Anti-microoganism Activity And Anti-microbial Mechanism Of Soybean Isoflavone

Soybean isoflavone(SI)is a kind of secondary metabolites during the growth of soybeans, mainly contained in the cotyledon and hypocotyl of soybean seeds, is regarded as one of the crude drugs. SI has many biological activities, many studies have investigated the effect of the antibacterial activities, the antibacterial mechanisms were reported less. The antibacterial mechanisms of SI were researched, for the development of high-performance and low toxicity of antibacterial drugs provide a theoretical basis, it is urgent to find more effective drugs to replace of the traditional ones, which will help to timely redirect drug use so as to diminish the development and spread of resistance. It is meaningful to provide theoretical basis for directional screening nature antibacterial drugs and developing SI as a kind of natural food additives. In this paper, the effect of soybean isoflavone on anti-microoganism is investigated, then the minimum inhibitory concentrations (MIC) experiment was carried out by utilizing 2, 3, 5-triphenyl tetrazolium chloride (TTC). The antibacterial mechanisms of SI were extensively investigated, it was carried out by utilizing scanning and transmission electron microscope(SEM and TEM), and further analyzing the respiratory metabolism and change of SDS-PAGE protein spectrums, considering the inhibition of the activity about DNA topoisomerase, 4'6-diamidino-2-phenylindole (DAPI) staining assay and the inhibition of the activity of DNA topoisomerase. The main results are followed:A: Anti-microoganism activity of soybean isoflavone1. Anti-microoganism effect of SI to different kinds of bacteria and fungiIt is found that soybean isoflavone has antibacterial and anti-fungi activity. It shows obvious inhibition for Staphylococcus aureus, Bacillus cereus, Micrococcus luteus, Bacillus subtilis, Bacillus pumilus, Candida albicans, Listeria monocytogens, Aspergillus oryzae, Absidia. But it has no inhibition for Escherichia coli or Saccharomyces cerevisiae.2. The minimum inhibitory concentrations (MIC) experiment of SIThe growth of Staphylococcus aureus was completely inhibited, when SI up to 0.3mg/mL.So the minimum inhibitory concentration to Staphylococcus aureus is confirmed. B: The antibacterial mechanisms of SI1. Effect of SI on morphology to Staphylococcus aureus The erose structures such as rugae and bubbles were observed on the surface of cells by SEM after 24h.2. Effect of SI on ultra-structure to Staphylococcus aureus Moreover, with the TEM, we detected the shrinkage of cytoplasm, the plasmolysis, and then the breach of wall and membrane along with the outflow of protoplasm in Staphylococcus aureus treated with the SI for 4h, 14h and 24h respectively.3. Effect of SI on membrane permeability Membrane permeability was indicated by changes of conductivity in liquid culture medium. Conductivity in liquid culture medium increased after SI added in medium (1h), the rate of change about conductivity was up to 4.61% after treated with SI, it indicates that some cytoplasm permeate from Staphylococcus aureus. It is speculated that membrane permeability was altered by SI.4. The respiratory inhibition experiment Notably, the respiratory inhibition experiment revealed that the superpose rate of soybean isoflavone to propanedioic acid, lodoacetic acid, trisodium phosphate were 8.6%,16.4% and 22.1% seperately.Because of the lowest superpose rate between SI and propanedioic acid, it indicates that SI mainly inhibits the TCA cycle of bacteria.5. Effect of SI on synthesis of proteinThe SDS-PAGE elucidated that the total expression of proteins were decreased in the cell treated with the SI, and especially the larger proteins were reduced with 90.1% which compared to control group.6. Measurement of quantity of DNA,RNAFluorescence microscopy and fluorescence spectrophotometer results showed that DAPI integrated with Staphylococcus aureus and the quantity of DNA and RNA were reduced by 33.53% and 39.82% respectively after treated with SI for 28h.7. Effect of SI on activity of DNA topoisomeraseSI has significant moderate effect on TOPOâ… and TOPOâ…¡, at the concentration of 6.4mg/mL, SI completely inhibited the pBR322DNA cleavage mediated by TOPOâ… and TOPOâ…¡.As the concentration of SI reached higher than 3.2mg/mL, it had a direct break effect on pBR322DNA.The result was that SI could obviously inhibit the growth of curtain bacteria, the antibacterial mechanisms of SI may include inhibiting of cell wall synthesis and cell membrane damage or inhibiting respiratory metabolism and both the activity of TOPOâ… and TOPOâ…¡, further to inhibit the synthesis of nucleic acid and relative proteins.