Thermosensitivity Hydrogel Nanoparticles Preparation And Application In Peptide Recognation

Thermosensitive hydrogel nanoparticle is a kind of high molecular polymerwhich has outstandingly temperature sensitive property. The particle is normallyreserved in water or aqueous solution and its phase transition happened when thecurrent temperature reaches its lowest critical solution temperature (LCST). Atpresent, the thermosensitive hydrogel nanoparticles (NPs) copolymerized byN-isopropylacrylamide (NIPAm) together with other ionic monomers have causedextensive attention.In this paper, a series of NPs were prepared while NIPAm, acrylic acid (AAc)and N-t-butylacrylamide (tBAm) as the structured monomer, electro-affinitymonomer and hydrophobical monomer in different molar ratio. Furthermore, thesurfactant, crosslinker, catalyzer and initiator in the NPs synthesis process wassodium dodecyl sulfate (SDS), N,N’-methylenebisacrylamide (BIS),N,N,N’,N’-tetramethylethylenediamine (Temed) and ammonium peroxydisulfate(APS), respectively. The prepared monodisperse NPs were characterized byscanning electron microscopy (SEM) and dynamic light scattering (DLS). In orderto control the particle size, we studied the dosage of SDS and AAc in theexperiment. The particle size decreased from460to70nm while SDS dosageincreased from0to50mg; the particle size increased from74to613nm whileAAc increased from5%to40%(mol%), meanwhile, the absolute z-potential valueof NPs increased and the system tended to stabilization. NPs maintained uniformstability and large particle size below15℃, but began to shrink rapidly when thetemperature rose to20-30℃.The NPs affinity to his-tagged peptide whose sequence isNH2-LEVLFQGPHHHHHH-COOH and HOOC-LEVLFQGPHHHHHH-NH2was studied using high performance liquid chromatography (HPLC) analysis inthis research. NPs with smaller particle size showed higher affinit y to the targetpeptide. It was also proved that the amount of AAc in the NPs, the electroaffinity monomer, gave great influence on the peptide recognition. NPs ofNIPAm/tBAm/AAc/BIS (43/40/15/2, mol%) had the highest absorption rate asmuch as78%to target peptide. Moreover, the absorption rate rised as buffersolution pH value droped.The NPs affinity to melittin whose sequence is NH2-GLY-ILE-GLY-ALA-VAL-LEU-LYS-VAL-LEU-THR-THR-GLY-LEU-PRO-ALA-LEU-ILE-SER-TRP-ILE-LYS-ARG-LYS-ARG-GLN-GLN-COOH was also studied byHPLC method, polyacrylamide gel electrophoresis （SDS-PAGE） andultraviolet-visible spectrophotometry. The adsorption rate to melittin increasedalong with the NPs size decreased. The amount of tBAm in the NPs, thehydrophobic monomer, gave great influence on the melittin recognition. However,the amount of AAc had little effects on the melittin recognition. NPs’ catch andrelease capability was also studied with adjusting the temperature consideringtheir thermosensitivity.