Synthesis And Liver Targeting Performance Of Dual-ligand Modified Chitosan As A Drug Carrier

At present, the researches of drug carrier in hepatic targeted drug delivery system in and abroad are focus on the single-ligand modified nanoparticles. However, the interaction of receptor-ligand of a single-ligand modified nanoparticles could are variable with the pathology and physiological conditions, which could reduce the specific recongnition ability and the binding activity of receptor. In view of this case, a dual-ligand modified drug carrier material is put forward to improve the reliability of the active targeting and enrich the content of the hepatic targeted drug delivery system.In this study, a novel material as a liver targeting vector, dual-ligand modified chitosan (GCGA) composed of chitosan (CTS), glycyrrhetinic acid (GA) and lactobionic acid (LA), was firstly synthesized by N-acylation reaction, and the structure has not been reported in the literature. We firstly prepared the FITC-labeled GCGA nanoparticles (FITC-GCGA NPs) and5-fluorouracil-loaded GCGA NPs (GCGA/5-Fu NPs) by Ionic cross-linking. Through advanced testing technology, we analysised the physical and chemical structure of GCGA, estimated the cytotoxicity and liver targeting performance, optimized preparation conditions of GCGA/5-Fu NPs, investigated the In vitro drug release characteristics of GCGA/5-Fu NPs, and studied the mechanism of active targeting. The conclusions are as follows.(1) The results of FTIR spectra,1H-NM R spectra, contact angle measurement and MTT assay show:GA and LA were introduced into CTS after N-acylation reaction chains, and the degree of substitution (DS) of GA coupled with CTS in GCGA was estimated to be13.77, and LA16.74mol%; GCGA is an amphiphilic drug vector with hydrophilic LA group and hydrophobic GA group and can form self-aggregated polymeric micelles; The cytotoxicity of CTS and GCGA was concentration dependent which was inverse proportion to the cell viability by MTT assay using L929cell line, and GCGA is no toxicity with a concentration range of0.005-0.05mg/ml.(2) The study of cellular uptake in vitro and the biodistribution in SD rats showed:GA and LA ligands on the FITC-GCGA NPs bind to cell-surface relevant receptors, which trigger internalization of the FITC-GCGA NPs into the cell; With the retardation of a receptor-mediated endocytosis due to certain pathophysiological reasons, there is still another direct interaction of ligand-receptor; Approximately a18.22-fold improvement in BEL-7402hepatoma cells uptake of the GCGA NPs than LO2normal liver cells, suggesting FITC-GCGA NPs has a higher affinity to hepatoma cells; The tissue biodistribution study in SD rats show: FITC-GCGA NPs had no toxic and side effects upon heart, brain and skeletal muscle; Compared with FITC-CTS NPs and FITC-GA-CTS NPs, the FITC-GCGA NPs resulted in higher levels of the bioaccumulation quantity of NPs in the liver. Therefore, GCGA showed great potential to be served as a liver targeting vector of drug.(3) We optimized the mean particle size and drug content of CTS/5-Fu NPs by central composite design and showed that5-Fu/CTS had obvious influence on total desirability function; With the CTS/5-Fu NPs model, GCGA/5-Fu NPs prepared had the nearly spherical shape with a mean particle size of239.9nm (major size range of100-500nm), polydispersity index of0.04, zeta potential of+21.2mV, and drug content of3.90%(w/v). In vitro drug release study of GCGA/5-Fu NPs in pH7.4PBS buffer solution showed a four phase composition: a first initial burst release, sustained release, the second burst release, a constant sustained release of the drug later. GCGA NPs, as a nano-drug carrier in the drug controlled release system, had the properties of a sustained release form.