INTRODUCTION:

Oral diseases such as dental caries, periodontal diseases and teeth loss have great influence on human health. Among these, dental caries is one of the most prevalent infectious diseases all over the world affecting people of all age groups and is caused by the interaction among oral microflora, diet, dentition and oral environment. It is a chronic disease and affects 60-90% of young population. A number of microorganisms such as mutans streptococci, lactobacilli and Actinomyces are involved in causing dental caries. Among these, Streptococcus mutans is considered as the main aetiological agent, although additional acidogenic microorganisms such as lactobacilli may also be involved. The cariogenicity of this pathogen is attributed to its ability to metabolize dietary carbohydrates, to adhere to and to form biofilm on tooth surfaces ^[1-4]. The development of resistance in cariogenic bacteria against most commonly used antimicrobials in dentistry and the cost and possible side effects associated with the synthetic antimicrobials provoked researchers to screen plants for developing novel antimicrobials with anticaries activity. In the present study, we have determined anticaries activity of methanolic extract of eleven plants (Table 1) against clinical isolates of S. mutans recovered from dental caries subjects.

MATERIALS AND METHODS:

Plant materials used in this study:

A total of 11 plants were used to screen their inhibitory efficacy against 7 clinical isolates of S. mutans isolates recovered previously from patients suffering from dental caries. Details of the plants used are shown in Table 1.

Table 1: Plants used in this study:

Plant	Part used	Family
Coscinium fenestraum	Stem	Menispermaceae
Putranjiva roxburghii	Seed	Putranjivaceae
Curcuma aromatica	Rhizome	Zingiberaceae
Nardostachys jatamansi	Whole plant	Valerianaceae
Abrus pulchellus	Leaf	Fabaceae
Polyalthia longifolia	Leaf	Annonaceae
Artocarpus lakoocha	Leaf	Moraceae
Lagerstroemia speciosa	Seed	Lythraceae
Emblica offinalis	Leaf	Euphorbiaceae
Tephrosia purpurea	Leaf	Fabaceae
Maesa indica	Leaf	Myrsinaceae

Extraction:

The shade dried plant materials were extracted using methanol in a soxhlet apparatus. The solvent extracts were filtered through 4-fold muslin cloth followed by Whatman filter paper No. 1, concentrated in rotary evaporator and dried in desiccator ^[5].

Anticaries activity of extracts of selected plants:

Inhibitory activity of selected plant extracts was tested against 7 clinical isolates of S. mutans (Sm) recovered from dental caries subjects by Agar well diffusion method. The bacterial isolates were inoculated into sterile Brain heart infusion broth (HiMedia, Mumbai) tubes and incubated overnight at 37^oC. The broth cultures of isolates were aseptically swabbed on sterile Brain heart infusion agar (HiMedia, Mumbai) plates using sterile cotton swabs. By using a sterile cork borer, wells of 6mm diameter were punched in the inoculated plates and 100µl of extracts (25mg/ml of 25% DMSO), standard (Streptomycin, 1mg/ml) and DMSO (25%) were transferred into respectively labelled wells. The plates were incubated at 37^oC for 24 hours and the zone of inhibition formed around the wells was measured. The experiment was repeated two times and the average reading was noted ^[5].

RESULTS:

Table 2 shows the result of anticaries activity of extracts of plants selected. Extracts of all plants showed inhibition of isolates of S. mutans but to varied extents. It was observed that stem extract of C. fenestratum, rhizome extract of C. aromatica and leaf extract of P. longifolia inhibited bacterial isolates to higher extent. Least inhibitory activity was observed in case of leaf extract of E. officinalis. Remaining plants showed moderate inhibition of bacterial isolates. Inhibition caused by streptomycin was higher than that of plant extracts. DMSO was not found to cause inhibition of bacterial isolates.

Table 2: Anticaries activity of extracts of plants selected:

Extract/ Standard	Zone of inhibition in cm
Extract/ Standard	Sm-1	Sm-2	Sm-3	Sm-4	Sm-5	Sm-6	Sm-7
C. fenestraum	3.0	2.9	2.7	2.7	2.3	2.5	2.4
P. roxburghii	1.6	1.6	1.4	2.1	1.6	1.7	1.9
C. aromatica	2.5	3.1	3.2	2.4	2.1	2.6	3.0
N. jatamansi	2.0	1.9	1.7	1.9	1.6	1.7	1.5
A. pulchellus	2.0	2.5	1.4	1.3	1.2	1.4	1.5
P. longifolia	3.4	3.2	2.9	2.8	3.1	2.9	3.3
A. lakoocha	2.1	2.1	1.6	1.4	1.9	2.0	2.1
L. speciosa	1.9	1.6	2.0	1.7	1.9	2.4	2.1
E. offinalis	1.2	1.2	1.0	1.1	1.3	1.2	1.3
T. purpurea	1.6	1.6	1.3	1.4	1.5	1.6	1.5
M. indica	1.3	1.5	1.8	1.6	1.7	1.3	1.4
Streptomycin	3.5	3.9	3.4	2.9	3.3	3.6	3.8

Infectious diseases are the major cause of death worldwide. On discovering antibiotics, it was thought that this problem would disappear. However, it was found that the bacteria have the ability to become resistant to antibiotics. The increase of antibiotic resistance among microorganisms can be attributed to the microbial characteristics, the selective pressure of antibiotic use and social and technical changes that enhance the transmission of resistant organisms ^[6]. Streptococcus mutans is considered one of the most important cariogenic species among human microflora. The suppression of S. mutans by antimicrobial agents is of clinical importance. The use of antibiotics and other antimicrobial agents such as chlorhexidine is routinely followed in modern dentistry to combat dental caries. However, the resistance against antibiotics and other antimicrobial agents has been observed in clinical isolates of S. mutans ^[7,8]. This situation stimulated researchers to screen antimicrobial agents from natural sources. Natural products, in particular plants, have been studied for their efficacy to control dental biofilm. A number of plants have shown to be effective against caries microflora ^{[2-5, 9-12]}. In the present study, extracts of all plants were found effective against clinical isolates but to varied extents. Among plants tested, leaf extract of P. longifolia displayed stronger anticaries activity.

CONCLUSION:

In the present study, we have demonstrated anticaries activity of eleven plant extracts against clinical isolates of S. mutans. These plants can be useful in the development of bioactive agents active against aetiological agents of dental caries.

ACKNOWLEDGEMENTS:

The authors are thankful to HOD, Department of Microbiology, Principal, SRNMN College of Applied Sciences, Shivamogga and NES, Shivamogga for providing facilities to conduct work and moral support. Authors also thank Dr. N. Mallikarjun, Associate Professor and Chairman, Department of Microbiology, Sahyadri Science College (A), Shivamogga for providing bacterial cultures.

REFERENCES:

1 Ferrazzano GF, Amato I, Ingenito A, De Natale A, Pollio A. Anti-cariogenic effects of polyphenols from plant stimulant beverages (cocoa, coffee, tea). Fitoterapia 80; 2009: 255-262.

2 Almeida AAP, Naghetini CC, Santos VR, Antonio AG, Farah A, Gloria MBA. Influence of natural coffee compounds, coffee extracts and increased levels of caffeine on the inhibition of Streptococcus mutans. Food Research International 49; 2012: 459–461.

3 Brighenti FL, Gaetti-Jardim E, Danelon M, Evangelista GV, Delbem ACB. Effect of Psidium cattleianum leaf extract on enamel demineralisation and dental biofilm composition in situ. Archives of Oral Biology 57; 2012: 1034-1040.

4 Narotzki B, Reznick AZ, Aizenbud D, Levy Y. Green tea: A promising natural product in oral health. Archives of Oral Biology 57; 2012: 429-435.

5 Kekuda PTR, Raghavendra HL, Mallikarjun N, Venugopal TM, Kumar AHS. Elemental composition, anticariogenic, pancreatic lipase inhibitory and cytotoxic activity of Artocarpus lakoocha Roxb pericarp. International Journal of Drug Development and Research 4(1); 2012: 330-336.

6 Dzidic S, Suskovic J, Kos B. Antibiotic Resistance Mechanisms in Bacteria: Biochemical and Genetic Aspects. Food Technology and Biotechnology 46(1); 2008: 11-21.

7 Leistevuo J, Helina J, Osterblad M, Leistevuo T, Huovinen P, Tenovuo J. Resistance to mercury and antimicrobial agents in Streptococcus mutans isolates from human subjects in relation to exposure to dental amalgam fillings. Antimicrobial Agents and Chemotherapy 44(2); 2000: 456–457.

8 Fani MM, Kohanteb J, Dayaghi M. Inhibitory activity of garlic (Allium sativum) extract on multidrug-resistant Streptococcus mutans. Journal of the Indian Society of Pedodontics and Preventive Dentistry 25(4); 2007: 164-168.

9 Jose S, Beegum GRJ. In vitro susceptibility of viridans streptococci to leaf extracts of Mangifera Indica. Indian Journal of Microbiology 47; 2007: 160-163.

10 Venugopal TM, Swathi D, Suchitha Y, Prashith Kekuda TR, Mallikarjun N, Soundarya S, Eyasu Ejeta, Raghavendra HL. Mineral composition, cytotoxic and anticariogenic activity of Scleropyrum pentandrum (Dennst.) Mabb. International Journal of Drug Development and Research 3(4); 2011: 344-350.

11 Yano A, Kikuchi S, Takahashi T, Kohama K, Yoshida Y. Inhibitory effects of the phenolic fraction from the pomace of Vitis coignetiae on biofilm formation by Streptococcus mutans. Archives of Oral Biology 57; 2012: 711-719.

12 Junaid S, Dileep N, Rakesh KN, Pavithra GM, Vinayaka KS, Prashith Kekuda TR. Anticariogenic Activity of Gnidia glauca (Fresen.) Gilg, Pothos scandens L. and Elaegnus kologa Schlecht. Journal of Applied Pharmaceutical Science 3(3); 2013: 20-23.

Received on 21.07.2013 Accepted on 01.08.2013

Asian J. Pharm. Tech. 2013; Vol. 3: Issue 3, Pg 107-108