Method and composition for inhibiting formation of biofilm
A biofilm and composition technology, applied in the biological field, can solve the problems of high cost, unclear inhibition mechanism, and unsatisfactory effect.
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[0103] Preparation of serum bottles with different oxygen partial pressures: Fill the serum bottle with sterilized water, and insert a needle into the rubber stopper to remove air bubbles. Insert two needles on the rubber stopper, one is connected to the drain pipe, the other is connected to the air pipe, and the water in the serum bottle is filled with high-purity nitrogen (99.99% N 2 ) replacement, continue to ventilate and exhaust for 3 minutes, so that the pressure in the bottle is balanced with the atmospheric pressure. According to the calculation based on the oxygen concentration in the air of 20.8%, when the final oxygen concentration is 0.1%, inject 0.29mL of air, and draw out an equal volume of nitrogen from the bottle according to the calculated volume before pumping the air, and so on to prepare different oxygen partial pressures gas bottle.
[0104] The TBST solution was prepared using 20 mM Tris, 137 mM NaCl, 0.1% Tween 20, and its pH was finally adjusted to 7.6...
Embodiment 1
[0116] Embodiment 1: the preparation of coumarin-chalcone complex (C1-12)
[0117] Coumarin-chalcone complexes (C1-C12) were synthesized by Claisen-Schmidt condensation reaction. Take complex C9 as an example. Add equivalents of 3-acetylcoumarin (0.01 mol) and substituted benzaldehyde (0.01 mol) into a 50 mL single-neck round bottom flask, add 10 mL of ethanol to the reaction mixture and stir for 5 min. Then 2~3 drops of piperidine were added to the mixture and refluxed for 12~15h. The progress of the reaction was checked by thin layer chromatography (TLC). In this example, the progress of all reactions was monitored on pre-coated silica gel plates (Merck, with fluorescent indicator UV254) using a hexane:ethyl acetate (70:30) system as the mobile phase by ultraviolet (254 nm) irradiation Visualize the blobs. After the reaction was complete, the reaction mixture was quenched in crushed ice; the solid product was filtered, washed with water, and recrystallized from ethanol. ...
Embodiment 2
[0118] Example 2: Biofilm assay
[0119] Inoculate 100 μL of Jensen liquid medium with 1% inoculum of the overnight cultured bacterial solution, and at the same time, add C1-C12 and no substances, and culture overnight at 30° C. in a 96-well PVC plate (costar).
[0120] The biomass of bacteria adsorbed on the 96-well plate was detected by the crystal violet method, and the steps were as follows: (1) The free and loosely adsorbed bacteria were sucked away with a row gun, and washed gently in normal saline for 3 times. (2) Add 120 μL of 0.1% crystal violet solution to stain the firmly adsorbed bacterial cells in the small wells, and let stand at room temperature for 30 minutes. (3) Take out the PVC board, suck away the crystal violet with a row gun, and wash off the unadsorbed crystal violet with normal saline. (4) Dissolve the crystal violet adsorbed on the PVC plate with 200 μL of 30% acetic acid solution, and measure the OD value of the acetic acid solution at 560 nm with a ...
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