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Research Article
 

Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves



S. Nirmalraj, M. Ravikumar, M. Mahendrakumar, B. Bharath and K. Perinbam
 
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ABSTRACT

The leaves of Justicia gendarussa are used in the folk medicine for the treatment of various ailments like rheumatic pain, jaundice, cephalgia, hemiplegia and eczema, etc. The leaves of the plant consists of important phytoconstituents including O-di-substituted aromatic amines, 2-aminobenzyl alcohol, O methyl ethers, friedelin, lupeol and β-sistosterol. In the present investigation, different solvent extracts of J. gendarussa leaves were screened and compared for membrane stability and antibacterial activities. Even at lower concentrations, methanolic extract of J. gendarussa shown good antibacterial and anti-inflammatory activity. At higher concentrations, methanolic extracts gave relatively more projecting antibacterial activity when compared with gentamycin. The anti-inflammatory effects of the crude drug were compared with Diclofenac as positive control. The methanolic extracts of J. gendarussa leaves possessed a broad spectrum antibacterial activity against both gram-negative and gram positive organisms like Staphylococcus aureus (MTCC 96), Staphylococcus mutans (MTCC 497), Bacillus subtilis (MTCC 441), Micrococcus luteus (MTCC 1538), gram negative organisms; Proteus vulgaris (MTCC 426), Klebsiella pneumoniae (MTCC 109), Escherichia coli (MTCC 443) and Shigella flexneri (MTCC 1457) with a zone of inhibition from 7-12 mm. The extract also showed good membrane stability to be considered to have significant anti-inflammatory action.

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  How to cite this article:

S. Nirmalraj, M. Ravikumar, M. Mahendrakumar, B. Bharath and K. Perinbam, 2015. Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves. Journal of Plant Sciences, 10: 70-74.

DOI: 10.3923/jps.2015.70.74

URL: https://scialert.net/abstract/?doi=jps.2015.70.74
 
Received: April 07, 2015; Accepted: May 27, 2015; Published: August 03, 2015



INTRODUCTION

Indian system of medicine is mainly depends on plant derivatives. Plants offer a large variety of secondary metabolites which play a significant role in their metabolic pathways. Some species of plants are used for particular disease in a specific region, the same plant is used in different ailment in some other region. Inflammation is injury of living tissues incurred by the result of various enzymes activities. Justicia gendarussa Burm. F. is an ever green shrub belongs to Acanthaceae family which grows up to 1.5-2 m. Justicia gendarussa is frequently grown in Indian gardens as hedge plant, which is propagated by cuttings. It hardly withstands heavy rain and thrives in shade. It is used as a tribal medicine for various ailments such as bronchitis, inflammations, vaginal discharges, dyspepsia, eye diseases and fevers, etc. The leaves were used in the form of decoction, infusions and paste (Anonymous, 1959; Kirtikar and Basu, 1935). The leaves of J. gendarussa contains phytoconstituents such as alkaloids, flavonoids, saturated steroidal saponins or triterpinoidal saponins, amino acids and aromatic amines are rich in potassium salts (Ratnasooriya et al., 2007). The leaves also contains 2-amino benzyl alcohol, 2 (2’-amino benzyl amino) benzyl alcohol and their respective 0-methyl ethers, friedelin, lupeol and β-sitosterol (Arokiyaraj et al., 2007; Herrera-Mata et al., 2002). Long-chain 2-amino alcohols display interesting biological effects like immunosuppresive, anti-inflammatory and cytotoxic activities, as well as inducing apoptosis (Constantinou-Kokotou, 2002). The detection of traditional medicine as an alternative form of healthcare and the development of microbial conflict to the available antibiotics has led to the investigation of the antimicrobial activity of medicinal plants. The increasing disappointment if chemotherapeutics and antibiotics resistance exhibited by pathogenic microbial infectious agents have lead to the screening of several medicinal plants for their potential antimicrobial activity (Elizabeth, 2005). Plant drugs are known to be good for combination therapy which as multidrug resistance modifiers (Hemaiswarya et al., 2008). In the present study, the antibacterial and anti-inflammatory activity of different solvent extracts of Justicia gendarussa were studied.

MATERIALS AND METHODS

Plant material: The leaf samples of J. gendarussa were collected from Semmedu hamlet, Kolli hills, Tamil Nadu, South India. The plant was authenticated by Botanical survey of India, Coimbatore.

Solvent extraction: Justicia gendarussa leaves were dried at room temperature and ground into fine powder. Hundred grams (100 g) of powdered plant material was extracted with 500 mL (1:5) of different solvents of increasing polarity such as hexane, diethyl ether, dichloromethane, ethyl acetate and methanol. The soxhlet extraction method was followed for 6 h with 10-12 cycles under laboratory conditions. The extracts were then concentrated under reduced pressure in a rotary vacuum evaporator (Superfit Rotavap Pub-6, India). All the obtained extracts were weighed, packed and stored at 4°C (Nabere et al., 2013).

Antibacterial study: Eight bacteria were used in this study consisted of both gram positive and gram negative such as, Staphylococcus aureus (MTCC 96), Staphylococcus mutans (MTCC 497), Bacillus subtilis (MTCC 441), Micrococcus luteus (MTCC 1538), Proteus vulgaris (MTCC 426), Klebsiella pneumoniae (MTCC 109), Escherichia coli (MTCC 443) and Shigella flexneri (MTCC 1457). Sensitivity of different bacterial strains to various solvent extracts was measured using disc diffusion method (Bauer et al., 1966; Chew et al., 2011). The plates containing Mueller- Hinton agar were spread with 0.2 mL of the overnight grown bacterial inoculum. About 6 mm diameter of sterile Whatman filter paper (No. 1) were soaked with 10 μL of extract (20 mg mL-1) and deposited in plates. The plates were incubated at 37°C up to 24 h and diameter of the resultant zone of inhibition was measured. For each combination of extract and the bacterial strain, the experiment was performed in triplicate. The bacteria with a clear zone of inhibition of more than 12 mm were considered to be sensitive. Sensitivity of different bacterial strains to DMSO (Dimethyl sulfoxide) was measured to evaluate this solvent toxicity. Antibacterial activity of different solvent extracts was compared with commonly employed antibiotic gentamycin (10 μg disc-1).

Anti-inflammatory (membrane stability) activity assay
HRBC membrane stabilization method: The anti-inflammatory activity of different solvent extracts of Justicia gendarussa leaves were assessed in vitro by Human Red Blood Cell (HRBC) membrane stabilization method. Blood was collected from healthy volunteers and was mixed with equal volume of Alsever’s solution (dextrose 2%, sodium citrate 0.8%, citric acid 0.05% and sodium chloride 0.42% in distilled water) and centrifuged with isosaline. To 1 mL of HRBC suspension, equal volume of test drug in three different concentrations, 1000, 500 and 250 g mL-1 was added. All the assay mixtures were incubated at 37°C for 30 min and centrifuged. The haemoglobin content in the supernatant solution was estimated spectrophotometrically at 560 nm (James et al., 2009). The percentage of haemolysis was calculated then by the equation:

Image for - Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves

The percentage of membrane protection can be hence calculated using the equation:

Image for - Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves

Here "OD of test" is optical density or the test sample’s absorbance and "OD of control" is optical density or absorbance of the negative control. Here, the negative control used was Alsever’s solution with blood without plant extract in it.

RESULTS AND DISCUSSION

Antibacterial activity: Table 1 depicted the antibacterial activity of different solvent extract of J. gendarussa via. hexane, diethylether, dichloromethane, ethyl acetate and methanol extract. Among the tested extracts, the better zone of inhibition was observed with methanol fractions of J. gendarussa (12 mm) against E. coli (MTCC 443) and Bacillus subtilis (MTCC 441). All the extracts of J. gendarussa exhibited inhibition effect on the eight bacteria tested in the present study. The maximum inhibition against Bacillus subtilis, Escherichia coli and this antibacterial activity could be endorsed to different compounds identified in the plant. Antibacterial activity is accredited to several phytochemicals such as flavonoids, saponins, tannins and terpenoids (Sonal et al., 2011), O-disubstituted aromatic amines, 2-aminobenzyl alcohol, O methyl ethers, friedelin, lupeol and β-sistosterol (Arokiyaraj et al., 2007). These various potentialities related to the compounds found in the extracts of our plant could explain their high use in South Indian traditional medicine in the treatment of infectious diseases.

Table 1: Anti bacterial activity of Justicia gendarussa Burm. F
Image for - Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves
DEE: Diethyl ether, DCM: Dichloromethane, Positive control: Gentamycin

Table 2: HRBC membrane stabilization activity Justicia gendarussa (Burm. F.)
Image for - Antibacterial and Anti-Inflammatory Activity of Justicia gendarussa Burm. F. Leaves

HRBC membrane stabilization assay: The lysosomal enzyme released during inflammation produces a variety of disorders. The extracellular activity of these enzymes is said to be related to acute or chronic inflammation. The non-steroidal drugs act either by inhibiting these lysosomal enzymes or by stabilizing the lysosomal membrane. Since, Human Red Blood Cell (HRBC) membrane is similar to lysosomal membrane, their study was undertaken to check the stability of HRBC membrane by the extracts to predict the anti-inflammatory activity in vitro (Varadarasu et al., 2007). Table 2 revealed that the Human Red Blood Cell (HRBC) membrane stability of J. gendarussa via. anti-inflammatory activity of different extracts. Justicia gendarussa showed significant anti-inflammatory activity in a concentration dependent manner. Justicia gendarussa methanolic extract was determined the concentration of 1000, 500 and 250 mg mL-1. The maximum concentration was observed the methanolic extract (1000 mg mL-1) 76.20, 72.46 and 73.62% protection of HRBC in hypotonic solution respectively. All the results were compared with standard Diclofenac at 50 mg mL-1, which showed 74.14% protection of HRBC in hypotonic solution.

CONCLUSION

The methanolic leaf extract of Justicia gendarussa possesses potent anti-inflammatory activity by inhibiting the release of prostaglandins or other inflammatory mediators from cell membrane by stabilizing membrane and anti bacterial activity by zone of inhibition. In further studies, it can be possible to formulate natural anti inflammatory, anti-bacterial drugs of Justicia gendarussa leaf extract.

ACKNOWLEDGMENTS

The authors are thankful to Dr. K. Elumalai and Dr. S. Uma from Center for Entomotoxicity Studies, Department of Zoology, Government Arts College, Nandanam, Chennai for their valuable comments and support in this study work.

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