Antimicrobial Activity of Caesalpinia Melanadenia (Rose) Standl (Fabaceae)
Total Page：16
File Type：pdf, Size：1020Kb
Int.J.Curr.Microbiol.App.Sci (2013) 2(11): 186-193 ISSN: 2319-7706 Volume 2 Number 11 (2013) pp. 186-193 http://www.ijcmas.com Original Research Article Antimicrobial activity of Caesalpinia melanadenia (Rose) Standl (Fabaceae) Adriana Pichardo1, Ximena Méndez1, Griselda Alvarado1, Pilar Ramirez1, RocioSerrano1, Marisol Avila1, Samuel Meraz1, Julieta Orozco1, Ana García-Bores2, J. Guillermo Avila2 and Tzasna Hernández1* 1Laboratorio de Farmacognosia, UBIPRO, Facultad de Estudios Superiores-Iztacala, Universidad Nacional Autónoma de México, Tlalnepantla 54090, Edo. de México, México 2Laboratorio de Fitoquímica, UBIPRO, Facultad de Estudios Superiores-Iztacala, Universidad Nacional Autónoma de México, Tlalnepantla 54090, Edo.de México, México *Corresponding author A B S T R A C T Infusions of the aerial part of Caesalpinia melanadenia (Fabaceae) are used by the inhabitants of San Rafael Coxcatlán, Puebla (México) for the treatment of K e y w o r d s gastrointestinal, respiratory and skin diseases. The aim of this work was to investigate the antimicrobial activity of the aerial parts of Caesalpinia Caesalpinia melanadenia, validate its use and contribute to the knowledge of medicinal flora melanadenia; from San Rafael municipality. Hexane and methanol partitions were used for Antibacterial; antimicrobial test. Eight Gram positive, nine Gram negative bacteria and nine Antifungal; fungal strains were used in the antimicrobial assay. The methanol partition does not Infectious show any activity while hexane partition showed antibacterial activity against all diseases; the bacterial and fungal strains. The most sensitive strains were E. feacalis (MIC= Infusion. 60 µg/mL), S. pneumoniae (MIC= 60 µg/mL), S. epidermidis (MIC= 250 µg/mL), E. aerogenes (MIC= 250 µg/mL) and C. neoformans (MIC = 125 µg/mL). The present study tends to confirm the use in folk medicine of Caesalpinia melanadenia against infectious diseases. Introduction Caesalpinia melanadenia (Rose) Standl Valley, México (Argueta and Cano, 1994; (Fabaceae) is commonly known as Davila et al., 2002). Ixcanelillo . In México infusions of the aerial part are used in Mexican traditional Some species of the Caesalpinia genus medicine by the inhabitants of the village have been chemically examined and of San Rafael Coxcatlán, Puebla, for the yielded diterpenes, triterpenes, flavanoids, treatment of gastrointestinal, respiratory quinones, and alkaloids; and also and skin diseases. C. melanadenia is a biological activities has been evaluated shrub, endemic to Tehuacán-Cuicatán (antibacterial, antifungal, analgesic, anti- 186 Int.J.Curr.Microbiol.App.Sci (2013) 2(11): 186-193 inflammatory, etc.), including C. ferrea, separating funnel. After solvent-solvent (Carvalho et al., 1996), C.pulcherrima extraction, the methanol phase was (Srinivas et al., 2003; Chakraborthy et al., removed from the hexane phase. Both 2009; Das et al., 2010), C. sappan (Pawar partitions, methanolic and hexanic were et al., 2008; Gan et al., 2010), C. concentrated under low pressure (12.0 g mimosoides (Chanwitheesuk et al., 2005; and 3.5 g respectively) and kept in the Yodsaoue et al., 2011), C. boduc (Ata et dark at 4 °C until tested. al., 2009), C. crista (Das et al., 2010; Santnami and Yadava, 2011), C. sappan Microbial Strains (Gan et al., 2010), and C. digyna (Srinivasan et al., 2010). The following strains of bacteria were used: Enterococcus faecalis ATCC 29212, The genus Caesalpinia (Caesalpiniaceae) Staphylococcus aureus ATCC 12398, S. has more than 500 species, many of which aureus ATCC 29213, Escherichia coli have not yet been investigated for ATCC 25922, Pseudomonas aeruginosa potential pharmacological activity (Zanin ATCC 27853, Salmonella typhy ATCC et al., 2012). C. melanadenia has no 19430. Bacillus subtillis, Staphylococcus chemicals or biological studies. The aim of epidermidis, Enterobacter aerogenes were the study was to investigate the donated by FES-Cuautitlán. B. subtilis, antimicrobial activity of the aerial part C. S. epidermidis, E.aerogenes donated by melanadenia (Fabaceae), validate its use the Clinical Analysis Laboratory of FES- and contribute to the knowledge of Iztacala. Streptococcus pneumoniae, medicinal flora from San Rafael Yersinia enterocolitica were isolated from Coxcatlán, municipality. a clinical case and donated by Hospital Angeles (Metropolitano). Vibrio cholera isolated from a clinical case, Vibrio Materials and Methods cholerae INDRE 206 (isolated from Plant Material polluted water), Vibrio cholerae (clinical strain pertaining to 01 group, Inaba Aerial parts of C. melanadenia were serotype, El Tor biotype, and obtained in November 2011 from San enterotoxin producer). These strains were Rafael Coxcatlán municipality. Dr. Edith maintained at 4 °C in Mueller Hinton agar López Villafranco of the IZTA Herbarium (Bioxon), submitted to sensitivity tests at the Factultad de Estudios Superiores (multidiscs Bigaux) and were subcultured Iztacala authenticated it. A voucher every month. specimen was deposited in the IZTA herbarium (Voucher no. HCM15/2011). The yeasts tested were Candida albicans ATCC 10231, C. albicans ATCC Extract preparation and partitions 14065,C. albicans isolated from a clinical case donated by the Clinical Analysis Aerial parts of C. melanadenia (182 g) Laboratory of FES-Iztacala, C. albicans, were shade-dried at room temperature, C. glabrata, C. tropicalis isolated from a ground into powder and sequentially clinical case and donated by Hospital extracted with methanol. The extract was Angeles (Metropolitano), C. albicans and filtered and successively concentrated. The Criptococcus neoformans donated by methanol extract was redissolved in FES-Cuautitlán. The stock culture was methanol and hexane was added to it in a maintained on Czapek Dox agar (Sigma). 187 Int.J.Curr.Microbiol.App.Sci (2013) 2(11): 186-193 Antibacterial Activity Antifungal Activity The antibacterial activity was measured by Yeast was assayed by the method the disk-diffusion method (Vanden Berghe described for bacteria, using Petri dishes and Vlietinck, 1991). The microorganisms containing CzapekDox Agar (20 mL), were grown overnight at 37 °C in 10 mL Nystatin (30 g/disc) was used as of Mueller Hinton Broth (Bioxon). The reference and appropriate controls with no cultures were adjusted with sterile saline partitions were used. Each experiment was solution to obtain turbidity comparable to repeated three times. The estimation of the that of McFarland no. 0.5 standard (1.0 x Minimal Inhibitory Concentration (MIC) 108 CFU/mL) (Lennette et al., 1987). Petri and Minimal Fungicide Concentration dishes containing Mueller Hinton agar (MFC) were carried out by the broth (Bioxon) were inoculated with these dilution method (VandenBerghe and microbial suspensions. Vlietinck, 1991). Dilutions of partitions from 2000 to 60 g/mL were used. Test Solutions of 200 mg/mL of each extract yeast culture was used at the concentration were prepared, disks of filter paper of 105 CFU/mL. MIC values were taken as (Whatman no. 5) of 5 mm diameter were the lowest partition concentration that impregnated with 10 L of each one (final prevents visible yeast growth after 24 h of doses per disk: 2000 µg of hexanic and incubation at 37 °C. Nystatin was used as methanolic partitions) and placed on the reference and appropriate controls with no agar surface. Disks impregnated with extract were used. Each experiment was hexane and methanol were used as made three times. negative controls. Disks with chloramphenicol (25 g) were used as The inactivation broth death kinetic positive controls. The plates were method was performed using appropriate incubated overnight at 37 °C and the concentrations of hexanic partition diameter of any resulting zones of (corresponding to ½ MIC, MIC and inhibition (mm) of growth was measured. MBC). Death kinetics expressed in log10 Each experiment was performed in reduction time kills plots (Christoph et al., triplicates. 2000). The estimation of the Minimal Inhibitory Phytochemical screening Concentration (MIC) was carried out by the broth dilution method (Vanden Berghe Preliminary phytochemical analysis was and Vlietinck, 1991). Dilutions of carried out using thin layer partitions from 2000 to 60 g/mL were chromatography on silica gel plates used. Test bacteria culture was used at the developed with a mixture of toluene-ethyl concentration of 105 CFU/mL. MIC values acetate (93:7). Spots were revealed by the were taken as the lowest extract following spray-reagents: Dragendorff and concentration that prevents visible Mayer reagent for alkaloids, vainillin- bacterial growth after 24 h of incubation at sulphuric acid for terpenes and flavonoids, 37 °C. Chloramphenicol was used as and 2% methanol solution of -naphtol reference and appropriate controls with no with concentrated sulphuric acid for extract were used. Each experiment was glicosides. The plates were dried, the made three times. presence of triterpenoids suggested by 188 Int.J.Curr.Microbiol.App.Sci (2013) 2(11): 186-193 violet spots and flavonoid by yellow or effect of the hexanic partition (in the orange spots, mono and sesquiterpenes by survival curve) on a fungal strain blue-violet, red-violet, grey-blue or blue (C. neoformans). Minimum inhibitory spots (Wagner et al., 2001; Sampietro et concentrations (MIC) had a fungistatic al., 2009). effect on the microbial population, while the minimum fungicidal concentrations Statistical analysis (MFC) had a lethal effect on the fungal strain within the first 24 hours. All experiments were