The feasibility of separation of activated prothrombin complex (APC) solution via ultrafiltration was investigated. To separate thrombin from high molecular weight impurities, polyethersulfone hollow fiber ultrafiltration membrane PES-HF-100 (manufactured by Institute of Physical Organic Chemistry of the National Academy of Sciences of Belarus) was used, further concentration of the target filtrate of PES-HF-100 was carried out using polysulfone membrane PS-PV-20. It was found that ultrafiltration methods are suitable for isolation of thrombin from APC. Comparative analysis of prepared samples with commercial analogs of the coagulation reagent “Thrombin” established that thrombin isolated via ultrafiltration is not inferior in purity to thrombin isolated by chromatography.