Clinical laboratory monitoring of a synthetic antithrombin agent, argatroban, using High Performance Liquid Chromatography and functional methods

Background. Argatroban is a peptidomimetic inhibitor of thrombin which is i n clinical trials for thrombotic complications. Clot-based assays measure t he cumulative anticoagulant effect of argatroban and its metabolites(s). To monitor the absolute concentrations of argatroban, a specific HPLC method was developed. Methods. Validation studies included normal volunteers administered with es calating doses of argatroban (ARG 102 Study), patients undergoing coronary interventional procedures (ARG 310), and patients receiving argatroban in c onjuction with streptokinase for acute myocardial infarction (ARG 230). Pla sma samples were extracted with acetonitrile and reconstituted in a mobile phase. UV detecion was made at 333 nn. Calibration curves were prepared wit h known standards of argatroban in normal human plasma, Results. The retention time for argaeroban was 6.0 +/- 0.5 min and the extr action efficiency was >98% (r(2)=0.99). In the ARG102 Study, argatroban lev els sere: 0.84+/-0.23 (day 1), 1.55+/-0.34 (day 2), 2.92+/-0.15 (day 3), an d 3.04+/-0.49 (day 4). In the ARG310 trial, the mean argatroban levels were : 0.23+0.09 mu g/ml (preinfusion), 5.77+/-0.92 mu g/ml (postinfusion/intrap rocedure), and 2.23+/-0.29 mu g/ml (postprocedure). In the ARG 230 Study, t he mean argatroban levels at 2-8 hrs were between 1.5-2.0 mu g/ml. Upon com pletion of the infusion, a time-dependent clearance of argatroban was noted . Conclusions. Since heparinization, hemodilution and hypofibrinogenemia due to thrombolysis influence the clotting tests, absolute quantitation of arga troban by HPLC in these patients provides a more reliable means of monitori ng this anticoagulant and helps in the dosage-optimization of this agent. T he current HPLC method is of value in the monitoring of paients who are sim ultaneously administered with thrombolytic drugs.