ABSTRACT
Objective To study the antibacterial mechanisms of ethyl acetate extract (B06e) from the fermentation liquid of an endophytic fungus Alternaria spp. Alternaria spp isolated from the medicinal plant Humata tyermanni. Methods Double dilution method was adopted to measure the minimum inhibitory concentration (MIC) of B06e against Escherchia coli. Then, the changes of electric conductivity of bacterial culture, nucleic acid and protein concentration before and after treated by B06e were analysed, respectively. Besides, flow cytometry, scanning electron microscopy, gel retardation experiments, circular dichroism spectrum and Real-time quantitative PCR were introduced to study the antimicrobial mechanisms of B06e against E. coli. Results The results showed that MIC value of B06e against E. coli was 25 μg/mL. The electric conductivity of 3 × MIC treatment group was 1.01 times the value of the control group. The β-galactosidase activity of 3 × MIC treatment group was 11.6 times more than the value of the control group. Flow cytometry analysis showed that PI positive cells ratio of 3 × MIC treatment group was 286.5 times the value of the control group. Scanning electron microscopy showed that cell surface becomes rough after the treatment of B06e, a large number of cell membrane collapse. These results suggested that B06e can increase the permeability of cell membrane, destroy the integrity of cell membrane. The results of gel retardation experiments and circular dichroism spectrum applied that B06e can be inserted into DNA structure at particular position, however, can not cause DNA degradation. Real-time quantitative PCR results showed that the expressions of recA and recN genes were both up-regulated with the values of 2.9 and 3.7 times the value of the control group, respectively. This result suggested that B06e can destroy the DNA structure, which force E. coli to initiate SOS repair. Conclusion B06e can kill E. coli cell by destroying the cell membrane and damaging DNA structure.
ABSTRACT
Objective: Huperzia serrata, whose growth is limited by high temperature, is a rare medicinal plant with the treatment function for Alzheimer's disease (AD). To research the effect of high temperature on the structure and function of cell membrane and chloroplast, and to provide the evidence for production practices. Methods: H. serrata was processed at 25, 30, 35, and 40°C, respectively, then the content changes of malondial dehyde (MDA) and conductivity rate, and the content changes of total chlorophyll, chlorophyll a, chlorophyll b, and chlorophyll a/b values were measured. The changes of the chloroplast ultra microstructure were observed under the transmission electron microscope (TEM). Results: The changes of MDA and conductivity rate in the process at 35 and 40°C were significantly higher than those of the control group; After processed at 40°C for 4 d, the total chlorophyll was decreased significantly, and became the lowest on the day 6, just was 58% compared to the control group; the change trends to the contents of chlorophyll a, chlorophyll b, and total chlorophyll were similar; TEM observation revealed that after processed at 35°C for 4 d, the chloroplast structure appeared deformation, and after processed at 40°C for 4 d, the chloroplast structure subjected obvious destruction capsule fuzzy, fracture in different degrees, thylakoid in disorder, matrix lamellar irregular, and so on. Conclusion: According to the changes of physiological index, ultramicroscopic structure, and external morphology of chloroplast, the suitable temperature for H. serrata is 25-30°C, 40°C is the limited temperature, causing death after 4 d stress, and 35°C has obvious impact on the growth, long-time stress in 35°C could also cause plant deaths.