This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, systematic supply, or distribution in any form to anyone is expressly forbidden. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, systematic supply, or distribution in any form to anyone is expressly forbidden. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, systematic supply, or distribution in any form to anyone is expressly forbidden. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources.
Publisher: Taylor & Francis Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK Natural Product Research: Formerly Natural Product Letters Publication details, including instructions for authors and subscription information: http://www.tandfonline.com/loi/gnpl20 Antimicrobial activity and phytochemical analysis of crude extracts and essential oils from medicinal plants N.C.C. Silva a , L. Barbosa b , L.N. Seito c & A. Fernandes Junior a a Department of Microbiology and Immunology, Institute of Biosciences, So Paulo State University, UNESP, Botucatu, So Paulo, Brazil b Department of Biostatistics, Institute of Biosciences, So Paulo State University, UNESP, Botucatu, So Paulo, Brazil c Department of Pharmacology, Institute of Biosciences, So Paulo State University, UNESP, Botucatu, So Paulo, Brazil Available online: 19 Oct 2011 To cite this article: N.C.C. Silva, L. Barbosa, L.N. Seito & A. Fernandes Junior (2011): Antimicrobial activity and phytochemical analysis of crude extracts and essential oils from medicinal plants, Natural Product Research: Formerly Natural Product Letters, DOI:10.1080/14786419.2011.564582 To link to this article: http://dx.doi.org/10.1080/14786419.2011.564582 PLEASE SCROLL DOWN FOR ARTICLE Full terms and conditions of use: http://www.tandfonline.com/page/terms-and- conditions This article may be used for research, teaching, and private study purposes. Any substantial or systematic reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to anyone is expressly forbidden. The publisher does not give any warranty express or implied or make any representation that the contents will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims, proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in connection with or arising out of the use of this material. D o w n l o a d e d
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Natural Product Research 2011, 15, iFirst SHORT COMMUNICATION Antimicrobial activity and phytochemical analysis of crude extracts and essential oils from medicinal plants N.C.C. Silva a , L. Barbosa b , L.N. Seito c and A. Fernandes Junior a * a Department of Microbiology and Immunology, Institute of Biosciences, Sao Paulo State University, UNESP, Botucatu, Sao Paulo, Brazil; b Department of Biostatistics, Institute of Biosciences, Sao Paulo State University, UNESP, Botucatu, Sao Paulo, Brazil; c Department of Pharmacology, Institute of Biosciences, Sao Paulo State University, UNESP, Botucatu, Sao Paulo, Brazil (Received 1 July 2010; final version received 16 February 2011) We aimed to establish a phytochemical analysis of the crude extracts and performed GC-MS of the essential oils (EOs) of Eugenia uniflora L. (Myrtaceae) and Asteraceae species Baccharis dracunculifolia DC, Matricaria chamomilla L. and Vernonia polyanthes Less, as well as determining their antimicrobial activity. Establishment of the minimal inhibitory concentrations of the crude extracts and EOs against 16 Staphylococcus aureus and 16 Escherichia coli strains from human specimens was carried out using the dilution method in MuellerHinton agar. Some phenolic compounds with antimicrobial properties were established, and all EOs had a higher antimicrobial activity than the extracts. Matricaria chamomilla extract and E. uniflora EO were efficient against S. aureus strains, while E. uniflora and V. polyanthes extracts and V. polyanthes EO showed the best antimicrobial activity against E. coli strains. Staphylococcus aureus strains were more susceptible to the tested plant products than E. coli, but all natural products promoted antimicro- bial growth inhibition. Keywords: plant crude extracts; essential oils; medicinal plants; minimal inhibitory concentration; antibacterial activity; phytochemical analysis 1. Introduction Eugenia uniflora L. (Myrtaceae) is used as anti-diarrhoeal, diuretic, anti-rheumatic, anti-febrile and anti-diabetic agents in Brazilian folk medicine and its leaf extracts possess anti-inflammatory (Schapoval, Silveira, Miranda, Alice, & Henriques, 1994) and antimicrobial activities (Oliveira, Lopes, Cabral, & Eberlin, 2006). Terpenoids and flavonoids are the chemical groups most commonly found in the genus Baccharis (Asteraceae) (Verdi, Brighente, & Pizzolatti, 2005) and Baccharis dracunculifolia DC also has phenolic compounds (Funari, Ferro, & Mathor, 2007) with antimicrobial activity (Duarte, Figueira, Pereira, Magalha es, & Delarmelina, 2004; Ferronato, Marchesan, Pezenti, Bednarski, & Onofre, 2007). *Corresponding author. Email: ary@ibb.unesp.br ISSN 14786419 print/ISSN 14786427 online 2011 Taylor & Francis http://dx.doi.org/10.1080/14786419.2011.564582 http://www.tandfonline.com D o w n l o a d e d
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Vernonia polyanthes Less (Asteraceae) are used in folk medicine in Brazil against flu, colds, coughs, bronchitis, bruises, haemorrhoids and infections of the uterus (Corre a, Bertolucci, Pinto, Reis, & Alves, 2004), showing antibacterial activity against mycobacterial strains (Oliveira et al., 2007). Matricaria chamomilla L (Asteraceae) is used as tea against gastrointestinal and nervous diseases (Fragoso, Esparza, Burchiel, Ruiz, & Torres, 2008) as well as antimicrobial activities (Romero et al., 2005). We aimed to establish the antimicrobial activity and phytochemical analysis of crude methanolic extracts and essential oils (EOs) from E. uniflora, B. dracunculifolia, V. polyanthes and M. chamomilla samples against Staphylococcus aureus and Escherichia coli strains isolated from human specimens. 2. Results and discussion Extracts dry weight, EO density and yield, phytochemical analysis of extracts and EO are presented in Table 1. Phenols, flavonoids and fixed strong acids were found in all extracts, which were previously reported for their antimicrobial activity (Burt, 2004). Most compounds identified in the oils were monoterpenes or sesquiterpenes, which may be involved in the EO antimicrobial effect (Cowan, 1999), although other compounds may also have increased their antimicrobial effects, in a synergistic way. The crude extracts showed around 10% of those obtained in the EO samples and were used for processing the minimum inhibitory concentration (MIC) values (mg mL 1 ) in the antibacterial assays. MIC 90% values are presented in Table 2, and found that S. aureus strains were susceptible to both extracts and EOs from all studied plants. The crude extracts from M. chamomilla, followed by V. polyanthes and B. dracunculifolia, were those with the highest activities against S. aureus strains, while in contrast, the EO from E. uniflora showed the highest inhibitory activity against this bacterium, with a value of MIC 90% 2.2 mg mL 1 . On the other hand, E. coli strains were less susceptible to plant products, except to E. uniflora extract, which MIC 90% (2.2 mg. ml 1 ) showed the best inhibitory effect, althought all EOs showed a similar result. Therefore, all plants showed an antimicrobial activity, either as extracts or oils, revealing their potential use as antimicrobial agents. Gonc alves, Alves Filho, and Menezes (2005) verified the antimicrobial activity of hydroalcoholic extract of E. uniflora against E. coli, S. aureus, Staphylococcus coagulase-negative, Streptococcus pyogenes, Proteus mirabilis, Providencia spp and Shigella sonnei. These data corroborate our results since E. uniflora had a potential as an antimicrobial agent among the assayed plants. Auricchio and Bacchi (2003) demonstrated the presence of flavonoids and tannins in E. uniflora crude extract, allowing us to conclude that these compounds are characteristic of this plant species, although further studies should characterise flavonoid and tannin groups. Terpenoids and flavonoids (Verdi et al., 2005) and phenolic compounds (Funari et al., 2007) were found in the genus Baccharis. Ferronato et al. (2007) found that B. dracunculifolia EO showed antimicrobial effect against E. coli, S. aureus and Pseudomonas aeruginosa, while our results showed higher MIC values assaying S. aureus and E. coli strains. Duarte et al. (2004) reported that the MIC for B. dracunculifolia extract against Enterococcus faecium strains was 10-fold higher for 2 N.C.C. Silva et al. D o w n l o a d e d
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the Gram-positive bacteria, although different bacterial strains were studied. The differences between the MICs may be due to the chemical composition and edaphoclimatic conditions (harvest season, time, location, among others), changing secondary compounds (Gobbo-Neto and Lopes, 2007). Souza, Sena, Maranho, Oliveira, and Guimara es (2008) found compounds such as fixed acids, alkaloids, coumarins, flavonic and saponin glycosides and the presence of alkaloids, coumarins Table 1. Physico-chemical characterisation of plant (crude extracts and EOs) antimicrobials. Vegetal source Density (mg mL 1 ) (EOs) Dry weight (mg mL 1 ) (crude extracts) Yield (%) (EOs) Phytochemical analysis of crude extracts GC-MS of EOs E. uniflora 924.0 145.0 0.19 Phenols, tannins, chalcones, aurones, fla- vones, cate- chins, flavo- noids, saponins, fixed strong acids, quater- nary com- pounds, free steroids and quinones Selina 1,3,7(11) trien-8-one (30.1%), Selina 1,3,7(11) trien- 8-one-epoxide (21.89%), cariofilene (6.51%) V. polyanthes 856.0 62.5 0.15 Phenols, tannins, chalcones, aur- ones, flavo- noids, fixed strong acids, saponins, free steroids, qui- nones and flavananois Germacrene D (27.79%), "-Cariofilene (16.2%), Germacrene B (15.01%) B. dracunculifolia 857.0 76.0 0.20 Phenols, tannins, flavones, cate- chins, flavo- noids, saponins, fixed strong acids, quater- nary bases and xanthones Nerolidol (18.77%), Germacrene D (10.45%), limo- neme (8.75%) M. chamomilla 940.0 100.0 0.17 Phenols, flavones, flavonoids, fixed strong acids, quater- nary com- pounds, qui- nones, xan- thones, free triterpenes Chamazulene (31.48%) -bisabolol and Bisabolone oxide (15.71%) Note: GC-MS Gas chromatography-mass spectrometry. Natural Product Research 3 D o w n l o a d e d
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and flavonoids in V. polyanthes samples. Oliveira et al. (2007) reported the anti-mycobacterial activity with V. polyanthes, crude extracts as well as viable count of inhibited cells after 3 h of contact in the time kill curve assay. Despite different methodologies performed, our results were also satisfactory with V. polyanthes crude extract. Matricaria chamomilla showed antibacterial activity against S. aureus and phenolic compounds were responsible for its antimicrobial properties (Romero et al., 2005). Nogueira, Diniz, and Lima (2008) found no inhibition against P. aeruginosa growth with chamomile EO, but 4% EO inhibited S. aureus and Candida albicans strains, with inhibition zones ranging between 10 and 12 mm. Thus, M. chamomilla extract and EO showed important inhibitory action against S. aureus strains, and phenolic compounds were found in the phytochemical analysis. However, we believe that other compounds are involved with the antimicrobial property together with the phenols. 3. Conclusion The results showed the potential use of these plants as antimicrobial agents and S. aureus strains were more susceptible to plant products than E. coli and all natural products promoted antimicrobial growth inhibition. Further studies should inves- tigate the toxicity and chemical characterisation, aiming its use in the treatment of infectious diseases, either alone or as an adjuvant with conventional antimicrobial drugs. Supplementary material Experimental details relating to this article are available online. Acknowledgements The authors thank Dr Margarida Saeki and Dr Julio Doyama for the chromatographic analysis of essential oils, and Dr Jose Mauricio Sforcin for the critical review of the manuscript. Table 2. Minimal inhibitory concentration (mg mL 1 ) values for 90% (MIC 90% ) of S. aureus and E. coli strains according to the crude extracts and EOs. S. aureus E. coli Vegetal source Crude extracts Essential oils Crude extracts Essential oils B. dracunculifolia 5.4 a 3.7 ac 32.4 ab 25.8 a V. polyanthes 3.3 ab 2.8 a 26.9 ac 24.1 a M. chamomilla 1.2 b 2.9 bc 43.4 b 28.2 b E. uniflora 24.1 c 2.2 b 15.9 c 27.6 c Note: Different letters in columns represent significant differences in antimicrobial activity between products when p 0.05. 4 N.C.C. Silva et al. D o w n l o a d e d
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