ABSTRACT
Objective To evaluate the inhibitory activity of Herba Taraxaci extract on Escherichia coli DH5α (E. coli DH5α) and to investigate proteomic response of E. coli. Methods Medicinal powder of Herba Taraxaci was extracted with the solvents of different polarity ( n-hexane, ethyl acetate, distilled water) , and then the obtained 8 different extracts were subjected to thin layer chromatography ( TLC) analysis. Microdilution method was performed to detect the minimum inhibitory concentration ( MIC) of different extracts and the growth curves were described. The protein expression profiles of E . coli treated with the extracts were analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis ( SDS-PAGE) and two dimensional electrophoresis (2-DE) . Results Water decoction of Herba Taraxaci could obviously suppress the growth of E. coli with a MIC of 1.95 mg/mL. The different extractions exhibited no antibacterial activity except ethyl acetate phase 3 with a MIC of 0.13 mg/mL, which was equal to 19.23 mg/mL of crude drugs. The results of TLC analysis showed that chlorogenic acid was undetectable in n-hexane extract and ethyl acetate phase 1 extract, and ethyl acetate phase 2 and 3 extracts showed obviously increased spots. The results of SDS-PAGE and 2-DE showed that water decoction of Herba Taraxaci had inhibitory effect on the expression of functional protein. The results of 2-DE showed that after treatment with ethyl acetate phase 3 at the concentration of 2 × MIC for 21 hours, the amount of protein spots were 92 less than those of the blank control group, the spots of E. coli DH5α soluble protein with expression amount down-regulated doubly were 24, and those with expression amount up-regulated doubly were 19. Ethyl acetate phase 3 extract had an effect on down-regulating the protein expression of E. coli DH5α soluble protein pH3-10, and water decoction of Herba Taraxaci had inhibitory effect on E. coli DH5αprotein expression. Conclusion Herba Taraxaci has significant antibacterial activity on E. coli DH5α, and the water-soluble fraction of chlorogenic acid and caffeic acid might be the active components. The possible antibacterial mechanism may be related with the regulation of bacterial protein expression.
ABSTRACT
【Objective】In-vitro antibacterial activity of the combination of Herba Violae(HV)and Herba Taraxaci(HT)in different proportions was compared and analyzed for the optimization of the combination proportion of Herba Violae and Herba Taraxaci and the influencing factors.【Methods】The minimum inhibitory concentration(MIC)of the combination of HV and HT was determined by agar double dilution method and the minimum bactericidal concentration(MBC)by broth dilution method,and bactericidal curve was traced.Meanwhile,the resistance of the combination of HV and HT induced by sub-inhibitory concentration was analyzed,and the influencing factors of in-vitro antibacterial activity were investigated.【Results】The combination of HV and HT in the proportion of 1∶4 had a better antibacterial activity on four kinds of gram-positive bacteria and on four kinds of gramnegative bacteria,and exerted an bactericidal effect on Escherichia coli and Klebsiella Pneumoniae(both MBC_(50)/MIC_(50) and MBC_(90)/MIC_(90) being 2).The bactericidal curve showed that 2?Log cfu/mL bacteria strain could be killed by the combination of the proportion being 1∶4 at 8~(th) hour,the effect being superior to that of other two combinations.Inoculation dosage,serum amount and pH value had no effect on in-vitro antibacterial activity.【Conclusion】The combination of HV and HT in the proportion of 1∶4 has a better antibacterial activity.