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Catechins are a class of polyphenols and have high anti-bacterial activity against various microorganisms. Here, we report the mechanism for antibacterial activity of epigallocatechin gallate (EGCg) against Gram-positive bacteria Bacillus subtilis, which is highly sensitive to EGCg. Transmission electron microscope analysis revealed that deposits containing EGCg were found throughout the cell envelope from the outermost surface to the outer surface of cytoplasmic membrane. Aggregating forms of proteins and EGCg were identified as spots that disappeared or showed markedly decreased intensity after the treatment with EGCg compared to the control by two-dimensional electrophoresis. Among the identified proteins included 4 cell surface proteins, such as oligopeptide ABC transporter binding lipoprotein, glucose phosphotransferase system transporter protein, phosphate ABC transporter substrate-binding protein, and penicillin-binding protein 5. Observations of glucose uptake of cells and cell shape B. subtilis after the treatment with EGCg suggested that EGCg inhibits the major functions of these proteins, leading to growth inhibition of B. subtilis.

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Keywords: monoglycerol fatty acid ester, sucrose fatty acid ester, antibacterial effect, bacterial spore, vegetative cell

Although most Gram-positive bacteria are sensitive to epigallocatechin gallate (EGCg), some species of lactic acid bacteria (LAB) are highly tolerant. The mechanism of LAB tolerance to the antibacterial action of EGCg was investigated. LAB strains with three different cell wall composition types were used: Lactobacillus plantarum NBRC15891 (meso-DAP-type), Lactobacillus fermentum NBRC15885 (Orn-type), and Lactobacillus delbrueckii NBRC3073 (Lys-type). The minimum inhibitory concentration of EGCg for L. plantarum NBRC15891, L. fermentum NBRC15885, and L. delbrueckii NBRC3073 were >1000, >1000, and 500 µg/mL at pH 6.5, respectively. The cell surface hydrophobicity (CSH), and contents of extracellular polymeric substances (EPS) and teichoic acid of these strains suggested that strains with low CSH and producing greater amounts of EPS are highly resistant to EGCg at pH 6.5. After EGCg treatment, the membrane potential decreased in strains with high susceptibility to EGCg. Our findings suggested that LAB characterized by high EPS level and low CSH are resistant to EGCg at pH 6.5.