Antimicrobial Investigation of Different Extracts of Dolichos lablab beans

 

Akash P. Dahake1*, Arun B. Joshi2 and Krunal M. Patel1

1Pharmacy college, Rampura, Kakanpur, Godhara, Panchmahal, Gujarat, India.

2N.G.S.M Institute of Pharm.Sciences, Paneer, Dearalaktte, Mangalore, India.

 

ABSTRACT

Increasing prevalence of multidrug resistant strains of microorganisms has initiated the exploration of alternate antimicrobial agent. Taking into account the medicinal importance of Dolichos Lablab (Fabaceae) in this respect, an attempt was made in the current study to investigate the antimicrobial potential of this plant. Antimicrobial activity of the ethanolic, petroleum ether, aqueous, chloroform, ethyl acetate and n-hexane extracts of Dolichos Lablab beans were determined against various pathogenic bacteria and fungi. The extracts were tested against various bacteria like Bacillus subtilis, Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli and fungi like Candida albicans by disk diffusion method. Minimum Inhibitory Concentration (MIC) values of each active extract were determined. It is concluded that n-Hexane and chloroform extract exhibited significant antimicrobial and antifungal activity.

 

Keyword- Dolichos Lablab, Antibacterial, Antifungal, Disk diffusion method

 

INTRODUCTION

There is a continuous and urgent need to discover new antimicrobial compounds with diverse chemical structures and novel mechanisms of action because there has been an alarming increase in the incidence of new infectious diseases. Another big concern is the development of resistance to the antibiotics in current clinical use1.

 

The multidrug resistant strain of many microorganisms has revealed exploration of alternative antimicrobial agent. Medicinal plants have become the focus of intense study in terms of validation of their traditional uses through the determination of their actual pharmacological effects. Synthetic drugs are not only expensive and inadequate for the treatment of diseases but also often with adulterations and side effects2.

 

Alternative System of Medicine viz. Ayurveda, Siddha, and Traditional Chinese Medicine have become more popular in recent years. According to one estimate, more than 700 mono and poly-herbal preparations in the form of decoction, tincture, tablets and capsules from more than 100 plants are in clinical use3.

 

Dolichos Lablab (Syn: Lablab Purpureus) is a plant belonging to family Fabaceae4. Traditionally it’s flowers are used as antivinous, alexiteric and carminative. It’s stem is used in the treatment of cholera. The juice from the pods is used to treat inflamed ears and throats. The seeds are anthelmintic, antispasmodic, aphrodisiac, astringent, digestive, febrifuge and stomachic5.

 

Due to the fact that the plant Dolichos Lablab is very useful, as found by above mentioned reports and there is a need to find out more about the potentiality of this plant as an antimicrobial agent. The present study is, therefore, designed to assess the potency of different extracts of beans of Dolichos Lablab on some selected microorganisms.

Table 1. Antimicrobial activity of different extracts of beans of Dolichos Lablab

S. No

Compound

Diameter of zone of inhibition (mm)

S.aureus

B.subtilis

P.aeruginosa

E. coli

C.albicans

1

Ethanolic extract

20

22

13

12

11

2

Pet. ether extract

11

13

11

14

14

3

Aqueous extract

18

17

12

12

13

4

Chloroform extract

19

17

18

25

14

5

Ethyl acetate extract

16

14

12

23

13

6

n-Hexane extract

22

21

17

22

15

7

Ampicillin

23

25

19

26

_

8

Fluconazole

_

_

_

_

19

 

 

 

 

 

 

 

 

 

 

 

 

Table 2. MIC values of four different active extracts of beans of Dolichos Lablab

 

Microorganisms

MIC values (µg/ml)

Ethanolic

extract

Aqueous

extract

Chloroform

extract

Ethyl acetate

extract

n-Hexane

 extract

S.aureus

15.62

15.62

15.62

31.25

15.62

B.subtilis

15.62

15.62

15.62

31.25

15.62

P.aeruginosa

31.25

62.50

15.62

62.50

15.62

E. coli

62.50

62.50

15.62

15.62

15.62

C.albicans

62.50

31.25

31.25

31.25

31.25

MATERIAL AND METHODS

Chemicals used

The following chemicals were used in study: Dimethyl formamide (Merck), Ampicillin (Cadilla pharmaceuticals). Fluconazole (Torrent pharmaceuticals) 

 

Plant material and extraction

The beans of Dolichos Lablab were obtained from Mangalore, Karnataka in India in March 2009 and its botanical identity was confirmed by a taxonomist Dr. Gopalkrishna Bhat, Department of Botany, Poornaprajna college, Udupi. A Voucher specimen was deposited in NGSM Institute of Pharmaceutical Sciences, Paneer, Mangalore, India.

 

Preparation of the extract

Beans were collected from Mangalore, India and dried. Dried beans were ground to fine powder and soxhlet extracted with ethanol, petroleum ether, water, chloroform, ethyl acetate and n-hexane. Each extract was filtered and evaporated to dryness on a rotary evaporator. The dried extracts were then re dissolved in 10% DMSO (v/v) to yield solutions containing 100.0 mg of extract per ml.

 

Microorganisms used

Test organisms (BACTERIA)

Gram positive organisms: Staphylococcus aureus, Bacillus subtilis.

Gram negative organisms: Escherchia coli, Pseudomonas aeruginosa.

Test organisms (FUNGI):    Candida albicans

 

Antimicrobial Screening

Antimicrobial screening of ethanolic, petroleum ether, aqueous, chloroform, ethyl acetate and n-hexane extracts of beans of Dolichos Lablab were performed against 4 pathogenic bacteria (2-gram positive and 2-gram negative) and 1 fungi by the standard disc diffusion method6-8.

 

Each Petridish was inoculated with one of the bacterial cultures suitably diluted to contain above 106 cells/ml by spreading 0.1 ml suspension of the organism with a sterile cotton swab. In each plate cups of 6 mm diameter were made at equal distances using sterile cork borer. One cup was filled with 0.1 ml of standard drug, another

 

with 0.1 ml of DMF, and others were filled with 0.1 ml of samples in sterile DMF. Ampicillin was used as an antibacterial standard while fluconazole as an antifungal standard drug. The Petridish were incubated at 37˚C for 48 hours. The diameter of zone of inhibition in mm was recorded after incubation. The experiment was performed in triplicates and average diameter of zone of inhibition was obtained.

 

Determination of MIC

The extracts that showed antimicrobial activity were subjected to minimum inhibitory concentration (MIC) assay by serial two fold dilution method9. A positive control and negative control were also prepared to confirm the nutritive and sterility properties of the prepared medium respectively. All the tubes were incubated at 37ºC for 24 hours. MIC was interpreted as the lowest concentration of the sample, which showed clear fluid without development of turbidity.

 

RESULTS

The antibacterial activities of the different extracts of Dolichos Lablab showed significant variations as shown in Table 1. Among the six extracts tested, n-Hexnae extract had greater antibacterial potential, followed chloroform, ethanolic, aqueous and ethyl acetate in the decreasing order.

 

The antifungal activities of the n-Hexane, chloroform, ethanolic, aqueous and ethyl acetate extracts of Dolichos Lablab also varied significantly among the test organism as shown in Table 1. n-Hexane had greater antifungal potential and remaining exrtacts showed mild antifungal activity.

 

Antimicrobial potency of the beans extract of Dolichos Lablab against the tested bacterial and fungal strains were expressed in MIC as presented in Table 2. The MIC values against these bacteria, and fungal strains ranged from 15.62 to 62.5µg/ml.

 

CONCLUSION

In our present study, a wide range of human pathogenic microorganisms were examined, including not only Gram-positive and Gram-negative bacteria, but also fungi. Significant antifungal activities also found against C. albicans in this study. This may indicate that the Dolichos Lablab extracts have broad inhibitory activities to

 

Fig.1: Effect of Different Extracts on Bacillus subtilis   

 

Fig.2: Effect of Different Extracts on E. coli

Fig.3: Effect of Different Extracts on Candida albicans

 

Fig.4: Effect of Different Extracts on Pseudomonas aeruginosa

 

Fig.5: Effect of Different Extracts on Staphylococcus aureus

Note:

1.        Ethanolic extract       2. Pet. ether extract

3.        Aqueous extract         4. n.Hexane extract

5.        Chloroform extract   6.Ethyl acetate extract

7.        Ampicillin                 8. Fluconazole

pathogenic microorganisms and promising to act as potential antibacterial and antifungal agents from natural plant sources.

 

Thus, there is now preliminary scientific validation for the use of beans of Dolichos Lablab for antibacterial activity. Further laboratory and clinical studies of this plant is required in order to understand better antibacterial principles which will allow the scientific community to recommend their use as an accessible alternative to synthetic antibiotics.

 

ACKNOWLEDGEMENT

The authors are thankful to B. Pharmacy College, Rampura, Godhara for providing us all necessary facilities.

 

REFERENCES

1.        Rojas R, Bustamante B, Bauer J et al. Antimicrobial activity of selected Peruvian medicinal plants. J Ethnopharmacol 88: 199-204, 2003.

2.        Sieradzki K, Wu Sw, Tomasz A, Micro. Drug Resist. 5(4): 253257 (1999).

3.        Ali M.A., Alam N.M., Yeasmin S., Khan A.M. and Sayeed M.A., Antimicrobial Screening of Different Extracts of Piper longum Linn., Research Journal of Agriculture and Biological Sciences 2007, 3(6), 852-857.

4.        http://www.uasbangalore.edu.in/asp/dolichosBean/html/ products-uses.html

5.        http://www.ibiblio.org/pfaf/cgi-bin/arr_html?Lablab+purpureus

6.        Barry A.L., Procedures for testing antimicrobial agents in agar media. In antibiotic in laboratory medicine, Williams and Wilkins Co., Baltimore, USA. 1980, 1-23.

7.        Bauer A.W., Kirby W.W.M., Sherris J.C. and Turck M., Antibiotic susceptibility testing by a standardized single disc method,  American Journal of Clinical Pathology, 1966, 45, 493-496.

8.        Rios J.J., Reico M.C., and Villar A., Antimicrobial screening of natural products, Journal of Ethnopharmacology, 1988, 23, 127-149.

9.        Florey H.W., Chain E. and Florey M.E., TheAntibiotic, Voli.I, Oxford University Press, NewYork 1989.

 

Received on 12.06.2009

Accepted on 12.08.2009        

© A&V Publication all right reserved

Research Journal of Pharmacognosy  and Phytochemistry. 1(2): Sept. - Oct. 2009, 123-125