Preparation and antithrombotic activity identification of Perinereis aibuhitensis extract: a high temperature and wide pH range stable biological agent

Abstract

In this work, a thermal and wide pH range (from 1 to 13) stable biological agent was extracted from Perinereis aibuhitensis (P. aibuhitensis), and the antithrombotic activity of this P. aibuhitensis extract (PaE) was investigated. In vitro antithrombotic activity test indicated that PaE could significantly extend the clotting time of recalcified rabbit blood: the clotting time of the 10 mg mL⁻¹, 20 mg mL⁻¹ and 40 mg mL⁻¹ PaE groups increased by 24.24%, 68.64% and 130.76%, respectively. Further platelet poor plasma (PPP) coagulation study proved that PaE could obviously inhibit the intrinsic coagulation pathway and slightly inhibit the extrinsic coagulation pathway. Moreover, PaE could effectively inhibit fibrinogen turning into fibrin induced by thrombin. More importantly, PaE exhibited thermal stability, since P. aibuhitensis was boiled in the PaE preparation process, as well as tolerance to strong acid and alkali, since its fibrinogen inhibition activity was retained after treatment with a wide range of pH value conditions. Finally, the antithrombotic activity ingredients in PaE were preliminarily identified, the results of which indicated that the ingredients were ethanol insoluble, insensitive to papain and pepsin, and could be degraded by H₂O₂. Additionally, we demonstrated that PaE at a concentration of not more than 20 mg mL⁻¹, the concentration at which PaE showed high anticoagulant activity, exhibited favorable biocompatibility via in vitro hemolysis rate test and MTT assay. In summary, the results evidenced that this conveniently prepared and stable PaE was a safe and effective antithrombotic agent, and had the potential to be utilized for antithrombotic food development.