The methodology of determining antibacterial activity of plant extracts with disk-diffusion method in vitro

During the process of developing herbal drugs with antimicrobial action, one of the most important phases is testing of antimicrobial activity in vitro. The disk-diffusion method is performed in Petri dish, on solid feeding surface. The disks with definite concentrations of antibiotics, pure substances or plant extracts are placed on the top of feeding plates previously inoculated with pure bacterial culture. Growth of the bacterial culture depends on its susceptibility to a tested substance; if the substance has antibacterial effect, a clear zone free of bacteria will form around the disk. Minimal inhibitory concentration (MIC) of the tested substance is determined by extrapolation of the regression line: concentration of the tested substance/radius or surface of area where growth of bacteria was inhibited. This method of testing microbial susceptibility to antibiotics, pure substances or plant extracts is highly sensitive and specific.