Purification And Characterization Of Analgesic Peptides From Naja Atra Venom Digested By Pepsin

Snake venoms are complex mixtures of pharmacologically active peptides and proteins,three-finger proteins belong to the family of proteins.In recent years,three-finger proteins have become a hot topic for non-addictive,undependent,small doses and long-acting analgesic effect.Reveal the mechanism of its analgesic effect and than develop an analgesic drug with high selective,good security and less side effect has become an urgent problem need to be addressed.In this dissertation,the author aims to explore analgesic peptide based on snake venom three-finger protein and research its analgesic effect.To find an analgesic peptide which is an oral medication and can quick onset through isolation and purification of naja atra venom digested by pepsin.Lay a foundation for the study of the analgesic mechanism of three-finger proteins and provide a reference for related research.In this research,firstly,cobra venom was processed in boiling water bath to obtain a high thermal stability of alkaline protein mixture containing three-finger proteins,and then digested cobra venom by simulated gastric conditions with different time gradient.The analgesic activity of the hydrolysate was observed by hot-plate test.Purification and identification of purity of the hydrolysate by HPLC.Collected the peptide,the analgesic activity of peptide was also observed by hot-plate test.The results showed that hydrolyzed cobra venom has significant analgesic effect after intragastric on mice,the analgesic activity of venom which was hydrolysed by pepsin for 6 hours was best,it could took effect in about 10 minutes,and the effect could remained stably for at least 48 hours;the results of intraperitoneal injection group was not,with the deepening of the degree of hydrolysis even to completely hydrolysis,it almost has no analgesic effect.After purified by HPLC,the component which was collected in 12 min was more consistent with previous results.Freeze-drying the component,Identification of the nature of the component by MALDI-TOF,the molecular weight is about 1267.727 and it's amino acid sequence was—KDHRGTRIER by comparison with the database.Synthesized the peptide,the analgesic activity of synthesized peptide was very consistent with previous results after intragastric on mice.The analgesic peptide was obtained by purification,characterization and synthesis,it can take orally and faster-working,but its analgesic mechanism needs further study.