Screening of Anti-Microbial activity of Leaf Extract of Crotalaria pulchra Andr.

 

Rama Devi M.*, Siva Subramanian N., Gupta V.R.M., Sandeep K. and Arun Kumar P.

Pulla Reddy Institute of Pharmacy, Annaram (V), Jinnaram (M), Medak (Dt), A.P, India.

*Corresponding Author E-mail: siva_subramanian2006@rediffmail.com

ABSTRACT:

The powdered dried leaf of crotalaria pulchra Andr, (fabaceae) were extracted by using solvents chloroform and methanol by maceration method, then their anti-microbial activity was screened against both bacteria and fungus by disc diffusion method at various concentration. The crude extracts exhibited significant zone of inhibition, which were compared with standard drug. The results revealed that the extracts may be used as phytomedicine against the microbial infections.

 

 

INTRODUCTION:

Anti-microbial substance destroys/ inhibits the growth of microorganism. The plant part possessing anti-microbial properties can be used as potential source of anti-microbial agents. Infectious diseases caused by number of micro organisms are the world’s major threat to human health and account for almost 50,000 deaths every day^[¹^]. The most critical problem related with health and causes morbidity and mortality rates in society infectious diseases. Since last decade, there has been an increasing evidence of bacterial and fungal infections due to pollution and this fact coupled with the resistance developed in microorganism to allopathic agents, anti biotic which increased toxicity in human being and animals during prolonged treatment with several anti microbial drug^[2].

 

Plants have the major advantage of being effective and cheaper alternative sources of drugs^[³^]. The anti-microbial compound from plants may inhibit microorganism through different mechanisms than the conventional anti biotic and therefore be of clinical value in the treatment of microbial infections.^[4] So, these facts motivated us to find out a new herbal drug, which can act as anti microbial agents to treat microbial infections.

 

Crotalaria pulchra Andr.; (fabaceae), commonly known as rattlebox. It is a viscous shurb, leaves are simple obavate-oblanceolate, coriaceous, entire, obtuse-rounded. Flowers are bright yellow, in terminal panicle racemes.

 

Pods are oblong, pubescent, hardly exserted the calyx; seeds7-8^[5]. Leafs were used by tribal people for inflammation, skin diseases like scabies and anthelmintic properties. Based on this, an attempt has been made to evaluate the anti-microbial potency of Crotalaria pulchra Andr.

 

MATERIALS AND METHODS:

Plant Material:

The leaves of Crotalaria pulchra Andr;  (fabaceae) was collected from the Tirupathi district in January 2009, and was authenticated by taxonomist Dr. K. Madhava chetty, Department of botany, S.V. University, Tirupathi. The herbarium specimen was deposited in the department of pharmacognosy at our college. The plant material was shade dried, pulverized and stored in air tight container and used for further extraction.

 

Preparation of Extract:

The chloroform and methanol extract of the leaves of Crotalaria pulchra Andr; were prepared by maceration methods. In this process, 250 gm of dried powder was extracted with 500ml of chloroform kept at a room temperature for 24 hrs, and then filtered solvent was allowed to evaporate in a rotary vacuum evaporator. Similar procedure carried out by using methanol. The dry extracts obtained were subjected to anti-microbial activity.

 

Microbial stains:

Two gram-positive bacteria, one gram-negative bacteria and two fungi were used in the study. Two gram-positive bacteria where Bacillus cereus (ATCC 11778), Staphylococcus aureus (ATCC 9144), gram-negative bacteria Escherichia coli (ATCC 25922) and the two fungi were Candida albicans (ATCC 2091) and Aspergillus niger (ATCC 9029).

Table-1

Extracts	Antibacterial activity Zone of  Inhibition in (mm)			Antifungal activity Zone of  Inhibition in (mm)
	Bacillus  cereus ATCC11778	Staphylococcus aureus ATCC9144	Escherichia  coli mutant ATCC25922	Candida  albicans ATCC2091	Aspergillus  niger ATCC9029
Chloroform
10 µg\ml	08	07	07	08	08
20 µg\ml	14	14	15	15	14
50 µg\ml	19	18	19	18	17
Methanol
10 µg\ml	10	11	10	12	11
20 µg\ml	17	18	18	16	18
50 µg\ml	20	21	20	21	20
Standard *	22	23	22	23	24
Control DMF	-	-	-	-	-

*Standard antibiotics ampicilline for bacteria, ketoconazole for fungus (10 µg\ml)

 

Standard antibiotics:

Ampicilline and ketoconazole were used as reference antibiotics against bacteria and fungus respectively.

 

Determination of anti-bacterial activity:

The chloroform and methanol extract of leaves of Crotalaria pulchra were tested for anti-bacterial activity by the disc diffusion method^[6,7]. Different concentration of extract (10, 20, 50µg/ml) were prepared by reconstituting with dimethyl formamide (DMF). The test micro organisms (10µg/ml) were seeded into respective medium by spread plate method with the 24h culture of bacteria growth in nutrient broth. After solidification the filter paper discs (5mm in diameter) impregnated with the extract were placed on test organism seeded plates. Ampicilline (10µg/ml), was used as positive control while dimethyl formamide (DMF) solvent (100µg/ml) was used as negative control. The antibacterial assay plates were incubated at 37ºC for 24h. The diameter of each zone of inhibition was measured in mm.

 

Antifungal activity:

The antifungal activity was tested by disc diffusion method^[8,9]. The potato dextrose agar plates were inoculated with each fungal culture (10 days old) by point inoculation. The filter paper disc (5mm in diameter) impregnated with (10, 20, 50µg/ml) concentration of the extracts was placed on test organism seeded plates. A blank disc impregnated with dimethyl formamide (DMF) was used as negative control and ketoconazole 10µg disc was used as positive control. The activity was determined after 72h of incubating at 28ºC. The diameter of inhibition zones was measured in mm.

 

RESULTS AND DISCUSSION:

Results obtained present study exhibits that the tested plant extract possess potential antibacterial and anti fungal activity against the microorganisms studied. (Table-1) when tested by disc diffusion method, the chloroform leaf extract of Crotalaria pulchra at 50 µg/ml showed significant effect against Bacillus cereus, Staphylococcus aureus, Escherichia coli (around 18 -19mm) and methanol extract at 50 µg/ml showed significant effect against Bacillus cereus, Staphylococcus aureus, Escherichia coli (around 20-21mm) zone of inhibition and standard ampicilline shows around 22-23 mm zone of inhibition. In case antifungal activity the chloroform extract at 50 µg/ml showed around 17-18 mm zone of inhibition against Candida albicans and Aspergillus niger. In chloroform extract at 50 µg/ml showed around 20-21 mm zone of inhibition against Candida albicans and Aspergillus niger and standard ketoconazole shows around 23-24 mm zone of inhibition. The both chloroform and methanol extract showed almost similar activity when compared with the ampicilline and ketoconazole for antimicrobial and anti fungal activity respectively. In summary methanolic extract exhibited more than chloroform extract. Further research in this pursuit focusing on the isolation of individual compounds and finally subjecting to clinical trials promises to open new avenues in the use of plants for their therapeutic purpose.

 

ACKNOWLEDGEMENT:

The authors are thankful to Chairman and Principal, Pulla Reddy Institute of Pharmacy for giving the required facilities for carrying our work.

 

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Received on 30.10.2010          Modified on 20.11.2010

Accepted on 03.12.2010         © RJPT All right reserved