Preparation And Functional Study Of PH-Enzyme Responsive Amorphous Calcium Phosphate Nanocarriers

Cancer is becoming an important factor that jeopardizes national public health security,making the integration of disease diagnosis or monitor and treatment to achieve theranostics popular for researchers.Conditionally responsive nano-drug carriers are favorite for their controlled-release properties.Second,carbon quantum dots are widely used in bio-imaging,photocatalysis and sensors owing to their excellent biocompatibility and photostability etc.Previously,amorphous calcium phosphate（ACP）nano-drug carriers were prepared in our group in which calcium nitrate（Ca（NO₃）₂）and ammonium phosphate（（NH₄）₂HPO₄）were as calcium and phosphorus sources,respectively,and their pH-responsive release properties were investigated.However,there were the problems of tumor-targeting and imaging capabilities about ACP,leading to no accurate treatment for lesions.Therefore,it is imperative to design a nano-drug carrier that can improve tumor-targeted,admirable fluorescence and conditionally responsive drug release performance.In this thesis,carbon quantum dots with similar properties were prepared by two methods and applied in drug carriers.In addition,the related mechanism and biological evaluation of composite nano-drug carriers were studied.Two water-soluble carbon quantum dots HA-FCNs-1 and HA-FCNs-2 were synthesized from hyaluronic acid（HA）through high-temperature dehydration and hydrothermal method,respectively.Additionally,multiple testing instruments were also introduced to investigate the basic properties.Transmission electron microscope（TEM）and X-ray diffractometer（XRD）were used to characterize their morphology and structure.Fourier transform infrared（FTIR）and ¹H nuclear magnetic resonance（¹H NMR）were applied to study their chemical composition.The biocompatibility and tumor-imaging performance were explored through experiments such as laser confocal microscopy imaging（CLSM）and cytotoxicity testing.The results showed that the prepared HA-FCNs-1 and HA-FCNs-2 both emitted bright blue fluorescence under the irradiation of 365 nm UV lamp,exhibited a graphene-like structure,retained the partial structure of HA,and had good biocompatibility,as well as tumor-targeting and bioimaging performance.Amorphous calcium phosphate composite nano-drug deliveries（HA-FCNs/ACP）were prepared by co-precipitation method,and curcumin（Cur）as template drug molecule was loaded by blending method.The optimal conditions,structure,morphology and particle size distribution of the synthesized HA-FCNs/ACP were tested by XRD,scanning electron microscopy（SEM）and laser particle analysis.The fluorescence performance was characterized using a 365 nm UV lamp,and FTIR analysis was adopted to infer the synthesis mechanism.Furthermore,drug release and targeted imaging performance were tested by drug sustained release and CLSM.The results showed that HA-FCNs/ACP presented an amorphous structure when the concentration of HA-FCNs was not less than 600μg/m L.Besides,the prepared HA-FCNs/ACP was spherical particles of about 172 nm,and the particle size distribution was relatively uniform.Under excitation with a lamp at 365 nm,HA-FCNs/ACP strongly emitted a blue color.In additionally,HA-FCNs/ACP had good biocompatibility,tumor-targeted imaging and pH-responsive drug release.To meet the application requirements of drug carriers,Cur and doxorubicin（DOX）were loaded by blending method,respectively.The hemolysis assay and drug release were used to characterize blood compatibility and release performance,respectively.The distributions of HA-FCNs and HA-FCNs/ACP in cancer cells were observed,and the effects of samples before and after loading drug on cancer cell migration were investigated by Transwell test.The results showed that HA-FCNs/ACP had outstanding blood compatibility,pH-responsive and enzymatic drug release performance before and after loading drug.HA-FCNs and HA-FCNs/ACP dispersed in the cytoplasm through the overexpressed CD44 receptor that actively targeted into A549 human lung cancer cells.On the other hand,the migration of A549 cells would be inhibited after cells were treated with drug-loaded nanomaterials.