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Antibacterial and Antifungal Activity of Litsea Monopetala Leaves on Selected Pathogenic Strains

Md. Fakhrul Hasan
Department of Pharmacy, Southeast University, Dhaka-1213, Bangladesh

Mohammed Ashraful Iqbal
Department of Chemistry, Fareast International University, Dhaka-1213, Bangladesh

Md. Sahab Uddin
Department of Pharmacy, Southeast University, Dhaka-1213, Bangladesh

Abstract/ Executive Summary:

Aims: To evaluate the antibacterial and antifungal activity of some solvent extracts of Litsea monopetala (LM) leaves. Study Design: Examination of antibacterial and antifungal activity of petroleum ether fraction (PEF), chloroform fraction (CLF), ethyl acetate fraction (EAF) and crude methanol extract (CME) of Litsea monopetala (LM) leaves. Place and Duration of Study: Department of Pharmacy, Southeast University, Dhaka-1213, Bangladesh, from September to November 2014. Methodology: Fresh LM leaves were extracted with methanol followed by fractionation. Antibacterial and antifungal activities of the crude extract were determined by using the Agar disk diffusion method against gram positive, gram negative bacterial and fungal strains. Results: All tested plant extracts, showed varying zones of inhibition against bacteria and fungi tested. The zone of inhibition for bacteria was found to be in the range from 5 to 15 mm and 5 to 13 mm for fungi. The CME and EAF of the methanolic extract of LM has greater antibacterial activity against all tested gram positive and gram negative bacteria compare to other fraction. CME and EAF has also strong antifungal activity against tested fungi except Candida albicans in which CLF showed maximum antifungal activity. Conclusion: The present study shows that LM leaves possess an excellent source of natural antibacterial and antifungal agents which could be developed in the treatment of bacterial and fungal diseases.

Keywords: Litsea monopetala; Antibacterial activity; Antifungal activity; Pathogenic strains.

Location: In Bangladesh.

Start Date:

End Date:

Program Area: Development of Host and Medicinal Plants

Commodity: Medicinal Plants

Objectives:

The present study was conducted to estimate the antibacterial and antifungal activities of petroleum ether fraction (PEF), chloroform fraction (CLF), ethyl acetate fraction (EAF) and crude methanol extract (CME) of LM leaves.

Methodology:

2.1 Chemicals All of the chemicals used in this study were analytical grade and purchased from Active Fine Chemicals Ltd., Bangladesh. 2.2 Collection and Identification of Plant Materials The fresh leaves of LM were collected from Comillah, Bangladesh, in September, 2014 and identified by expert of Bangladesh National Herbarium, Mirpur, Dhaka, Bangladesh. Accession number: DACB-35517 for LM. 2.3 Drying and Grinding of Plant Materials The fresh leaves of the plants were first washed with water to remove adhering dirt. Then fruits were cut into small pieces, sun dried for 10 days and finally dried in an oven at temperature not more than 50°C for better grinding. After drying, the entire portions were ground into coarse powder by a grinding machine and stored in an airtight container for further use. 2.4 Extraction and Fractionation of Plant Materials Powdered sample having a weight of 250 g was taken in an amber colored glass bottle and soaked in 500 ml of 95% methanol at 25°C. The bottle with its contents were sealed and kept for a period of about 7 days with occasional shaking and stirring. The whole mixture was then filtered through cotton and then through Whatman No.1 filter paper. Then the filtrate was concentrated with a rotary evaporator under reduced pressure at 50°C temperature to give crude extracts. An aliquot of the concentrated methanol extract was fractionated by petroleum ether, chloroform and ethyl acetate. Concentrated extracts and different fractions were stored until further use and yield value of these were recorded. 2.5 Antimicrobial Activity 2.5.1 Test microorganism Microorganisms used in this study were five gram positive bacteria strains Bacillus subtilis, Bacillus cereus, Bacillus megaterium, Sarcina lutea, Staphylococcus aureus; five gram negative bacteria strains Pseudomonas aeruginosa, Vibrio minicus, Salmonella paratyphi, Shigella dysenteriae, Escherichia coli and three fungal strains Aspergillus niger, Candida albicans and Saccharomyces cerevisiae were obtained from the microbiology lab of the Department of Pharmacy, Southeast University, Dhaka. 2.5.2 Preparation of inoculums. The colony suspension method was used to prepare the inoculum of the test organisms. Active cultures for experiments were prepared by transferring a loopful of cells from the stock cultures to test tubes of Mueller-Hinton broth (MHB) for bacteria and Sabouraud dextrose broth (SDB) for fungi that were incubated without agitation for 24 hrs at 37°C and 25°C respectively. The bacterial and fungal strains were adjusted to a turbidity of 0.5 McFarland standards (approximately 108 CFU/ml for bacteria and 105 or 106 CFU/ml for fungi) with the addition of sterile saline (0.9% NaCl) based on the optical density measurement at 530 nm. 2.5.3 Antibacterial activity Antibacterial activity of the plant extracts was determined by the disc diffusion method [14]. As media MHB was used. Into sterile petri dish 15 ml of molten media was taken to make plates. The plates were allowed to solidify for 5 minutes. Using the micropipette, 100 μl inoculum suspensions that were previously standardized were swabbed uniformly and the inoculum was allowed to dry for 5 minutes. The crude extract at a concentration of 0.1 g/ml was dissolved in 100% dimethyl sulfoxide (DMSO) followed by sterilization using a 0.2 mm Millipore disposable filter. Whatman filter paper (No.1) discs of 6 mm diameter were made by punching the paper. The extracts at a concentration of 500 μg/ml were loaded on 6 mm sterile disc. The loaded disc was placed on the surface of the medium and the compound was allowed to diffuse for 5 minutes and the plates were kept in incubation at 37°C for 24 hrs. Kanamycin was used as positive control at the dose of 30 μg/disc. At the end of incubation, inhibition zones formed around the disc were measured with a transparent ruler in millimeter. DMSO added disc was taken as negative control to determine the possible inhibitory activity of the diluent of extract. The zone of inhibition (ZI) was measured in millimeters. 2.5.4 Antifungal activity. The disc diffusion method was used for the determination of the antifungal activity of the plant extracts. SDB was used as a media. The plates were made by discharging 15 ml molten media into sterile petri dish. Then the plates were permitted to harden for 5 minutes. Using the micropipette, 100 μl inoculum suspensions (previously standardized) were rubbed homogeneously and the inoculum was permitted to dry for 5 minutes. The crude extract with a concentration of 0.1 g/ml was dissolved in 100% DMSO and sterilized by filtration using a Millipore disposable filter having 0.2 mm diameter in size. Whatman filter paper (No.1) discs having 6 mm in diameter were prepared by punching the paper. The extracts at a concentration of 500 μg/ml were occupied on 6 mm sterile disc. The filled disc was positioned on the surface of the medium and the compound was permitted to diffuse for 5 minutes and the plates were reserved for incubation at 25°C for 72 hrs. Griseofulvin at the dose of 30 μg/disc was used as positive control. After the period of incubation, inhibition zones formed around the disc were measured by using a transparent ruler in millimeter. DMSO added disc was considered as negative control to determine the possible inhibitory activity of the diluent of extract. The ZI was measured in millimeters.

Source of Information: European Journal of Medicinal Plants 12(4): 1-8, 2016, Article no.EJMP.23658 ISSN: 2231-0894, NLM ID: 101583475

Article Link (Online Journal): DOI: 10.9734/EJMP/2016/23658

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The results of the present study provide an important basis for the use of the CME and EAF extracts from the leaves of LM for the treatment of bacterial and fungal diseases. The crude extract found to be active in this study could also be useful for the development of new antibacterial and antifungal drugs. However, further pharmacological and toxicity studies will be necessary to establish if they could be safely used as antibacterial and antifungal agents.

Knowledge Management: Journal