Study On The Antibacterial Activities Against Putrefactive Bateria In Meats And The Effective Components Of Herbal Medicine In Zhejiang

According to statistics, about 20% of meat and meat products were polluted by food spoilage every year in the world, which not only causes great economic losses, but also is extremely detrimental to human health. Previous studies showed that a major of food deterioration was caused by microbial contamination. In this work, we investigated the antibacterial activities of ethanol extracts from six kinds of herbal medicines in Zhejiang including Euphorbia humifusa Willd, Scrophularia ningpoensis Hemsl., Atractylodes macracephal Koidz, Paeonia lactiflora Pall, Curcuma wenyujin Y.H.Chen et C.Ling, Fritillaria thunbergiiMiq. The results showed that Paeonia lactiflora Pall has significant inhibitory effects on the all tested microorganism, and was, therefore, chosen to track the antibacterial activity and purified the active ingredients by modern chromatographic techniques. The structure of purified compounds was analyzed and identified. Meanwhile, the antibacterial activities of the purified compounds were evaluated. The main results are shown as follows:(1) The results of antibacterial screening showed that the ethanol extracts of Paeonia lactiflora Pall and Euphorbia humifusa had strong inhibitory effects on Staphylococcus aureus, Micrococcus luteus, Escherichia coli., Enterobacter aerogenes, Staphylococcus saprophyticus, Macrococcus caseolyticus,Pseudomonas fluorescens, Proteusbacillus vulgaris, Bacillus subtilis, Monilia albican.; the ethanol extracts of Atractylodes macracephal Koidz had strong inhibitory effects on Bacillus subtilis, Staphylococcus aureus, Monilia albican, Pseudomonas fluorescens; the ethanol extracts of Curcuma wenyujin Y.H.Chen et C.Ling, had strong inhibitory effects on Escherichia coli., Enterobacter aerogenes, Staphylococcus aureus, Micrococcus luteus, Pseudomonas fluorescens; the ethanol extracts of Scrophularia ningpoensis Hemsl. had certain inhibition on Staphylococcus aureus.(2) The MIC value of Paeonia lactiflora Pall extracts to the all tested microorganism were further examined. The results showed that the ethanol extracts of Paeonia lactiflora Pall had strong inhibitory effects on Staphylococcus aureus, Micrococcus luteus; the MIC of Paeonia lactiflora Pall extract to Staphylococcus aureus and Micrococcus luteus were 12.5 mg/mL respectively; while the MIC of the extract to Escherichia coli., Enterobacter aerogenes, Staphylococcus saprophyticus, Macrococcus caseolyticus and Pseudomonas fluorescens were 25 mg/mL; the MIC of the extract to Proteusbacillus vulgaris, Bacillus subtilis and Monilia albican were 50 mg/mL.(3) The effects of heat treatment and UV irradiation on antibacterial activity of Paeonia lactiflora Pall extract were examined. The result showed that the antibacterial activity to Staphylococcus aureus was stable to high temperature and UV irradiation.(4) The inhibition of petroleum ether, ethyl acetate, n-butanol and water phases of Paeonia lactiflora Pall extract on Staphylococcus aureus and Micrococcus luteus was further examined, and the results showed that the ethyl acetate phases had strong inhibition on the two tested microorganisms. The MIC value of Paeonia lactiflora Pall ethyl acetate phases to Staphylococcus aureus and Micrococcus luteus were 12.5 mg/mL, which were similar to Paeonia lactiflora Pall ethanol extract; the MIC values of Paeonia lactiflora Pall n-butanol phases were 50 mg/mL and 100 mg/mL to Staphylococcus aureus and Micrococcus luteus respectively, which were higher than Paeonia lactiflora Pall extract. While the petroleum ether phases and water phases almost had no antibacterial activity to Staphylococcus aureus and Micrococcus luteus. The result showed that the main antimicrobial active substances of Paeonia lactiflora Pall extract were in ethyl acetate phases.(5) The chemical constituent in the petroleum ether extract of Paeonia lactiflora Pall was separated and purified by modern chromatographic techniques including silica gel, silica gel thin-layer, Sephadex LH-20,and HPLC. Six compounds were purified and eventually identified as 4′-O-benzoylpaeoniflorin(CompoundⅠ),(+)-catechin(CompoundⅡ),(-)-catechin(CompoundⅢ), ethyl gallate(CompoundⅣ), 4-methoxy-2′,4′-dihydroxychalcone(CompoundⅤ), β-Sitosterol(CompoundⅥ) in order.(6) The inhibitory effects of the purified compounds on the bacteria were examined by MTT Colorimetric Assay, and the results indicated that all of the compounds had the antibacterial activity to Staphylococcus aureus and Micrococcus luteus, and CompoundⅠ showed the highest inhibition on both bacteria. Furthermore, we tested the antibacterial activities of CompoundⅠ with different concentrations. Strong inhibitory effects on Staphylococcus aureus and Micrococcus luteus were obtained when the concentration is more than 100 μg/mL. But there was little inhibitory effects when the concentration was less than 12.5 μg/mL. The median effective inhibitory concentrations on Staphylococcus aureus and Micrococcus luteus were 47.81 μg/mL and 30.99 μg/mL respectively by fitting linear equation.To sum up, the results of this work indicated that Paeonia lactiflora Pall could be used as a kind of natural source of bacteriostatic agent with high antibacterial activities, and the main antimicrobial active substances were in ethyl acetate phases. Moreover, we tracked out that CompoundⅠ might be one of main inhibiting activity substances. The results provided a scientific basis for further exploring new natural antiseptic afterwards.