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Evaluation of anticoagulant effects of direct thrombin inhibitors, dabigatran and argatroban, based on the Lineweaver–Burk plot applied to the Clauss assay
  1. Yuta Fujimori1,
  2. Masatoshi Wakui2,
  3. Hisako Katagiri1,
  4. Kentaro Ohira1,
  5. Nobuko Shimizu1,
  6. Mitsuru Murata2
  1. 1Central Clinical Laboratory, Keio University Hospital, Tokyo, Japan
  2. 2Department of Laboratory Medicine, School of Medicine, Keio University, Tokyo, Japan
  1. Correspondence to Dr Masatoshi Wakui, Department of Laboratory Medicine, School of Medicine, Keio University, Shinjuku-ku, Tokyo 160-8582, Japan; wakuism{at}

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Direct thrombin inhibitors (DTIs) represented by dabigatran were expected to be available for therapeutic use without the need for routine monitoring, in contrast to warfarin. However, clinical concerns regarding impacts of plasma dabigatran concentrations on the rate of major bleeding have been raised.1 Clinical and basic aspects of DTIs aid in studying how best to address concerns regarding bleeding risk in therapeutic use. Based on enzymatic examinations with a synthetic substrate, S-2238, it has been well recognised that dabigatran and another DTI argatroban reversibly and competitively inhibit thrombin reaction.2 ,3 However, it still remains unclear whether the type of inhibition by DTIs of thrombin reaction with S-2238 can be applied to the anticoagulant effects.

The Clauss assay is classical but most popular even now for quantification of plasma fibrinogen concentrations.4 This is based on the quantitative relationship between fibrinogen concentrations and time for fibrin clot formation by thrombin. As a high concentration of thrombin ranging from 40.25 to 115 IU/mL is added to dilute test plasma, the clotting time depends exclusively on concentrations of fibrinogen but not of other coagulation factors.

The Lineweaver–Burk plot, which is one of classical linear transformations of the Michaelis–Menten equation for enzymatic reaction kinetics, presents curve-linearity between the reciprocal of the substrate concentration versus the reciprocal of the rate of reaction.5 When used for determining the type of enzyme inhibition, this plot can distinguish competitive, uncompetitive, non-competitive and mixed inhibitors.

To assess how DTIs affect in vitro clotting from the enzymatic perspective, the Lineweaver–Burk plot was …

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  • Contributors YF and MW equally contributed to this work. MW conceived the study. YF, MW, HK and NS designed the study. YF and KO performed the experiments. YF and MW analysed the data. YF, MW, HK and MM participated in the discussion. YF and MW wrote the manuscript. All authors have given final approval for this version of the manuscript to be published.

  • Funding This work was supported by the JSPS Grant-in-Aid for Scientific Research (A) (research project number 23249030) for MM and by the Keio Gijuku Academic Development Funds for HK.

  • Competing interests None declared.

  • Provenance and peer review Not commissioned; externally peer reviewed.