The Separation And Purification Of Small Peptides From The Venom Of Akigstrodon Acutus And Study Their Anti-platelet Activities

Objective Thrombotic disease is a disease with a high incidence and mortality rates in the world,the small peptides of snake venom have an effect in anti-platelet aggregating.Anti-platelet plays an important role in cure thrombotic disease,we intend to isolate and purify the small peptides from the venom of snake for the purpose of thrombotic disease.Methods ① The small peptides from the venom of Agkistrodon acutus are isolated through Sephadex G-50 and RP-HPLC and identified with MS and NMR analysis;②Assess the inhibitory activity of small peptides on human platelet aggregation;③ Use ADP to induce the acute pulmonary thrombosis in mice,assess the time of breathing palpitations,the number of platelets and the number in the microthrombusin lung tissue;④ HPLC measurements assess the inhibitory activity of small peptides on angiotensin-converting enzyme(ACE)with by RP-HPLC;⑤ Assess the risk of small peptides by cutting tail test on mice.Results ① Two tripeptides,Pt-A with sequence Glu-Asn-Trp(429 Da)and Pt-B with sequence Glu-Gln-Trp(443 Da),in venom of Agkistrodon acutus were isolated through Sephadex G-50 and RP-HPLC and identified with MS and NMR analysis;② These two small peptides showed inhibitory activity in human platelet aggregation,especially Pt-A displayed a potent anti-platelet activity with a IC50 value of 66μM,while the value of Pt-B was 203μM;③In vivo studies showed that Pt-A prevented mice from acute pulmonary thrombosisat a dose of 20 mg/kg,and the dose of Pt-B was about 40 mg/kg;④Pt-A and Pt-B had a weak inhibitory activity of ACE,the IC50 value of Pt-A was 0.34mM,and the value of Pt-B was 1.08mM;⑤Tail bleeding indicated that Pt-A and Pt-B were devoid of hemorrhagic activity at a dose of 40 mg/kg.Conclusion Peptide A and peptide B from the venom of Agkistrodon acutus had a good effect of prevention and treatment of thrombosis;this may be related to the ability in anti-platelet aggregation,and they had no bleeding tendency.