Preparation And Characterization Of 5-fluorouracil-Fe/Fe₃O₄ Magnetic Liposome And Study On Anti-tumor Effect In Vitro

Tumor had a serious threat to human health. Traditional cancer treatment method mainly had surgery, radiotherapy and chemotherapy. But surgical larger traumatic and radio-chemotherapy relatively strong side effects made traditional treatments in bottleneck. Nano magnetic liposome of targeted drugs, which had a certain amount of magnetic and good biocompatibility, had become recent cancer research hotspots. In recent years, both at home and abroad, the widespread use of magnetic nano-materials mainly had iron monomer, ferrite, such as Fe₃O₄, Fe₂O₃, carbon iron compound, manganese zinc ferrite compound and so on. Iron monomer magnetic was stronger than other magnetic materials, but its nature was extremely unstable in nanometer scale and easy oxidation. The nature of ferrite was stable, had good biocompatibility, and could regularly be excluded from body, but its magnetic response ability was weak. Therefore, it had a great significance for magnetic liposome drug research on preparation high magnetic and biocompatibility magnetic particles.5-fluorouracil-Fe/Fe₃O₄ magnetic liposome (5-Fu-Fe/Fe₃O₄-MLP) was designed and prepared in this experimental. Magnetic-materials were Fe/Fe₃O₄ core-shell structural nano-particles, which had high magnetic and good biocompatibility. We tested 5-Fu-Fe/Fe₃O₄-MLP and Fe/Fe₃O₄ nano-particles about particle size, encapsulation efficiency, stability and saturation magnetization strength, then researched cytotoxicity of Fe/Fe₃O₄ nano-particles and anti-tumor effect of 5-Fu- Fe/Fe₃O₄-MLP in vitro.Methods1. Preparation, characterization and cytotoxicity of Fe/Fe₃O₄ nano-particlesThe chemical co-precipitation method was used to prepare Fe₃O₄, The reduction-oxidation method was used to prepare Fe/Fe₃O₄ core-shell structural nano-particles. That was, Fe₃O₄ was reduced to Fe in atmosphere reactor. After that, Fe was slowly oxidized to Fe/Fe₃O₄ nano-particles in atmosphere reactor. The characterization of Fe/Fe₃O₄ nano-particles was determined by X-ray diffraction (XRD), transmission electron microscopy (TEM) and vibrating specimen magnetometer (VSM).Under 4250 gaussian magnetic field, LO2 human normal liver cells incubated with the leach liquor of Fe/Fe₃O₄ nano-particles in different concentrations for 48 hours, the cell viability was examined by MTT colorimetric assay.2. Preparation, characterization and anti-tumor effect of 5-Fu-Fe/Fe₃O₄-MLPUsing reverse-evaporation method to prepare 5-Fu-Fe/Fe₃O₄-MLP, and the best formula of 5-Fu-Fe/Fe₃O₄-MLP was obtained through the four factors of three levels orthogonal experiment. The characterization of 5-Fu-Fe/Fe₃O₄-MLP was measured with particle size analyzer (PCS) and TEM. The entrapment efficiency was determined by gel-chromatography. Magnetic field was used to detect the magnetic responsiveness of 5-Fu-Fe/Fe₃O₄-MLP in vitro.Under 4250 gaussian magnetic field, HepG2 human hepatocarcinoma cells incubated with the standard solution of 5-Fu, 5-Fu-Fe/Fe₃O₄-MLP and Fe/Fe₃O₄-ML- P in different concentrations and time, cell viability was examined by MTT colorimetric assay. Results1. Fe/Fe₃O₄ nano-particles was similar to a globe with the average particle size of 70 nm, XRD and TEM observation shown, its shell was Fe₃O₄ and kernel was Fe, Magnetic saturation intensity of Fe/Fe₃O₄ nano-particles was 107.54emu/g. which was stronger than Fe₃O₄(77.72emu/g). LO2 normal liver cells incubated with leach liquor of Fe/Fe₃O₄ nano-particles in larger concentration range from 0 to 100mg/ml, The determined by MTT results are shown, the cytotoxicity of Fe/Fe₃O₄ nano-particles grading was 0 1 grade, which had good biocompatibility.2. The average particle size of 5-Fu-Fe/Fe₃O₄-MLP was 202.5 nm with 33.5% entrapment efficiency, which was placed a month in 4℃refrigerator and had a 30% of encapsulation efficiency, there was no precipitation in the bottle. In vitro 4250 gaussian magnetic field, 5-Fu-Fe/Fe₃O₄-MLP gathered at a faster pace than 5-Fu- Fe₃O₄ magnetic liposome, which showed a good magnetic responsiveness.The result of MTT colorimetric assay demonstrated that under the action of the 4250 gaussian magnetic field, the 5-Fu-Fe/Fe₃O₄-MLP had certain inhibition to the growth of HepG2 cell, the cells relative inhibition rate of which containing 5-Fu from 0.6 to 6μg/ml were range from16.5% to 54.7% in 48h, the group of 5-Fu-Fe/Fe₃O₄-MLP, which had concentration dependence, had a better relative inhibition rate than the group of 5-Fu (P<0.05). Within 12 to 60 hours, the cells relative inhibition rate of 5-Fu-Fe/Fe₃O₄-MLP containing 5-Fu 6μg/ml gradually strengthened and stability as incubation time prolongs, which had time dependence and slow-releasing potential.ConclusionsWe successfully prepared the Fe/Fe₃O₄ magnetic nano-particles, the preparation method is simple and practicable, and preparing magnetic particles had a good biocompatibility. 5-Fu-Fe/Fe₃O₄-MLP had good magnetism responsiveness and anti-tumor effect, which can be stored in 4℃refrigerator for a long time, and had a better inhibition rate than 5-Fu crude , and had concentration and time dependence and slow-releasing potential. This experiment provided a novel beneficial attempt for improving the magnetic targeted ability of magnetic liposome, and contributed to magnetic targeted drug carrier application in cancer treatment.