Research On Biosynthesis Silver Nanoparticles Using Trichoderma Hamatum

This experiment reported on isolation and identification of a fungi with ability to synthesis silver nanoparticles by adding Ag+ into soil suspension, which was collected in Maoer Mountain, Northeast Forestry University. We employed morphological, physiological and molecular experiments to identify the fungi and biosynthesized silver nanoparticles using this fungi. UV-vis, TEM-. HRTEM-. XRD and FTIR were used to characterized silver nanoparticles. In addition, we detected antibacterial ability of silver nanoparticles to E.coli and Bacillus subtilis and synthesis mechanism were discussed by analysising proteins in cell-free filtrate using SDS-PAGE.(1) Isolation and identification of fungi with ability to synthesis silver nanoparticlesThis experiment focused on isolation of fungi with ability synthesis silver nanoparticles by adding Ag+ into soil suspension. Morphological, physiological and molecular experiments were employed to identify the fungi. Morphological method showed that the colony was white and medium was yellow on PDA plate. The colony grew slowly with smooth surface. Hypha had branch and aerial hyphae was little. Cell nucleus can not be observed obviously. Molecular experiments showed that ITS sequence was 577 bp with Accession No. KM054532 in Genbank. Phylogenetic tree showed that the fungi exhibited 99.5% similarity with Trichoderma hamatum. On basis of morphological, physiological and molecular experiments, the fungi strain was identified as Trichoderma hamatum and named as NYZJ03. The fungi was preserved in China General Microbiological Culture Collection Center and the number was CGMCC No.8464.(2) Biosynthesis and characterization of silver nanoparticlesThis research reported on biosynthesis silver nanoparticles using Trichoderma hamatum mixed with AgNO3 and silver nanoparticles were characterized by UV-vis analysis, XRD analysis, TEM and FTIR analysis. Thermogravimetric analysis and atomic absorption were used to detected productivity of silver nanoparticle and conversion rate of Ag+. Results indicated that silver nanoparticles were cubic and pure; TEM images suggested that silver nanoparticles are monodisperse with shape in spherical. Most of them were under 9 nm and the average size was 6.69 nm. FTIR spectrum revealed that proteins must be present on the surface of silver nanoparticles. Thermogravimetric analysis showed that the productivity of silver nanoparticles was 67.12% and atomic absorption showed conversion rate of Ag+was 84.41%.(3) Research on antibacterial ability and synthesis mechanismMBC of silver nanoparticle for E.coli was 10 μg/mL and MIC was 7 μg/mL, while for Bacillus subtilis was 5 μg/mL and 4 μg/mL. Fatality rate of E.coli was low at the beginning and increased significantly between 20-40 min, which was 86.70%. At 70 min, fatality rate reached to 99.23% and the bacteria must be basically killed. Fatality rate of Bacillus subtilis was 96.22% at 40 min and 99.84% at 60 min. Fatality rate of Bacillus subtilis was much greater than that of E.coli.After adding AgNO3, secreted proteins decreased significantly, which contained three kinds of proteins with relative molecular mass of 40 kDa,55 kDa and 70 kDa, respectively. Proteins attached to silver nanoparticles can be separated after treated with urea and SDS. The protein was 40 kDa and may be essential for synthesis and stability of silver nanoparticles.