| Objective:A novel amphiphilic core-shell micell based on a pectin-doxorubicin conjugates macromolecular pro-drug were designed and synthesized for efficient antineoplastic drug delivery. Besides, we further reported the evaluation of its sustained-release characteristics, toxicity, biocompatibility and in vivo pharmacokinetic of PDC-M.Methods: 1. Synthesis of PDC: pectin was conjugated to doxorubicin via an amide condensation reaction. The structure of targetedproduct was confirmed by NMR and FTIR. 2. Preparation and characterization of PDC-M: PDC-M were prepared by self-assembled in situ, with pectin as the hydrophilic shell and doxorubicin(DOX) as the hydrophobic core. Single factor analysis was used to optimize the preparation conditions. Scanning electron microscopic was used to observe morphology of nano-micelles. The micells size, distribution and stability were analyzed by nano ZS90. Besides, drug loading efficiency was measured by UV spectrophotometer. 3. The in vitro study on anti tumor effect and sustained release of PDC-M: the drug release performance in different p Hs in vitro were studied under simulated conditions; Anti proliferative effect of PDC-M and DOX in HepG2 was determined by CCK8. Absorption of PDC-M and DOX in HepG2 was observed by flow cytometry. 4. The biocompatibility study of PDC-M in vitro: cell viability of PDC-M and DOX in endothelial cells was evaluated by CCK8; Hemolysis activity was investigated in rabbits and protein adsorption of PDC-M and DOX was tested by bovine serum albumin. 5. Pharmacokinetics study of PDC-M: sprague Dawley(SD) rats were treated by tail vein i.v. injection with PDC-M and DOX, the blood concentration was detected by High Performance Liquid Chromatography(HPLC), and the pharmacokinetic parameters were obtained by running software 3p97.Results: The chemical structure of the PDC macromolecular pro-drug was identified by both Fourier transform infrared spectroscopy(FTIR) and nuclear magnetic resonance spectroscopy(1H-NMR), and proved that doxorubicin combined well with the pectin and formed macromolecular pro-drug. Then, it was found that PDC-M was only formed under certain condition:p H5.0, 50℃, 100 rpm. The PDC-M were observed to have an unregularly spherical shape and were uniform in size and the average particle size was about 140 nm. The encapsulation efficiency and drug loading were 57.82% ± 3.7%(n=3) and 23.852% ± 2.3%(n=3), respectively. In vitro drug release study showed that PDC-M displayed a prolonged slow release profile in different p Hs. Assays for antiproliferative effects and flow cytometry of the resulting PDC-M in HepG2 cell lines demonstrated greater properties of delayed and slow release as compared to free DOX. A cell viability study against endothelial cells further revealed that the resulting PDC-M possesses excellent cell compatibilities and low cytotoxicities in comparison with that of the free DOX. Hemolysis activity was investigated in rabbits, and the results also demonstrated that the PDC-M has greater compatibility in comparison with free DOX. The pharmacokinetic results indicate that PDC-M can extended half-time, reducing toxic and side effects, improved bioavailability.Conclusion: A self-assembled amphiphilic core-shell micelle based on a macromolecular pro-drug was successful in an in-situ one step fabrication of the hydrophobic drug DOX as the core and pectin as the hydrophilic shell. The resulting PDC-M displayed a slow release and stable properties. Besides, the PDC-M possessed better biocompatibilities and lower cytotoxicities in comparison with free DOX. Otherwise, PDC-M can extended half-time, reducing toxic and side effects, improved bioavailability. This demonstrates a great potential for use as a drug delivery system in cancer chemotherapy. |
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