| ObjectiveMagnetite nanoparticles were respectively surface-modified with gluconic acid (GLA) and chitosan oligosaccharide (CSO) to improve their hydrophilicity and bio-affinity. Moreover, the adsorption property of the magnetic nano-adsorbent for BSA was also investigated under the different conditions including adsorption time, pH and concentration of bovine serum albumin. Meanwhile biocompatibility GLA-Fe3O4 and two CSO-Fe3O4 magnetic nanoparticles were studied. The study is the possible mechanism of mouse macrophages RAW264.7 in vitro interact with GLA-Fe3O4 and CSO-Fe3O4. This study provides experimental evidence for the application of magnetic nanoparticles in the field of medicine and biological adsorption.MethodsFe3O4 magnetic nanoparticles were fabricated by a chemical co-precipitation method. Chitosan oligosaccharide was grafted onto the surface of Fe3O4 by employing (3-chloropropyl) trimethoxysilane as a coupling agent. Infrared spectroscopy (FT-IR), and thermal gravimetric analysis (TGA) can detect the composition of the nanoparticles and coverage of modified substances on the surface of the nanoparticles. X-ray diffraction (XRD) can measure the structure of crystals. Nano-sizer is used to test the surface potential of the nanoparticles, morphology of the magnetic nanoparticles is observed by scanning electron microscopy (SEM).Under the univariate variables (time, pH and initial concentration of BSA) conditions, BSA interacted with Fe3O4, GLA-Fe3O4 and CSO-Fe3O4. Using UV spectrophotometer, BSA concentration in the supernatant was measured wavelength at 595nm by Coomassie blue method (Bradford method). And calculate the amount of magnetic nanoparticles adsorption of BSA.The biocompatibility of GLA-Fe3O4 and CSO-Fe3O4 were evaluated in vitro via MTT assay and hemolysis test. The nanoparticles showed no cytotoxicity to mouse macrophage cells (RAW264.7) and the hemolysis rates came well within permissible limits (<5%) at a certain concentrations under the tested conditions. Further study was observed by a transmission electron microscope(TEM), it included the method of particles entering the cells, the distribution in cells and the changes of cellular structures.ResultsMagnetic nanoparticles size of 20 nm, uniform shape, good dispersion were prepared. Grafting ratio of GLA and CSO on the surface of Fe3O4 were 20.03% and 15.40% respectively.The experiment of BSA protein adsorption showed that:(1) GLA-Fe3O4 and CSO-Fe3O4 interaction with BSA after 60 min reached the highest adsorption capacity, (2) changes in the initial concentration of BSA can affect the adsorption amount. When the initial concentration of 0.6 mg/mL, the optimal adsorption effect can be reached. (3) changes in pH, BSA adsorption amount of nanoparticles was also changed. When the pH is less than the isoelectric point of BSA, the amount of GLA-Fe3O4 adsorbing BSA is higher than CSO-Fe3O4. When the pH is greater than the isoelectric point of BSA, the results were reversed.MTT results showed that within 24 h GLA-Fe3O4 and CSO-Fe3O4 were not toxicity on RAW264.7 macrophages cells. Hemolysis test of GLA-Fe3O4 and CSO-Fe3O4 was negative. Two kinds of nanoparticles were uptake by RAW264.7 macrophage cells. Morphology of cells and internal structure were no obvious abnormalities.Conclusion1. High dispersion and biological affinity GLA-Fe3O4 and CSO-Fe3O4 were prepared. BSA adsorption behavior on nanoparticles had a variety of factors, and the surface of charged nature and pH of the environment were particularly important. BSA adsorption capacity of GLA-Fe3O4 and CSO-Fe3O4 was different on both sides of pI.2. This suggests the electrostatic force played an important role on the BSA adsorption behavior. MTT results showed that within 24 h GLA-Fe3O4 and CSO-Fe3O4 were not toxicity on RAW264.7 macrophages cells.3. Hemolysis test of GLA-Fe3O4 and CSO-Fe3O4 was negative. So GLA-Fe3O4 and CSO-Fe3O4 can be safely used in biological and medical other fields. |
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