| Eupatorium odoratum Linn. has medicinal functions of antibacterial, anti-inflammatory and regulating immunity. Its main active components are the total flavonoids. Escherichia coli is the most common pathogen of diarrhea in livestock with serious drug resistance. In the present study, the total flavonoids were extracted from E. odoratum and the in-vitro antibacterial activity of the flavonoids extract were screened to determine which microbial strains were sensitive to these flavonoids. 90 Kunming mice were randomly assigned into 5 groups, 18 mice in each group as the high-dose group(160 mg/kg), medium-dose group(40 mg/kg), the low-dose group(10 mg/kg), the self-healing group, and the blank control group. The mice were intraperitoneally(ip) injected with E.coli to induce diarrhea in all groups except the blank control group. The mice in high-dose group, middle-dose group and the low-dose group were orally given the flavonoid extract once a day post infection. The mice in self-healing group were treated with physiological saline as the substitute once a day post infection. At the same time, the mice in blank control group were also treated with physiological saline as the substitute. By examining the rate of diarrhea in mice, changes in body weight gain, the number of E.coli in feces and serum Ig M, Ig A, IgG, spleen index, changes in duodenal morphology and structure, and changes in mRNA expression of TGF-β1, the in vivo anti-infectious mechanism of the total flavonoids extract on diarrhea was explored. The results are as follows:1. The inhibition zone diameter of the total flavonoids extract from E. odoratum against E.coli was 11.00 mm, along with the MIC of 0.1250 g/mL and the MBC of 0.2500 g/mL. The inhibition zone diameter against Bacillus subtilis was 13.67 mm, MIC was 0.0625 g/mL and the MBC was 0.1250 g/m L; The inhibition zone diameter against Staphylococcus aureus inhibition zone was 19.50 mm, the MIC was 0.0156 g/mL and MBC was 0.0313 g/mL; The inhibition zone diameter against Salmonella inhibition zone was 13.33 mm, MIC was 0.0313 g/mL and MBC was 0.1250 g/mL.2. The mice with ip injection of 3×108 E. coli were successfully established as the diarrhea model that induced diarrhea in all the animals(100%) and the diarrhea index was 0.99 ± 0.36.3. The mice orally administered with the flavonoid extract showed enhanced body weight gain, reduced rate of diarrhea, mortality rate and fecal coliform count, in addition to enhanced serum Ig A and IgG. The body weight gain of the mice in the high-dose group was the highest and the rate of diarrhea and mortality the lowest. The fecal coliform count was reduced(P<0.01), and serum Ig A(P<0.01) and IgG(P<0.05) were enhanced in the high-dose group.4. The morphological study of the duodenal mucosa showed that duodenum mucosa of mice in the experimental group orally given the total flavonoids extract regenerated faster than in the self-healing group. The best recovery of the duodenal mucosa was observed in mice in the high-dose group, whose duodenal mucosa morphology returned to normal.5. In the early diarrhea mice, compared with the blank control group, the duodenal TGF-β1 mRNA expression levels were significantly increased(P<0.01). The mice orally given the high-doses of the total flavonoids extract performed the best with high TGF-β1 mRNA expression level observed among all of the treatments(P<0.05). In later period of diarrhea, the duodenal TGF-β1 mRNA expression level of the diarrhea mice declined back to normal, but the TGF-β1 mRNA expression level in the mice orally given the high-dose of flavonoids remained significantly higher than those in the blank control group and the self-healing group(P<0.05).In summary, the total flavonoids extract from E. odoratum showed broad-spectrum antimicrobial effect and its antibacterial activity against Escherichia coli reached moderate sensitivity. In the mice orally given the total flavonoids extract, decreased the diarrhea incidence, regulated the structure of intestinal microflora, improved antibody levels, and enhanced expression of TGF-β1 mRNA, promotion of repair of the intestinal mucosa were observed. The addition of 160 mg/kg of the total flavonoids extract was the optimal dosage that decreased the diarrhea rate and enhanced the body weight gain of the mice. The possible pharmaceutical mechanism of the total flavonoids from E. odoratum aganist diarrhea in mice with E.coli infection could be: The total flavonoids extract directly killed or inhibited the E.coli in the intestine of mice. The total flavonoids extract improved immune function and increased serum Ig A and IgG and this probably gave protection from E.coli infection. The total flavonoid extract promoted the expression of TGF-β1 mRNA and accelerated the repair speed of intestinal mucosa. It is concluded that all of the above effects were responsible for the pharmaceutical effect of the flavonoid extract to prevent the diarrhea. |
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