| ObjectiveTo explore the optimal extraction process conditions of Eucalyptus oil with supercritical CO2 extraction apparatus; to investigate the LD50, establish the method of GC-MS to determine the concentration of 1,8-cineole in rats plasma and research pharmacokinetic parameters of 1,8-cineole; to study antibacterial activity in vitro, establish the model of bacteria diarrhea with mice and study the therapeutic effect of eucalyptus oil for bacteria diarrhea in mice. Methods(1) To optimism the extraction process conditions of eucalyptus oil with supercritical CO2 extraction apparatus.After single factor experiment, response surface analysis( Design-Expert software) was used to analysis and optimize the supercritical CO2 extraction conditions such as time, temperature and pressure. Eucalyptus oil yield was chosen as the index to screen out the optimal extraction conditions.(2)Determination of LD50.Kunming mice were chosen as the subject. The method of lavage was adopted for high, medium and low dose of eucalyptus oil. Observe the status of deaths and calculate LD50.(3)Establish the method to detect the content of 1,8-cineole in plasma sample with GC-MS.Liquid-liquid extraction method was used to extract eucalyptus oil ingredient in plasma sample, butyl acetate was chosen as internal standard to establish the method to detect the content of 1,8-cineole in plasma sample by GC-MS.(4)Study on the pharmacokinetic parameters of 1,8-cineole in vivo.Plugging the plasma drug concentration data that had measured into pharmacokinetic software, DAS3.0, to calculate the pharmacokinetic parameters of 1,8-cineole in mice.(5)Study on anti-Escherichia coli effect of eucalyptus oil in vitro.Eucalyptus oil acted on E. coli, and setting negative control group and positive control group. That the clarification dreege of culture medium was the index to detect MIC.(6)Make diarrhea model.Healthy Balb/c mice were chosen as the research object. E. coli was injected into mice by intraperitoneal injection to establish the model of bacterial diarrhea. The diarrhea index was chosen as index, observing the situation of diarrhea within 4h.(7)The therapeutic effect of eucalyptus oil on bacterial diarrhea.Mice were randomly divided into blank group, model group, eucalyptus oil group, amoxicillin group. After successfully making the diarrhea model for 1h, the last three groups were given physiological saline, eucalyptus oil, amoxicillin. Observe the curative effect of eucalyptus oil on diarrhea. Results(1) The optimal extraction technology conditions, extracting eucalyptus oil from eucalyptus leaves, were that extraction temperature was 75.72℃, extraction time was 8h, extraction pressure was 402.78 bar, extraction ratio was 6.83 %.(2) The LD50 of eucalyptus oil was 5046.90 mg/kg. LD50 of 95% confidence limit was 4979.14-5356.68 mg/kg.(3) Results showed that the ratio of concentration and peak area of 1,8-cineole had a good linear relationship within the concentration range of 0.5mg/m L ~ 8mg/m L.(4) Dealing with pharmacokinetic software DAS3.0, it showed that Cmax was 17.46 mg/L,Tmax was 10 min, t1/2 was 155.70 min.(5) Applying eucalyptus oil to the study of antibacterial activity, results showed that MIC of eucalyptus oil to E. coli was 2.08 mg/m L.(6) E. coli was injected into mice by intraperitoneal injection at a dose of 0.3m L/10 g. Within 4h, the mice persisted diarrhea and eat less, the spirit was listless, the fur was loose and bleak. It showed that the model of diarrhea was successful.(7) In the study, we found that eucalyptus oil group, amoxicillin group compared with model group, according to the inspection level(α=0.05), had statistically significance. It showed that eucalyptus oil and amoxicillin had therapeutic effect on diarrhea caused by E. coli. Eucalyptus oil group compared with amoxicillin group, according to the inspection level(α=0.05), P(P = 0.662) > 0.05 showed that there hadn’t been statistically significance. In the treatment of diarrhea caused by E. coli, there was no significant difference between eucalyptus oil and amoxicillin. Conclusion(1) Compared with traditional methods(steam distillation), supercritical CO2 extraction method significantly improves the eucalyptus oil extraction rate;(2) Bacteriostasis experiment in vitro proves that the eucalyptus oil has inhibitory effect on E. coli;(3) Research shows that eucalyptus oil has certain effects on E. coli in vivo and in vitro. |
PDF Full Text Request