In vitro efficacy of bioactive extracts of 15 medicinal plants against ESbetaL-producing multidrug-resistant enteric bacteria

Alcoholic crude extracts and some fractions from 15 traditionally used Indian medicinal plants were investigated for their ability to inhibit the growth of extended spectrum beta-lactamases (ESbetaL)-producing multidrug-resistant enteric bacteria. The test bacteria Eschrichia coli and Shigella were resistant to 16-23 antibiotics with intermediate or resistance to beta-lactams (minimum inhibitory concentration (MIC) value range 16-1024 microg/ml). The crude plant extracts demonstrated zone of inhibition in the range of 11-29 mm against one or more test bacteria. On the basis of promising activity, 12 plants were selected to determine their efficacy in terms of MIC, which ranged from 0.64 mg/ml to 10.24 mg/ml. The extracts of Acorus calamus, Hemidesmus indicus, Holarrhena antidysenterica and Plumbago zeylanica demonstrated relatively high activity as compared to other plant extracts and were fractionated into acetone, ethyl acetate and methanol. Acetone fraction in most of the cases exhibited higher potency (low MIC value) as compared to ethyl acetate and methanol fraction. However, in Plumbago zeylanica, ethyl acetate fraction was most active. Synergistic interactions among crude extracts were demonstrated in the 12 different combinations against ESbetaL-producing E. coli (ESbetaL-02). Certain combinations exhibited significant synergy with enlargement of combined inhibition zone size by 5 mm. Interaction of crude extracts with five antibiotics (Tetracycline, ciprofloxacin, nalidixic acid, chloramphenicol and streptomycin) demonstrated synergistic interaction with tetracycline and ciprofloxacin by 10 and 3 plant extracts respectively. Phytochemical analysis and thin layer chromatography (TLC) bioautography of crude extracts showed the presence of alkaloids, phenols and flavonoids as active phytoconstituents. Most active fractions of four plants were subjected to Infrared spectroscopy and the major groups of compounds were detected. The plant extracts were further tested for their in vitro haemolytic activity to sheep erythrocytes and demonstrated no haemolysis at recommended doses. Further activity-guided fractionation of active fractions is needed to isolate and characterize the active principle in order to establish the mode of action against the ESbetaL-producing multidrug-resistant enteric bacteria and the mechanism of synergy.     

Antibacterial and cytotoxic activities of extracts from (Tunisian)Rhamnus alaternus (Rhamnaceae)

The petroleum ether, chloroformic, ethyl acetate, methanolic, Total Oligomers Flavonoids (TOF) enriched extracts, water extract as well as its fractions A₁, A₂, A₃ obtained from aerial parts ofRhamnus alaternus, a Tunisian-Mediterranean medicinal species, were investigated for the contents of phenolic compounds, cytotoxic activity against the K562 human chronic myelogenous leukaemia cell line and L1210 leukaemia murine cells and for antibacterial activity against Gram positive and Gram negative bacterial reference strains. A pronounced cytotoxic effect on both the cell lines was shown in the TOF, ethyl acetate, methanolic, aqueous extracts and A₂ fraction, with respectively IC₅₀ values 75, 232, 298, 606 and 571 μg/ml on K562 cells and 198, 176, 767, 560 and 614 μg/ml on L1210 cell line. Significant activity against bacterial reference strains:Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Salmonella enteritidis andSalmonella typhimurium was shown with ethyl acetate, TOF extracts and A2 fraction. The antimicrobial and cytotoxic activities showed byR. alatemus depended on the chemical composition of the tested extracts.     

In vitro antibacterial activity of Psidium guajava Linn. leaf extract on clinical isolates of multidrug resistant Staphylococcus aureus

In the present study, antibacterial activity of aqueous and organic extracts of Psidium guajava leaves was evaluated against multidrug resistant (MDR) clinical isolates of Staphylococcus aureus strains collected from hospitals in northern (Malabar region) Kerala. The strains which exhibited resistance against all the antibiotics tested was selected for antibacterial assays. Minimum inhibitory concentration (MIC) for methanolic and aqueous extracts was found to be 625 ug/ml and 7.5 mg/ml, respectively. Minimum bactericidal concentration (MBC) recorded for methanolic and aqueous extracts was 1.25 and 12.5 mg/ml, respectively. Methanolic extract at minimum bactericidal concentration inhibited the growth of MDR strain by 80%. Time-kill assay revealed that methanolic extract (4 mg/ml) killed MDR bacteria within 10 hr. Total polypeptide profiling of bacterial cultures by SDS-PAGE indicated a high degree of protein degradative activity of the extract. Finally, a human RBC based haemolytic assay showed absence of haemolysis even at concentrations higher than that of MBC, advocating thereby its safety in therapeutic use.     

In vitro evaluation of antibacterial, antioxidant, cytotoxic and apoptotic activities of the tubers infusion and extracts of Cyperus rotundus

The in vitro antibacterial, antioxidant, cytotoxic and apoptotic activities from tubers extracts of Cyperus rotundus (Cyperaceae) were investigated. Antibacterial activity of different extracts was evaluated against five bacterial reference strains. A marked inhibitory effect was observed against Salmonella enteritidis, Staphylococcus aureus and Enterococcus faecalis with total oligomers flavonoids (TOFs) and ethyl acetate extracts. In addition to their antibacterial activity, the same extracts showed a significant ability to inhibit nitroblue tetrazolium reduction by the superoxide radical in a non-enzymatic superoxide generating system. Apoptosis, a highly organized physiological mechanism to eliminate injured or abnormal cells, is also implicated in multistage carcinogenesis. It was observed that TOF and ethyl acetate extracts suppressed growth and proliferation of L1210 cells derived from murine lymphoblastic leukaemia. Morphological features of treated cells and characteristic DNA fragmentation revealed that the cytotoxicity was due to induction of apoptosis. This study confirms that TOF and ethyl acetate extracts of C.rotundus possess antibacterial and antioxidant properties and provoke DNA fragmentation, a sign of induction of apoptosis. These results were correlated with chemical composition of the tested extracts.     

紫穗槐种子提取物的抑菌活性研究

目的研究紫穗槐种子提取物的抑菌活性。方法将紫穗槐种子乙醇提取物分别通过石油醚、乙酸乙酯和正丁醇萃取,选择金黄色葡萄球菌和肺炎克雷伯杆菌为供试菌,采用试管二倍稀释法测定紫穗槐种子提取物的最小抑菌浓度（MIC）和最小杀菌浓度（MBC）,涂布平板法绘制杀菌曲线,电镜下观察药物对细菌超微结构的影响。结果紫穗槐种子提取物经乙酸乙酯萃取后对供试菌抑制作用较强,其中对金黄色葡萄球菌的MIC和MBC分别为2．5、5．0mg／mL;对肺炎克雷伯杆菌的MIC和MBC分别为5．0、10．0mg／mL;杀菌曲线结果表明,药物对供试菌的抑制作用存在浓度和时间依赖性;电镜结果说明,药物的作用可能与破坏菌体细胞壁、改变细胞膜通透性有关。结论紫穗槐种子提取物具有显著的抗菌活性。     

Anti-inflammatory and enzyme inhibitory activities of a crude extract and a pterocarpan isolated from the aerial parts of Vitex agnus-castus

A new compound, 6a,11a-dihydro-6H-[1] benzofuro [3,2-c][1,3]dioxolo[4,5-g]chromen-9-ol was isolated from the ethyl acetate fraction of Vitex agnus-castus. The structure of this compound was identified with the help of spectroscopic techniques ((13)C NMR, (1)H NMR, HMBC, HMQC, NOESY and COSY). The compound showed low urease- (32.0%) and chymotrypsin- (31.4%) inhibitory activity, and moderate (41.3%) anti-inflammatory activity. The crude extract and various fractions obtained from the aerial parts of the plant were also screened for possible in vitro hemagglutination, antibacterial and phytotoxic activities. No hemagglutination activity against human erythrocytes was observed in crude extracts and fractions of V. agnus-castus. The fractions and crude methanolic extract showed moderate and low antibacterial activity. Exceptions were the CHCl(3) fraction, which showed significant antibacterial activity against Klebsiella pneumonia (81% with MIC(50)=2.19 mg/mL), the n-hexane fraction, which exhibited no activity against Salmonella typhi, and the CHCl(3) and aqueous fractions, which showed no activity against Bacillus pumalis. Moderate phytotoxic activity (62.5%) was observed by n-hexane fraction of V. agnus-castus against Lemna minor L at 1000 μg/mL.     

Evaluation of anti-methicillin-resistant Staphylococcus aureus (MRSA) activity and synergy of some bioactive plant extracts

Anti-methicillin-resistant Staphylococcus aureus (MRSA) activity of ethanolic extracts of four medicinal plants namely Acorus calamus (rhizome) Hemidesmus indicus (stem), Holarrhena antidysenterica (bark), and Plumbago zeylanica (root), were detected with inhibition zone size ranged from 11 to 44 mm and minimum inhibitory concentration (MIC) varied from 0.32 to 3.25 mg/mL. Further, ethyl acetate, acetone and methanol fractions of above plants demonstrated antibacterial activity. The potency of these fractions based on zone of inhibition and MIC value was relatively higher in P. zeylanica (ethylacetate fraction), followed by acetone fractions of H. indicus, A. calamus, and H. antidysenterica. Time kill assay with most promising fractions of these plant extracts, demonstrated concentration-dependent killing of MRSA within 9-12 h of incubation. Interestingly, synergistic interaction among alcoholic extracts and some fractions of above four plants was evident against MRSA. Further, synergistic interaction of these extracts was detected with one or more antibiotics tested (tetracycline, chloramphenicol, ciprofloxacin, cefuroxime and ceftidizime). The findings also validate the traditional uses of above plants against infectious diseases. Phytochemical studies demonstrated flavonoids and phenols as major active constituents. Further investigations are needed to characterize the active principle and its interaction mechanism with antibiotics.     

Antibacterial activity of marine-derived fungi

A total of 227 marine isolates of ubiqituous fungi were cultivated on different media and the secondary metabolite content of the extracts (ethyl acetate/chloroform/methanol 3 : 2 : 1) characterized by HPLC. The fungi were secured from animals, plants and sediments of Venezuelan waters (0–10 m) including mangroves and lagoonal areas. The extracts were tested for antibacterial activity. A total of 7 were active towards Vibrio parahaemolyticus and 55 towards Staphylococcus aureus, representing 18 different fungal species from 8 ascomycetous genera. For 61 strains of Penicillium citrinum antibacterial activity correlated well with content of secondary metabolites as measured by HPLC. Thirteen isolates of Penicillium steckii produced very similar profiles of secondary metabolites and 6 of these had activity against either V. parahaemolyticus or S. aureus or both. This revised version was published online in June 2006 with corrections to the Cover Date.     

Biological activities from extracts of endophytic fungi isolated from Viguiera arenaria and Tithonia diversifolia

A total of 39 endophytic fungi have been isolated from Viguiera arenaria and Tithonia diversifolia, both collected in S?o Paulo State, Brazil. The isolates were identified based on their ribosomal DNA sequences. The ethyl acetate (EtOAc) extracts of all endophytic fungi were evaluated for their antimicrobial, antiparasitic and antitumoral activity. Antimicrobial screening was conducted using an agar diffusion assay against three pathogenic microorganisms: Staphylococcus aureus, Escherichia coli and Candida albicans. Antiparasitic activity was determined by enzymatic inhibition of gGAPDH of Trypanosoma cruzi and adenine phosphorybosiltransferase (APRT) of Leishmania tarentolae. Antitumoral activity was tested against human T leukemia cells by the Mosmann colorimetric method. All extracts showed activity in at least one assay: 79.5% of the extracts were cytotoxic against leukemia cells, 5.1% of the extracts were active against S. aureus, 25.6% against E. coli and 64.1% against Candida albicans. Only one extract showed promising results in the inhibition of parasitic enzymes gGAPDH (95.0%) and three were found to inhibit APRT activity. The cytotoxic extract produced by the strain VA1 (Glomerella cingulata) was fractionated and yielded nectriapyrone and tyrosol. Nectriapyrone showed relevant cytotoxic activity against both human T leukemia and melanoma tumor cell lines.     

Diversity and exploration of bioactive marine actinomycetes in the Bay of Bengal of the Puducherry coast of India

The present study was designed to investigate the Puducherry coast of the Bay of Bengal, India for the diversity of bioactive actinomycetes. A total of 50 actinomycete strains were isolated from the marine sediments and most of the strains were belongs to Streptomyces. These strains were identified by means of morphological physiological, biochemical and cultural characteristics. The isolates were subjected to shake flask fermentation and the secondary metabolites were extracted with ethyl acetate and screened for cytotoxicity, hemolytic activity and antimicrobial activity against selected bacterial and fungal pathogens. The cytotoxic activity was evaluated using HeLa cell lines by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a tetrazole (MTT) assay, hemolytic activity on mouse erythrocytes and the antifungal activity was evaluated by MTT cytotoxic assay against Aspergillus niger, Aspergillus fumigatus and Candida albicans. The antibacterial activity was studied against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Klebsiella pneumoniae. The cytotoxicity and antimicrobial activity of secondary metabolite was found to be concentration dependent and nearly 24% of isolates showed significant antimicrobial, hemolytic and cytotoxic activity. The results of our study indicate the diversity and bioactive potential of marine actinomycetes isolated in the Puducherry coast.     

香椿嫩芽和老叶萃取物抗菌活性的比较研究

采用琼脂扩散法对香椿嫩芽和老叶萃取物的氯仿、乙酸乙酯和正丁醇萃取部分的抗菌活性进行了研究.嫩芽萃取物的抗菌活性明显高于老叶萃取物,其中嫩芽萃取物的乙酸乙酯部分表现出最强的抗菌活性,其最小抑菌浓度(MIC)为1.3 mg/mL.实验结果表明,香椿叶中的抑菌物质随着生长时间的延长而逐渐降低.     

Phytochemical, antimicrobial, antioxidant and antigenotoxic potentials of Cyperus rotundus extracts

The aqueous, ethyl acetate, methanolic and Total Oligomer Flavonoids (TOF) enriched extracts, obtained from the aerial parts of Cyperus rotundus, were investigated for their contents in phenolic compounds. Antioxidative activity using the NBT/riboflavin assay system, antimicrobial activity against Gram positive and Gram negative bacterial reference strains as well as antigenotoxic activity tested with the SOS chromotest assay were also studied. Significant antibacterial activity against reference strains; Staphylococcus aureus, Enterococcus faecalis, Salmonella enteritidis and Salmonella typhimurium, was detected in the presence of ethyl acetate and TOF enriched extracts. In addition to their antimicrobial activity, the same extracts showed a significant ability to inhibit nitroblue tetrazolium reduction by the superoxide radical in a non enzymatic O₂^.− generating system, and were also able to reduce significantly the genotoxicity induced by nifuroxazide and Aflatoxin B1. The antioxidant, antimicrobial and antigenotoxic activities exhibited by C. rotundus depend on the chemical composition of the tested extracts.     

Comparative activity against pathogenic bacteria of the root,stem, and leaf of <Emphasis Type="Italic">Raphanus sativus</Emphasis> grown in India

Aqueous, methanol, ethyl acetate, and chloroform extracts of the root, stem, and leaf of Raphanus sativus were studied for antibacterial activity against food-borne and resistant pathogens. All extracts except the aqueous extracts had significant broad-spectrum inhibitory activity. The ethyl acetate extract of the root had the potent antibacterial activity, with a minimum inhibitory concentration (MIC) of 0.016–0.064 mg/ml and a minimum bactericidal concentration (MBC) of 0.016–0.512 mg/ml against health-damaging bacteria. This was followed by the ethyl acetate extracts of the leaf and stem with MICs of 0.064–0.256 and 0.128–0.256 mg/ml, respectively and MBCs of 0.128–2.05 and 0.256–2.05 mg/ml, respectively. The ethyl acetate extracts of the different parts of R. sativus retained their antibacterial activity after heat treatment at 100°C for 30 min, and their antibacterial activity was enhanced when pH was maintained in the acidic range. Hence this study, for the first time, demonstrated that the root, stem, and leaf of R. sativus had significant bactericidal effects against human pathogenic bacteria, justifying their traditional use as anti-infective agents in herbal medicines.     

杨梅果实不同溶剂提取物抑菌特性的研究

目的探讨4种溶剂杨梅果实提取物对14种致病菌的抑菌作用,并对其中活性成分的含量进行测定和比较。方法利用常压浸提法制备杨梅果实的水、乙醇、乙酸乙酯和丙酮提取物;采用管碟法、琼脂平板稀释法测定各种杨梅提取物的抑菌活性,采用常规含量测定方法对各提取物中的主要成分及含量进行研究。结果4种杨梅果实提取物对革兰阴性和阳性菌均表现出了较强的抑菌活性,其中水提取物的抑菌活性最强。含量测定结果表明4种提取物的主要成分中总酚、黄酮含量以及总酸度相差较大,单宁含量差异较小。结论推测提取物中的主要抑菌成分为单宁类物质并确定杨梅果实的最佳提取溶剂为水。     

Antibacterial efficacy of Withania somnifera (ashwagandha) an indigenous medicinal plant against experimental murine salmonellosis

In the present study, we evaluated the antibacterial activity of ashwagandha [Withania somnifera L. Dunal (Solanaceae; root and leaves)], an Indian traditional medicinal plant against pathogenic bacteria. Both aqueous as well as alcoholic extracts of the plant (root as well as leaves) were found to possess strong antibacterial activity against a range of bacteria, as revealed by in vitro Agar Well Diffusion Method. The methanolic extract was further subfractionated using various solvents and the butanolic sub-fraction was found to possess maximum inhibitory activity against a spectrum of bacteria including Salmonella typhimurium. Moreover, in contrast to the synthetic antibiotic (viz. chloramphenicol), these extracts did not induce lysis on incubation with human erythrocytes, advocating their safety to the living cells. Finally, the antibacterial efficacy of the extracts isolated from plant (both root and leaves) was determined against experimental salmonellosis in Balb/C mice. Oral administration of the aqueous extracts successfully obliterated salmonella infection in Balb/C mice as revealed by increased survival rate as well as less bacterial load in various vital organs of the treated animals.     

Antiviral activity of crude extracts of Eugenia jambolana Lam. against highly pathogenic avian influenza (H5N1) virus

Crude extracts of leaves and bark of E. jambolana were tested for antiviral activity against highly pathogenic avian influenza virus (H5N1) by CPE reduction assay in three different layouts to elucidate virucidal, post-exposure and preexposure antiviral activity of the extracts. The cold and hot aqueous extracts of bark and hot aqueous extract of leaves of E. jambolana showed significant virucidal activity (100% inhibition) which was further confirmed in virus yield reduction assay (-98 to 99% reduction) and by egg based in ovo assay. The selective index (CC50/EC50) of hot aqueous extract (248) and cold aqueous extract (43.5) of bark of E. jambolana showed their antiviral potential against H5N1 virus. The significant virucidal activity of leaves and bark of E. jambolana merits further investigation as it may provide alternative antiviral agent for managing avian influenza infections in poultry farms and potential avian-human transmission.     

无叶假木贼和盐爪爪提取物的抗菌活性

无叶假木贼和盐爪爪地上部分乙醇提取物、不同溶剂萃取部分对供试病原细菌和真菌均表现出较好的抑制作用,其乙酸乙酯萃取部分和正丁醇萃取部分的抗菌活性明显强于石油醚萃取部分和水部分。结果表明,无叶假木贼中抗菌活性成分主要为极性中等的化合物,且很可能是具弱碱性、易与酸成盐的生物碱类。盐爪爪中抗菌活性成分同样为极性中等的化合物。     

Antibacterial and antilarval activity of deep-sea bacteria from sediments of the West Pacific Ocean

Deep-sea microorganisms are a new source of bioactive compounds. In this study, crude ethyl acetate extracts of 176 strains of deep-sea bacteria, isolated from sediments of the West Pacific Ocean, were screened for their antibacterial activity against four test bacterial strains isolated from marine biofilms. Of these, 28 deep-sea bacterial strains exhibited antibacterial activity against one or more of the bacteria tested. Active deep-sea bacterial strains belonged mainly to the genera of Pseudomonas, Psychrobacter and Halomonas. Additionally, antilarval activity of 56 deep-sea bacterial strains was screened using Balanus amphitrite larvae. Seven bacterial strains produced metabolites that had strong inhibitive effects on larval settlement. None of these metabolites showed significant toxicity. The crude extract of one deep-sea Streptomyces strain could completely inhibit larval settlement at a concentration of 25 microg ml(-1).     

Antimicrobial activities of extracts of Cyperus rotundus L. rhizomes against some bacterial and fungal pathogens of strawberry and tomato

Antimicrobial activities of rhizome extracts of Cyperus rotundus were investigated on selected plant pathogenic bacteria and fungi. Ethyl acetate and hexane extracts showed antibacterial activity against three isolates of Clavibacter michiganensis subsp. michiganensis at concentrations of 900 and 1000 μg/ml. However, Gram-negative bacterial pathogens of tomato; Pseudomonas syringae pv. tomato and Ralstonia solanacearum were not inhibited from the extracts. Ethyl acetate extracts at 100 μg/ml inhibited mycelial growth and spore germination of the two strawberry isolates of Botrytis cinerea; however, no significant inhibition was found in tomato fungal pathogen, Fusarium oxysporum f. sp. radicis lycopersici. Minimum inhibitory concentrations were determined as 0.0625 and 0.125 mg/ml against C. michiganensis subsp. michiganensis for ethyl acetate and hexane extracts of rhizomes, respectively. This study shows the potentials of extracts of C. rotundus rhizomes as antimicrobial agents that are effective against the tested plant pathogenic bacteria and fungi.     

Antibacterial and antifungal activities of teucrium royleanum (Labiatea)

The crude methanolic extract and subsequent fractions of Teucrium royleanum (Labiatea) were screened for antibacterial and antifungal activities. Against tested pathogens, crude extract and subsequent fractions demonstrated moderate to excellent antibacterial activities. Highest antibacterial activity was displayed by the ethyl acetate fraction against S. typhi (100%), against E.coli (76.7%) and against P. aerugenosa (70.8%) followed by the chloroform fraction against S. typhi (85.7%). Similarly, the crude extract and its subsequent fractions showed mild to excellent activities in the antifungal bioassay with maximum antifungal activity against M. canis (87%) by the chloroform fraction followed by the ethyl acetate (71%) and n-butanol (70%) fractions.