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Pharmacogn Mag. 2010 Jul-Sep; 6(23): 204–207.
PMCID: PMC2950383

Antimicrobial activity of Bauhinia tomentosa and Bauhinia vahlii roots

Abstract

The hexane, ethylacetate and methanol extracts from Bauhinia tomentosa and Bauhinia vahlii roots were tested for their antimicrobial activity against Gram-positive bacteria (four strains), Gram-negative bacteria (three strains) and three fungi strains using microdilution methods, for the determination of minimal inhibition concentration (MIC) and the minimal microbicidal concentration (MMC). The MIC values of hexane extracts of B. tomentosa and B. vahlii roots were more than 250 µg/ml. The MIC values of ethylacetate and methanol extracts of B. tomentosa roots varied from 7.81 to 31.25 µg/ml and 31.25 to 62.50 µg/ml, respectively. The MIC values of ethylacetate and methanol extracts of B. vahlii roots varied from 15.63 to 62.5 µg/ml and 62.5 to 250 µg/ml, respectively. MMC values obtained are two times greater than the corresponding MIC values. The activities of ethylacetate extracts are attributed to the presence of flavonoids and that of methanol extracts are attributed to the presence of tannins.

Keywords: Antibacterial, antifungal, Bauhinia tomentosa, Bauhinia vahlii, minimal inhibition concentration, minimal microbicidal concentration

INTRODUCTION

The relatively large Bauhinia genus (family: Fabaceae) consisting of trees, climbers and shrubs is distributed in a wide range of geographic locations. Certain Bauhinia species have a long history of traditional medicinal applications.[1] Bauhinia tomentosa is a scrambling, many-stemmed shrub or small tree, the branches often dropping, with many slender twigs.[2] It is called as adavimandaramu in Telugu and phalgu in Sanskrit. It has been reported to contain amino acids,[3] proteins,[3] fatty acids,[4] minerals,[5] lectins,[6] protocatechuic acid,[7] phytohemagglutinins,[8] rutin,[9] quercetin[9] and isoquercetin.[10] Bauhinia vahlii is the largest creeper in India and is called adattige in Telugu and asamantaka in sanskrit. It has been reported to contain amino acids,[11] proteins,[11] minerals[12] and flavonoids.[13] Despite the very encouraging traditional medicinal applications of some species of Bauhinia, prior investigations to validate the traditional medicinal applications have not appeared in literature. The aim of this investigation was to evaluate the antimicrobial activity of the crude extracts of B. tomentosa and B. vahlii roots.

MATERIALS AND METHODS

Plant materials

The roots of B. tomentosa and B. vahlii were collected in July 2007 in Thalakona and Tirumala, respectively. Botanical identification of the plants was done by Dr. K. Madhava Chetty, Department of Botany, S.V. University, Tirupati. Specimens of B. tomentosa (Voucher number: 1821) and B. vahlii (Voucher number:1716) were conserved in S.V. University herbarium.

Extraction of plant materials

The air-dried and powdered roots of B. tomentosa and B. vahlii (2 kg each) were extracted in Soxhlet apparatus sequentially with hexane, ethylacetate and methanol. The extracts were concentrated in a rotary evaporator under vacuum at a temperature not more than 50°C.

Microbial strains

Three Gram-negative bacteria, four Gram-positive bacteria and two fungi obtained from American Type Culture Collection (ATCC) were used in this study. Three Gram-negative bacteria were Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853) and Proteus vulgaris (ATCC 12454); four Gram-positive bacteria were Bacillus subtilis (ATCC 10774), Bacillus pumilus (ATCC 14884), Eneterococcus faecalis (ATCC 2912) and Staphylococcus aureus (ATCC 25923) and the two fungi were Candida krusei (ATCC 6258) and Candida albicans (ATCC 10231).

Culture media

Nutrient agar (NA) containing bromocresol purple was used for culturing of Bacillus species while NA was used for other bacteria. Sabouraud glucose agar was used for culturing fungi. Nutrient broth containing 0.05% phenol red and supplemented with 10% glucose (NBGP) was used for the determination of minimal inhibition concentration (MIC) and the minimal microbicidal concentration (MMC). The Muller Hinton agar (MHA) was also used for the determination of MMC.

Reference antibiotics

Chloramphenicol (Sigma-Aldrich, USA) and Nystatin (Sigma-Aldrich) were used as reference antibiotics (RA) against bacteria and yeast, respectively.

Minimal inhibition concentration and minimal microbicidal concentration determinations

The MIC of test samples and RA were determined as follows. The test sample was dissolved in dimethylsulfoxide (DMSO). The test and RA solutions obtained were added to NBGP and made to a final concentration 250 µg/ml. These solutions were serially diluted twofold to obtain concentration ranges of 0.49–250 µg/ml. Inoculums were standardized at 1.5 × 106 CFU/ml by adjusting the optical density to 0.1 at 600 nm in a spectrophotometer. Each concentration was added in a well of 96-well microplate containing 95 l of NBGP and 5 µl of inoculum.[14] The final concentration of DMSO in the well was less than 1%. Preliminary analysis with 1% (v/v) DMSO/NBGP did not affect either the growth of the test organisms or the change in color due to microbial growth. The negative control well consisted of 195 µl of NBGP and 5 µl of the standard inoculum.[14] The plates were covered with a sterile plate sealer, then agitated to mix the content of the wells using a plate shaker and incubated at 37°C for 24 h. The assay was repeated thrice. Microbial growth was determined by observing the change of color in the wells. The color was red where there was no growth and yellow when there was growth. The lowest concentration showing no color change was considered as MIC.

For the determination of MMC, 5 µl of liquid from each well that showed no change of color was plated on MHA and incubated at 37°C for 24 h. The lowest concentration that yielded no growth after this subculturing was taken as MMC.

RESULTS AND DISCUSSION

The antibacterial and antifungal activities of the crude extracts were evaluated and the results are reported in Tables Tables11 and and2.2. The MIC values of hexane extracts from B. tomentosa and B. vahlii roots on all microbial strains were greater than 250 µg/ml. The MIC values of ethylacetate and methanol extracts of B. tomentosa and B. vahlii roots were less than 250 µg/ml. For ethylacetate extract of B. tomentosa roots, MIC values on Pr. vulgaris, S. aureus and other strains were 7.81, 31.25 and 15.63, µg/ml respectively. For ethylacetate extract of B. vahlii roots, MIC values on Pr. vulgaris, S. aureus and other strains were 15.63, 62.5 and 31.25 µg/ml, respectively. For methanol extract of B. tomentosa roots, the MIC values on S. aureus and other strains were 62.50 and 31.25 µg/ml, respectively. For methanol extract of B. vahlii roots, MIC values on Pr. vulgaris, S. aureus and other strains were 62.5, 250 and 125 µg/ml, respectively. The results of MIC [Table 1] and MMC [Table 2] showed that the MMC values obtained are two times higher than MIC on the corresponding sensitive microorganisms.

Table 1
MICs (g/ml) of the hexane, ethylacetate and methanol extracts from B. tomentosa and B. vahlii roots and reference antibiotics
Table 2
MICs (g/ml) of the hexane, ethylacetate and methanol extracts from B. tomentosa and B. vahlii roots and reference antibiotics

In conclusion, it can be said that the extracts of B. tomentosa roots were more potent than the respective extracts of B. vahlii roots. The relative potencies of extracts are in the order of ethylacetate > methanol > hexane.

Preliminary chemical examination showed the presence of sterols and terpenoids in hexane extracts, flavonoids in ethylacetate extracts, glycosides and tannins in methanol extracts of B. tomentosa and B. vahlii roots.

Only the antimicrobial activity of B. tomentosa leaves has been reported in the literature.[15] To the best of our knowledge, antimicrobial activitiy of B. tomentosa and B. vahlii roots is being reported for the first time. Nevertheless, this study supports the traditional use of B. tomentosa roots as a vermifuge[16,17] and as a mouth gargle.[18] In addition, a good number of research papers documented the antimicrobial potency of some species of Bauhinia1 and the activity is attributed to the presence of flavonoids[19] and phenolic compounds.[2021]

Considering the medicinal importance of the tested microorganisms, the results of this study are considered to be very promising in the perspective of new drug discovery from plant sources. Ps. aeruginosa has emerged as one of the most problematic Gram-negative pathogens, with the alarmingly high antibiotic resistance rates.[22] Even with the most effective antibiotics against this pathogen, the resistance rates were detected as 15–20.4% amongst 152 Ps. aeruginosa strains. This pathogen was found to be sensitive to the crude extracts of B. tomentosa and B. vahlii roots. Bacillus species and Es. coli are agents of food poisoning.[23,24]

C. albicans and other Candida species, causing candidiasis, are increasingly important as they are distributed worldwide, and also they are frequent opportunistic pathogens in AIDS patients.[25] Proteus species are widespread in environment and account for 3% of nosocomial infections in the United States.[26] The incidence of enterococcal bacteremia due to En. faecalis is continuously increasing.[27] S. aureus has been associated with persistent and antibiotic-resistant infections.[28]

Our investigation showed that the antimicrobial activity of ethylacetate extracts from B. tomentosa and B. vahlii roots might be related to the presence of flavonoids. Flavonoids are known to exhibit antimicrobial activity through formation of a complex with the bacterial cell wall. The antimicrobial activity of the methanol extracts from B. tomentosa and B. vahlii roots might be attributed to the presence of tannins. The probable mechanism of these phenolic compounds activity includes enzyme inhibition by oxidized compounds, possibly through reaction with sulfhydryl groups or through more nonspecific interactions with proteins.[29]

Finally, antimicrobial activity of the extracts from B. tomentosa and B. vahlii roots may be due to the presence of both antifungal and antibacterial compounds in them. The present study provides an important basis for the use of extracts from these plants for the treatment of infections associated with the studied microorganisms. Isolation and characterization of bioactive compounds from these two species of genus Bauhinia is currently being carried out in our laboratory.

Acknowledgments

The authors acknowledge International Medical University, Malaysia, and University Grants Commission, India, for their financial support.

Footnotes

Source of Support: International Medical University, Malaysia, and University Grants Commission, India

Conflict of Interest: None declared.

REFERENCES

1. Valdir CF. Chemical Composition and Biological Potential of plants from the genus Bauhinia. Phytother Res. 2009;23:1347–54. [PubMed]
2. Orwa C, Mutua A, Kindt R, Jamandass R, Anthony S. Agroforestree Database: A tree reference and selection guide, version 4.0. Available from: http://www.worldagroforestry.org /sites/treedbs/treedatabases.asp [cited 2010 Jan 31]
3. Agbede JO. Chemical analysis of leaf meal and processed seed flours of an aesthetic plant: Bauhinia tomentosa. J Food Agric Environment. 2007;5:233–5.
4. Daulatabad CD, Mulla GM, Mirajkar AM. Vernolic acid from Lagerstoroemia thomsonil and Bauhinia tomentosa seed oils. J Oil Technol assoc Inida. 1991;23:53–4.
5. Chowdhury AR, Banerji R, Misra G, Nigam SK. Fatty acid composition and mineral composition of the seeds of some species of Bauhinia. Feete Seifen Anstrichmittel. 1984;86:237–9.
6. Rao CK, Satyanand N, Rama G. Pollen lectins. J plant sci res. 1991;6:1–5.
7. Nageshwar G, Anuradha SM, Radha Krishnaiah M, Narayana LL. Distribution pattern of phenolic constituents in species of Bauhinia Linn. and its taxonomic significance. Proceedings - Indian academy of Sciences - Plant Sci. 1986;96:1–7.
8. Khanna AL. Lectins: Biology, Biochemistry and Chemical Biochemistry. 1982;2:701–9.
9. Row LR, Viswanadham N. Coloring matter of the flower petals of Bauhinia tomentosa.Proceedings – Indian Academy of Sciences, Section A. 1954;39:240–2.
10. Subramanina SS, Nair AG. Isolation of isoquercitrin from flowers of Bauhinia tomentosa. Indian J Chem. 1963;1:450.
11. Rajaram N, Janardhanan K. Chemical composition and nutritional potential of tribal pulses Bauhinia purpurea, B. racemosa and B. vahlii. J sci food agric. 1991;55:423–46.
12. Sastri BV, Shenolikar IS. Nutritive value of two unusual foods, adda (Bauhinia vahlii) and marking nut (Semecarpus anacardium) kernels. Indian J med res. 1974;62:1673–7. [PubMed]
13. Sultana S, Ilyas M, Kamil M, Shaida WA. Chemical investigation of Bauhinia vahlii (Leguminoceae) J Indian chem soc. 1985;62:337–8.
14. Victor K, Robert M, Bathélémy N, Armelle MT, Francois N, Ghislain WF, et al. Antimicrobial activity of the methanolic extracts and compounds from Treculia obovoidea (Moraceae) J Ethnopharmacol. 2007;112:531–6. [PubMed]
15. Mythreyi R, Murgugan M, Muthusamy P. Antimicrobial activity of the leaves of Bauhinia tomentosa linn. Indian J Pharm Sci. 2005;67:732–4.
16. Nadakarni KM. Vol. 1. Bombay: Popular Prakasan; 2005. The Indian Materia Medica; pp. 183–4.
17. Kirtikar KR, Basu BD. Dehradun: International book distributors and publishers; 1999. Indian Medicinal Plants. Vol. 2; pp. 892–4.
18. Ganesan S. Traditional oral care medicinal plants survey of Tamil Nadu. Nat pro Radiance. 2008;7:166–72.
19. Achenbach H, Stocker M, Constenia MA. Flavonoid and other constituents of Bauhinia manca. Phytochemistry. 1988;27:1835–41.
20. Ali MS, Azhar I, Amtul Z, Ahmad VU, Usmanghani K. Antimicrobial screening of some Caesalpiniaceae. Fitoterapia. 1995;70:299–304.
21. Kumar RS, Sivakumar T, Sunderam RS, Gupta M, Mazumdar UK, Gomathi P, et al. Antioxidant and antimicrobial activities of Bauhinia racemosa L.stem bark. Braz j Med Bio res. 2005;38:1015–24. [PubMed]
22. Savafi L, Duran N, Savaif N, Önlen Y, Ocak S. The prevalence and resistance patterns of Pseudomonas aeruginosa in intensive care units in a University Hospital. Turk J Med Sci. 2005;35:317, 22.
23. Sleigh DJ, Timbury MC. 5th ed. Edinburgh: Churchill Livingstone; 1998. Note on Medical Bacteriology.
24. Besser RE, Lett SM, Weber JT, Doyle MP, Barrett TJ, Well JG, et al. An out break of diarrhea and hemolytic spelling doubt uremic syndrome from Escherichia coli O157:H7 in fresh-pressed apple cider. J Am Med Assoc. 1993;269:2217–20. [PubMed]
25. Cowan MM. Plant products as antimicrobial agents. Clin Microbiol Rev. 1999;12:564–82. [PMC free article] [PubMed]
26. Mohr Ó Hara C, Brenner FW, Michael MJ. Classification, identification and clinical significance of Proteus, Prvidencia and Morganella. Cli Microbiol Rev. 2000;13:534–6.
27. Noskin GA, Peterson LR, Warren JR. Enterococcus faecium and Enterococcus faecalis bacteremia: Acquisition and out come. Clin infect dis. 1995;20:296–301. [PubMed]
28. Lyczak JB, Cannon CL, Pier GB. Lung infection associated with cystic fibrosis. Clin Microbiol Rev. 2002;15:194–222. [PMC free article] [PubMed]
29. Mason TL, Wasserman BP. Inactivation of red beet beta-glucan synthase by native and oxidized phenolic compounds. Phytochemist. 1987;26:2197–202.

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