Antimicrobial activity of some Indian spices against food borne pathogens.
Abstract: Antimicrobial activity of 25%, 50%, 75% and 100% alcohol extract of Cinnamomum verum and Syzygium aromaticum has been evaluated against bacterial strains of Pseudomonas lundensis and Bacillus cereus and fungal strains of Aspergillus flavus and Aspergillus niger. Extracts from both Cinnamomum verum and Syzygium aromaticum showed excellent antimicrobial activity against all the test organisms. In the 25% and 50% concentration Cinnamomum verum showed the highest at 19 mm and 16 mm antibacterial zone against Bacillus cereus and Pseudomonas lundensis respectively. In the 75% and 100% concentration Syzygium aromaticum showed the highest at 15 mm and 21 mm antimicrobial zone against Aspergillus niger and Bacillus cereus respectively. Of all the results obtained the maximum antimicrobial zone formation with minimal concentration was recorded at 100% extract of Syzygium aromaticum against Bacillus cereus with 21 mm antibacterial zone and at 100% extract of Syzygium aromaticum with 18 mm of antifungal zone against Aspergillus niger.

Keywords: Spices extract, antimicrobial activity, Pseudomonas lundensis, Bacillus cereus, Aspergillus niger and Aspergillus flavus.
Article Type: Report
Subject: Antibacterial agents (Research)
Foodborne diseases (Care and treatment)
Spices (Health aspects)
Authors: Hema, R.
Kumaravel, I.S.
Martina, D.
Pub Date: 03/22/2011
Publication: Name: Australian Journal of Medical Herbalism Publisher: National Herbalists Association of Australia Audience: Academic Format: Magazine/Journal Subject: Health Copyright: COPYRIGHT 2011 National Herbalists Association of Australia ISSN: 1033-8330
Issue: Date: Spring, 2011 Source Volume: 23 Source Issue: 1
Topic: Event Code: 310 Science & research
Product: Product Code: 2834880 Bacteriostats; 2099930 Spices NAICS Code: 325412 Pharmaceutical Preparation Manufacturing; 311942 Spice and Extract Manufacturing SIC Code: 2834 Pharmaceutical preparations; 2099 Food preparations, not elsewhere classified
Geographic: Geographic Scope: Australia Geographic Code: 8AUST Australia
Accession Number: 254971769
Full Text: Introduction

Food borne pathogens such as Pseudomonas lundensis, Bacillus cereus, Aspergillus niger and Aspergillus flavus are widely distributed in nature causing considerable mortality and morbidity in the population. It is well known that pseudomonads are ubiquitous bacteria in nature. Due to their ability to utilise a wide range of organic compounds they occupy an important ecological position in the carbon cycle, therefore the ecology of pseudomonads in the biosphere has been a matter of interest.

Bacillus cereus has been recognised as an agent of food poisoning since 1955. Between 1972 and 1986, fifty two outbreaks of food borne disease associated with B. cereus were reported (Todar 2008). Bacillus food poisoning strains from 39 outbreaks were identified. B. cereus in 23 outbreaks, B. thuringiensis in 4, B. mycoides in one and mixed strains of Bacillus in 11 outbreaks (McIntyre 2008). A. flavus produce aflatoxin which can cause acute hepatitis, immunosuppression and hepatocellular carcinoma (Klich 2007). The absence of any regulation of screening for the fungus leads to a high prevalence of viral hepatitis and highly increases the risk of hepatocellular carcinoma (Crawford 2005).

Aspergillus niger, if inhaled with large amounts of spores, causes the serious lung disease aspergillosis. A. niger is one of the most common causes of otomycosis (fungal ear infections), which can cause pain, temporary hearing loss and in severe cases damage to the ear canal and tympanic membrane.

These bacteria have a broad host range and have often been isolated from humans with diarrhea (Janda 1998). Since the introduction of antibiotics there has been tremendous increase in the resistance of diverse bacterial pathogens (Cohen 1992, Gold 1996). This shift in susceptibility greatly affects our ability to successfully treat patients empirically. Plant derived products have been used for medicinal purposes for centuries. At present it is estimated that about 80% of the world population rely on botanical preparations as medicines to meet their health needs. Spices are generally considered safe and are proven to be effective against certain ailments (Hora 1944). They are also extensively used particularly in many Asian, African and other countries. In recent years, in view of their beneficial effects, use of spices has been gradually increasing in developed countries.

In the present study we have evaluated the antibacterial effect of the extracts of two widely used spices in India, Cinnamomum verum and Syzygium aromaticum against two bacterial strains of food borne pathogens, Pseudomonas lundensis and Bacillus cereus, and two fungal strains of food borne pathogens, Aspergillus niger and Aspergillus flavus.

Materials and methods

Microorganisms

Pseudomonas lundensis, Bacillus cereus, Aspergillus niger and Aspergillus flavus were the pathogenic microorganisms included in the study. All cultures were obtained in pure form from the culture collection of the Institute of Microbial Technology (IMTECH), Chandigarh, India.

Preparation of spice extracts

The fresh spices were obtained from the local market. The spices were cleaned, descaled where necessary and washed in sterile distilled water. To obtain the spice extracts about 100 g of washed spice was crushed with mortar and pestle. The extracts were sieved through a fine mesh cloth and sterilised using membrane filter (0.45 micron sterile filter). This extract was considered as the 100% concentration of the extract.

The concentrations of 75%, 50% and 25% were made by diluting the concentrated extract with appropriate volumes of sterile distilled water.

Antimicrobial sensitivity testing using filter paper method

Filter paper discs of 7 mm diameter were prepared and sterilised. Using an ethanol dipped and flamed forceps, these discs were aseptically placed over nutrient agar plates seeded with the respective test organisms (Srinivasan 2001). One hundred microlitres of the various spice extracts (100%, 75%, 50%, 25%) were aseptically transferred to these discs. The plates were incubated in an upright position at 37[degrees]C for 24 hours. The diameter of inhibition zones were measured in mm and the results were recorded. Inhibition zones with a diameter less than 12 mm were considered as having no antimicrobial activity. Diameters between 12 and 16 mm were considered moderately active, and those with >16 mm were considered highly active.

All the media used in the present investigation were obtained from Hi-Media Laboratories Ltd, Mumbai India.

Results and discussion

Both the spices tested against the four food borne pathogens showed excellent antimicrobial activity. The results of the antimicrobial activity against the tested pathogens are given in the table.

In the 25% concentration, Cinnamomum verum and Syzygium aromaticum showed the highest of 19 mm and 14 mm with 11 mm and 14 mm antimicrobial zones against Bacillus cereus and Aspergillus flavus respectively. In the 50% concentration, Cinnamomum verum showed the highest of 16 mm antibacterial zone against Pseudomonas lundensis and a maximum of 15 mm antifungal zone against Aspergillus niger and Aspergillus flavus, where Syzygium aromaticum showed the highest antibacterial zone of 11 mm against Bacillus cereus and the highest of 14 mm antifungal zone against Aspergillus niger. In the 75% concentration, the antibacterial zone was not found so significant, whereas Cinnamomum verum showed a prominent antifungal zone against Aspergillus niger, where the antifungal zone was prominent for Cinnamomum verum with 11 mm against Aspergillus niger and 15 mm against Aspergillus flavus. In the 100% concentration, the antibacterial zone was found significant only with Syzygium aromaticum against Bacillus cereus, where the antifungal zone was prominently found with 16 mm against Aspergillus niger, and Syzygium aromaticum with 21 mm against Bacillus cereus. From all the results obtained the maximum antimicrobial zone formation with minimal concentration was recorded with 100% extract of Syzygium aromaticum against Bacillus cereus with 21 mm of antibacterial zone and with 100% extract of Syzygium aromaticum 18 mm of antifungal zone against Aspergillus niger.

References

Cohen ML. 1992. Epidemiology of drug resistance, implications for a post antimicrobial era. Sci 257;1050-5.

Crawford JM. 2005. Liver and Biliary Tract. Pathologic Basis of Disease, Ed. Kumar V et al. Philadelphia: Elsevier Saunders.

Gold SG, Moellering RC. 1996. Antimicrobial drug resistance. N Engl J Med 335;1445-53.

Hora SL, Nair KK. 1944. Pollution of streams and conservation of fisheries. Proc Natl Inst Sci India 10;147-66.

Janda JM, Abbot SL. 1998. Evolving concepts regarding the genus Aeromonas: an expanding panorama of species, disease presentations and unanswered questions. Clin Infect Dis 27:332-4.

Klich MA. 2007. Aspergillus flavus: the major producer of aflatoxin. Molec Plant Pathol 8:6;713-22.

McIntyre L, Bernard K, Beniac D, Isaac-Renton JL, Naseby DC. 2008. Identification of Bacillus cereus group species, associated with food poisoning outbreaks in British Columbia Canada. Appl & Enviro Microbiol 1:11.

Srinivasan D, Sangeetha N, Suresh T, Lakshmanaperumalsamy P. 2001. Antimicrobial activity of certain Indian medicinal plants used in folkloric medicine. J Ethnopharm 74;217-20.

Todar K. 2008. Ph D Bacillus cereus Food Poisoning pg 1. Online textbook of Bacteriol .

* Hema R, Kumaravel IS

Indian Institute of Crop Processing Technology, Thanjavur

Martina D

PRIST University, Thanjavur

* Corresponding author email: hema.scientist@gmail.com
Table 1: Antibacterial activity of different concentrations of
spice extracts

                         Diameter of inhibition zone in
                             mm against various
                        concentrations of spice extracts

Pathogenic organisms          Cinnamomum verum

                        25 %    50 %    75 %    100 %

Pseudomonas lundensis   2 mm    16 m     --     8 mm
Bacillus cereus         19 mm   5 mm    2 mm     --
Aspergillus niger       2 mm    15 m    11 mm   16 mm
Aspergillus flavus      14 mm   15 m    2 mm    10 mm

                         Diameter of inhibition zone in
                             mm against various
                        concentrations of spice extracts

Pathogenic organisms         Syzygium aromaticum

                        25 %    50 %    75 %    100 %

Pseudomonas lundensis   3 mm     --     3 mm    11 mm
Bacillus cereus         3 mm    11 mm   1 mm    21 mm
Aspergillus niger       10 mm   12 mm   10 mm   18 mm
Aspergillus flavus      13 mm   14 mm   15 mm   10 mm
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