Study On Self-assembly Of Chiral Nonapeptides With Rapid Hemostasis Activity

Peptide self-assembly is widely spread in nature.In recent years, amino acid residue isoften used to design peptides, which has become one of the hottest topics in the field ofnanomaterials. Through the rational design of the amino acid residue sequence, peptidemolecules can take advantage of electrostatic interactions, hydrophobic interactions, hydrogenbonding, π-π stacking interactions and other non-covalent interactions to form self-assemblednanostructures spontaneously. In addition, the D-form amino acid has good biocompatibilityand can not be hydrolyzed by naturally occurring L-form amino acid degrading enzymes.Therefore, the use of the D-form amino acid self-assembled technology to build variousfunctional materials in biological engineering, drug delivery, tissue engineering and otherfields has great application prospects.The pre-hospital control of bleeding wound has becomethe focus of the medical profession.The peptide,consists of amino acids, is different from thetraditional hemostatic material.The peptide hemostatic materials show less prones to rejectionor immune response in vivo,so that they are expected to replace the traditional hemostaticmaterials to achieve rapid hemostasis.In this thesis,a pair of self-assembled nonapeptides（D-form nonapeptide and L-form nonapeptide） were designed and synthesized, and thephysicochemical properties of these nonapeptides were studied.In addition, the self-assembledstructures and morphologies in NaCl solution were also investigated.Finally,their rapidhemostasis activity was further proved.The amino acid residue sequence of L-form and D-form nonapeptides isAc-Pro-Glu-Phe-Arg-Phe-Asn-Phe-Gln-Pro-NH₂, which only comprises a pair of chargedamino acid residues: arginine （Arg） and glutamic acid （Glu）.The two prolines （Pro） are placedin the head and the tail of the nonapeptide chain,respectively. The remaining amino acidresidues are mainly composed of polar amino acid residues. The physicochemical propertiesand morphologies of L-P9and D-P9were characterized by circular dichroism （CD）spectroscopy, atomic force microscopy （AFM）, dynamic light scattering （DLS） andfluorescence spectrophotometer. In aqueous solution, the critical micelle concentration （CMC）of L-P9and D-P9is about0.23mM and0.4mM, respectively. Their secondary structureswere neither typical α-helix, nor neat β-sheet, but mixed secondary structures. When theconcentrations of L-P9and D-P9aqueous solution exceed their CMC values, nanofibers wereformed. With the concentration of nonapeptide increasing, nanofibers intertwined together toform nanofiber networks. In the NaCl solution, the CMC values of L-P9and D-P9reduce to 0.1mM and0.125mM,but their secondary structures were not affected,indicating that theNaCl solution does not affect L-P9and D-P9self-assemble into nanofibers,but accelerate theformation of self-assembled nanostructures.Furthermore,at high concentration of NaClsolution,L-P9and D-P9formed into thicker nanofiber to achieve stability.The results showthat physicochemical properties and self-assembling behaviors of L-P9and D-P9can beappropriately controlled by changing the concentration of NaCl solution.Based on the study of physicochemical properties, self-assembling structures andmorphologies of L-P9and D-P9, we used Sprague-Dawley rat kidney and liver hemostasismodels to study their rapid hemostasis activity. It was found that L-P9and D-P9could stopkidney and liver bleeding at9.2s,6.7s,10.2s and9s, respectively.When compared with thecotton swabs control,It was revealed that L-P9and D-P9can suppress wound bleedingquickly and effectively.Based on the above results,we suggested the possible hemostaticmechanism of the nonapeptides was discussed,and "a coagulation bed" bleeding model wasproposed.This study also provides a theoretical basis to the further design of such chiralpeptide nanomaterials.