Link (1) suggested in 1948 that the coumarin compound warfarin be used as a rodenticide. Its first human use was in a suicide attempt in 1952 (2). Although warfarin has been employed increasingly in the treatment of thromboembolic disorders (3)(4)(5), no published reports of its physiologic disposition in man have appeared, probably owing largely to the lack of a suitable method for its measurement in biologic fluids (6).Weiner, Brodie, and Burns (7,8) stated that the half-life of warfarin in man is 90 hours, but did not include the assay method, or give supporting data. In 1953, Yuyama, Goto, and Umezu (9) described a colorimetric method for the estimation of warfarin in rat plasma, but apparently this method was never applied to man. Since 1954, several investigations of warfarin in the rat have appeared, including the studies of Garner (10), Eble (11), and Lin (12). Their methods, however, were unpublished except as doctoral theses, or were not readily adaptable for studies in man.Recently, we described a spectrophotometric method for the estimation of warfarin in biologic fluids (13), and have used this method in the present study to investigate the pharmiacodynamics of warfarin in man. We determined the concentrations of warfarin in the plasma and of a warfarin metabolite in the urine of normal subjects after both oral and intravenous administration of the drug. Analysis of the data provided information on its absorption, elimination, apparent volume of distribution, and excretion. Differences in its biologic effect were evaluated by simultaneous measurements of its plasma concentration and of prothrombin complex activity. * Work supported by U. S. Public Health Service grant H-2754. METHODSThe subjects, all volunteers, were 14 normal men and women, ages 27 to 63 years, and one patient, age 72 years, with coronary artery disease who was included because he was unusually resistant to the prothrombinopenic effects of the drug.Warfarin sodium 1 was administered either orally in the form of tablets (starch base) or by iv injection. The tablets were swallowed whole in the morning by subjects in the postabsorptive state, and no food was taken for at least 2 hours thereafter. For iv administration, lyophilized warfarin sodium was reconstituted in distilled water. The total dose was injected into the antecubital vein in less than 1 minute. The dose of warfarin given by either route was based on body weight in an attempt to equalize the effect of the drug on subjects with different plasma volumes, assumed to be a function of body weight. A standard dose of 1.5 mg per kg of body weight was selected so that drug levels and prothrombin complex responses would be in a clearly measurable range for several days after administration of warfarin.Test specimens were prepared as follows. Blood obtaimed by clean venipuncture was mixed in glass tubes in a proportion of 9: 1 with 3.2% sodium citrate in 0.7%c saline and centrifuged at 2,000 rpm for 20 minutes at 4°C.

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SummaryA spectrophotometric method is described for the determination of warfarin in biologic fluids. The warfarin is extracted from the specimen with acidified ethylene dichloride, washed with mild alkali and recovered from the organic phase with strong alkali. The extinction is then measured spectrophotometri-cally at the ultraviolet absorption maximum for warfarin. Results of UV absorption spectrophotometry and countercurrent distribution analysis showed that the assay is specific for unchanged warfarin in plasma; in addition, it measures a probable metabolite of warfarin in urine. Further studies indicated that the metabolite may be hydroxy-warfarin.The method was applied in determining the absorption and elimination of warfarin after oral and intravenous administration. Gastrointestinal absorption of the drug was rapid and complete. With both routes the plasma concentrations and prothrombinopenic response were similar, and the rate of disappearance from the plasma followed an exponential decay curve. A correlation was found between rate of elimination and biologic response — the slower the rate of elimination, the more pronounced the depression of prothrombin complex activity. The application of this dual approach, the simultaneous study of the metabolic fate of the drug and the prothrombinopenic response, should provide a more rational basis for the therapeutic use of anticoagulant drugs.

SummaryThe pharmacodynamic and prothrombinopénie effects of the coumarin anticoagulant drugs warfarin and dicumarol were compared in man by means of specific chemical assays for the drugs and the Quick prothrombin time for the response. The absorption of warfarin was rapid and complete, and the prothrombinopénie response was similar after administration by any route. Absorption of dicumarol as a solution or powder was rapid while as whole tablets it was often slow. Dicumarol in solution taken orally produced higher maximum concentrations of drug in plasma, more rapid absorption rates, less unchanged drug in stool and an earlier and greater prothrombin complex response than the tablets. Only a slight effect was produced by varying the formulation of warfarin. In all subjects 15 to 30% of the oral dose of dicumarol was recovered as unchanged drug in the stool. Unchanged warfarin was not recovered from the stool in any experiment.The apparent volume of distribution (Vd) for warfarin and high doses of dicumarol was about 12.5% of body weight, approximately the size of the albumin space. The long half-time of disappearance (t½) of both drugs, their prolonged duration of action, small volume of distribution, and lack of excretion of unchanged drug in the urine are probably related to their high degree of binding to plasma proteins. The t½ and Vd for warfarin are independent of dose size; the unusual dependency of these parameters on dose size with dicumarol suggests tissue uptake, perhaps by the reticuloendothelial system.A good correlation was found between the t½ of warfarin and dicumarol and the magnitude and duration of response. Up to a certain point increasing the dose of these drugs hastened the occurrence of hypoprothrombinemia ; beyond a dose size characteristic for each individual no further hastening could be obtained.A substance extracted from the stool of subjects receiving dicumarol intravenously and orally is probably a metabolite of the drug.