomers, of warfarin. As a consequence, the total serum clearance values for warfarin enantiomers remained unchanged and the hypoprothrombinaemic response produced by warfarin was not affected.
Assuntos
Antiulcerosos/farmacologia , Anticoagulantes/farmacologia , Anticoagulantes/farmacocinética , Omeprazol/farmacologia , Varfarina/farmacologia , Varfarina/farmacocinética , Algoritmos , Animais , Anticoagulantes/urina , Biotransformação , Interações Medicamentosas , Hidroxilação , Técnicas In Vitro , Indicadores e Reagentes , Masculino , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Oxirredução , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Estereoisomerismo , Varfarina/urinaRESUMO
OBJECTIVE: Our objective was to investigate population differences in the metabolic activity of cytochrome P450 (CYP) 2C9 between genotypically matched Caucasian and Japanese patients by using the unbound oral clearance of S-warfarin as an in vivo phenotypic trait measure. METHODS: Ninety Japanese and 47 Caucasian patients receiving maintenance warfarin therapy were studied. Steady-state plasma unbound concentrations of S-warfarin were measured by a chiral HPLC method coupled with an ultrafiltration technique, and unbound oral clearance for S-warfarin was estimated. By combining plasma unbound concentrations of S-warfarin with the urinary excretion rates of S-7-hydroxywarfarin, the formation clearance of S-7-hydroxywarfarin was also determined. Genotyping of CYP2C9 was performed for 6 distinct alleles (CYP2C9*1, CYP2C9*2, CYP2C9*3, CYP2C9*4, CYP2C9*5, and a T/C transition in intron 2). RESULTS: The frequency distribution of unbound oral clearance for S-warfarin obtained from Japanese patients was shifted toward higher values as compared with that in Caucasian patients. Japanese patients had lower allelic frequencies for the 5 variants than Caucasian patients. When interpopulation comparisons of CYP2C9 activity were made for genotype-matched subjects, Japanese patients with the homozygous CYP2C9*1 (wild-type) genotype (n = 85) had significantly (P <.01) greater median values for unbound oral clearance and formation clearance than Caucasian patients with the corresponding genotype (n = 26), 10.4 mL x min(-1) x kg(-1) versus 4.25 mL x min(-1) x kg(-1) and 0.015 mL x min(-1) x kg(-1) versus 0.010 mL x min(-1) x kg(-1), respectively. In addition, Japanese patients heterozygous for the CYP2C9*3 genotype (n = 4) showed a significantly (P <.05) reduced unbound oral clearance for S-warfarin, by 63%, as compared with Japanese patients possessing the homozygous CYP2C9*1 genotype. By contrast, in Caucasian patients, no significant differences were observed in this parameter between CYP2C9(*)1 homozygous subjects and those with heterozygous CYP2C9(*)2 or CYP2C9(*)3 genotypes. CONCLUSIONS: These findings indicate that population differences in the frequencies of known variant CYP2C9 alleles account only in part for the variability observed in in vivo CYP2C9 activity in different populations. In addition, a gene-dose effect of defective CYP2C9 alleles on the in vivo CYP2C9 activity is evident in Japanese patients but not in Caucasian patients. Further studies are required to identify currently unknown factor(s) (eg, transcriptional regulation) responsible for the large intrapopulation and interpopulation variability in CYP2C9 activity.
Assuntos
Anticoagulantes/farmacocinética , Hidrocarboneto de Aril Hidroxilases/genética , Hidrocarboneto de Aril Hidroxilases/metabolismo , Povo Asiático/genética , Varfarina/farmacocinética , População Branca/genética , Administração Oral , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticoagulantes/administração & dosagem , Anticoagulantes/sangue , Anticoagulantes/química , Anticoagulantes/urina , Cromatografia Líquida de Alta Pressão , Citocromo P-450 CYP2C9 , Feminino , Genótipo , Humanos , Japão , Masculino , Taxa de Depuração Metabólica , Pessoa de Meia-Idade , Polimorfismo Genético , Varfarina/administração & dosagem , Varfarina/sangue , Varfarina/química , Varfarina/urinaRESUMO
OBJECTIVE: To clarify the mechanism(s) for the interaction between warfarin and benzbromarone, a uricosuric agent, and to predict changes in the in vivo pharmacokinetics of (S)-warfarin from in vitro data. METHODS: Warfarin enantiomers and benzbromarone in serum, 7-hydroxywarfarin in urine, and serum unbound fractions of warfarin enantiomers were measured in patients with heart disease given warfarin with (n = 13) or without (n = 18) oral benzbromarone (50 mg/d). In vitro inhibition constants (K(i)) of benzbromarone for (S)-warfarin 7-hydroxylation were determined with use of human CYP2C9 and liver microsomes. The magnitude of changes in the formation clearance for 7-hydroxylation (CLf), the unbound oral clearance (CL(oral,u)), and the oral clearance (CL(oral)) for (S)-warfarin were predicted by equations incorporating the in vitro Ki, the theoretical maximum unbound hepatic benzbromarone concentration, and the fractions of warfarin eliminated through metabolism and of CYP2C9-mediated metabolic reaction susceptible to inhibition by benzbromarone. RESULTS: The patients given warfarin with benzbromarone required a 36% less (P < .01) warfarin dose than those given warfarin alone (2.5 versus 3.9 mg/d) to attain similar international normalized ratios (2.1 and 2.2, respectively), and the former had 65%, 53%, and 54% lower (P < .05 or P < .01) CLf, CL(oral),u, and CL(oral) for (S)-warfarin than the latter, respectively. In contrast, no significant differences were observed for (R)-warfarin kinetics between the groups. Benzbromarone was found to be a potent competitive inhibitor (Ki < 0.01 micromol/L) for (S)-warfarin 7-hydroxylation mediated by CYP2C9. The average changes in the in vivo CLf, CL(oral),u, and CL(oral)values for (S)-warfarin induced by benzbromarone were largely predictable by the proposed equations. CONCLUSION: Benzbromarone would intensify anticoagulant response of warfarin through an enantioselective inhibition of CYP2C9-mediated metabolism of pharmacologically more potent (S)-warfarin. The magnitude of changes in the in vivo warfarin kinetics may be predicted by in vitro data.
Assuntos
Anticoagulantes/farmacocinética , Hidrocarboneto de Aril Hidroxilases , Benzobromarona/farmacocinética , Esteroide 16-alfa-Hidroxilase , Uricosúricos/farmacocinética , Varfarina/farmacocinética , Idoso , Anticoagulantes/sangue , Anticoagulantes/urina , Benzobromarona/sangue , Benzobromarona/urina , Citocromo P-450 CYP2C9 , Sistema Enzimático do Citocromo P-450/metabolismo , Sinergismo Farmacológico , Feminino , Humanos , Masculino , Microssomos Hepáticos/enzimologia , Pessoa de Meia-Idade , Estereoisomerismo , Esteroide Hidroxilases/metabolismo , Uricosúricos/sangue , Uricosúricos/urina , Varfarina/sangue , Varfarina/urinaRESUMO
Enantiomers of warfarin and 7-hydroxywarfarin in human plasma and urine, respectively, were determined by high-performance liquid chromatography using a cellulose-derivative column with UV or fluorescent detection, and their absolute configuration was determined simultaneously by a circular dichroism spectropolarimeter connected in series. Enantiomers of warfarin and its major metabolites [i.e., (R)-6-hydroxywarfarin, (S)-7-hydroxywarfarin and (RS)-warfarin alcohol] were well resolved. The method was precise and sensitive: within- and between-day coefficients of variation were <9.6% for warfarin enantiomers in plasma and <7.1% for 7-hydroxywarfarin enantiomers in urine, respectively, and the lower detection limits were 20 ng/ml for (R)-warfarin, 40 ng/ml for (S)-warfarin, 2.5 ng/ml for (R)-7-hydroxywarfarin and 4.5 ng/ml for (S)-7-hydroxywarfarin in 0.5 ml of both plasma and urine. The ultrafiltration technique was used for determining unbound concentrations of warfarin enantiomers in plasma using [14C]warfarin enantiomers resolved by the present HPLC system. Clinical applicability of the method was evaluated by determining unbound concentrations of warfarin enantiomers in five consecutive plasma samples obtained from a patient exhibiting an unstable anticoagulant response to warfarin (4 mg/day, p.o.). Results indicated that the present method would be useful in clarifying factors responsible for a large intra- and inter-patient variability in warfarin effects with regard to unbound plasma enantiomer pharmacokinetics.