Preparation And Properties Of Thrombus-targeted Chitosan Modified MWCNT Drug Carrier

This study intends to use multi-walled carbon nanotubes（MWCNTs）as the carrier matrix,RGD short peptides as target molecules,low molecular weight chitosan（CS）as the connecting arm,and urokinase（UK）as the model drug.A new thrombolytic drug delivery system for thrombus targeting and sustained release function to improve the therapeutic effect of the first-generation thrombolytic drugs,prolong the action time,reduce the dosage and side effects,and is a new drug delivery system for carbon nanotubes.The study provides experimental evidence.The carboxylated multi-walled carbon nanotubes（MWCNT-COOH）were prepared by oxidation of the original MWCNTs by mixed acid oxidation method.UV-visible spectrophotometry（UV）,infrared spectroscopy（FT-IR）and transmission electron microscopy（TEM）were used.The surface morphology and surface groups of MWCNT-COOH were characterized by other methods.The surface carboxyl group content was quantitatively analyzed by titration method.The factors affecting the carboxyl content of the product by acid oxidation method were studied.Low molecular weight chitosan was prepared by degrading large molecular weight chitosan by hydrogen peroxide（H₂O₂）oxidation method.The relative molecular mass of chitosan before and after degradation was determined by viscosity method and end group method.The RGD short peptide was covalently attached to chitosan by catalyst EDC and NHS to prepare chitosan RGD derivative（CS-RGD）.The CS-RGD was characterized by infrared spectroscopy and elemental analysis.The CS-RGD and MWCNT-COOH are self-assembled by electrostatic action to form a complex,and the thrombolytic drug urokinase（UK）is loaded onto the surface and inside of the carrier byπ-πstacking to form MWCNT/CS-RGD/UK drug carrier.The complex was analyzed by UV-Vis spectrophotometry,Zeta potential analysis,thermogravimetric analysis（TGA）and transmission electron microscopy（TEM）.The in vitro release of MWCNT/CS-RGD/UK drug in pH 7.4 medium was investigated,the release data was fitted,and the encapsulation efficiency and drug loading of MWCNT/CS-RGD/UK were determined.The in vitro thrombolytic effect of the MWCNT/CS-RGD/UK drug delivery system was investigated using the fibrin plate method.MWCNT-COOH with good water dispersion was successfully prepared by oxidative purification modification of MWCNT and characterization of its products.Infrared spectroscopy and titration analysis showed that the degree of carboxylation was affected by reaction temperature,reaction time and ultrasonic oscillation time.The temperature was 60°C,the reaction time was 2 h,and the ultrasonic vibration time was 30 min.The macromolecular chitosan was oxidatively degraded by H2O2,and the molecular weight was changed from 106 Da to 1000 Da,and the water-soluble low molecular weight chitosan was successfully prepared.After catalytic preparation,CS-RGD was successfully prepared.The results of infrared analysis and elemental analysis confirmed that chitosan andRGD were covalently bound.MWCNT/CS-RGD/UK was successfully prepared by molecular self-assembly method.TGA and Zeta potential analysis indicated that CS-RGD was bound to MWCNT-COOH surface,and TEM analysis showed that UK was loaded onto MWCNT/CS-RGD surface.The release of drug-loaded system in pH 7.4 medium has a certain sustained release characteristics,and the fitting results show that it is consistent with the first-order release;the encapsulation efficiency and drug loading are 83.45%and 13.12%,respectively.The fibrin plate showed that the UK in MWCNT/CS-RGD/UK had better fibrinolytic activity,and there was no significant difference compared to fresh UK.This study successfully constructed a MWCNT/CS-RGD/UK delivery system with a certain sustained release effect,which has a high encapsulation efficiency for urokinase.The delivery system has obvious in vitro thrombolytic effect,which can improve the therapeutic effect of the first-generation thrombolytic drug,prolong the action time,and reduce the side effects.This study laid the foundation for the construction of sustained-release and targeted preparations of thrombolytic drugs with aromatic structures such as urokinase,and provided new ideas for the research of carbon nanotube-based novel delivery systems.