INTRODUCTION:

The objective of endodontic therapy is to remove diseased tissue, elimination of bacteria present in the canals and dentinal tubules and to prevent post-endodontic recontamination. E. feacalis is one of the primary organisms in patients with post treatment endodontic infection. The major cause of endodontic failure is the survival of microorganisms in the apical portion of root filled teeth. E. feacalis can adhere to the root canal walls, accumulate, and form communities organized in biofilm, which helps it resist destruction by enabling the bacteria to become 1000 times more resistant to phagocytosis, antibodies, and antimicrobials than non-biofilm producing organisms. Although many advances have been made in different aspects of endodontics within the last few years, the main objective remains as elimination of microorganisms from the root canal systems.¹Herbal medicines have been used for many years. There are innumerable types of indigenous plants that have been used by people for centuries in the treatment of many ailments. The history of such usage is long and well documented. There have been numerous reports of the use of traditional plants and natural products for the treatment of oral diseases.^2,3The present study was to evaluate the antibacterial activity of ethanolic root extract of Glycyrrhiza glabra on E. feacalis

Glycyrrhiza glabra, also known as liquorice and sweet wood, is native to the Mediterranean and certain areas of Asia. It was one of the most widely known medicines in ancient history. It has been found useful for arthritis and allergies. In addition licorice has been used for mild Addison’s disease and other adrenal insufficiencies, such as hypoglycemia.⁴Licorice also acts like the hormone, ACTH, causing sodium retention, potassium depletion, and water retention. The herb contains glycyrrhizin, glycyrrhetinic acid, flavonoids, asparagine, iso-flavonoids,and chalcones.⁵The glycoside, glycyrrhizin has a similar structure and activity as the adrenal steroids.⁶ It also possess good anti-bacterial,⁸anti-fungal,⁹anti oxidant,¹⁰ antitussive,¹¹ hepatoprotective¹² and anti-inflammatory activity.¹³Historically, the dried rhizome and root of this plant were employed medicinally as an expectorant and carminative.

MATERIALS AND METHODS:

Plant material:

The ethanolic root extract of Glycyrrhiza glabra was obtained from Green Chem Herbal Extract and Formulations, Bangalore.

Test microorganisms:

Enterococcus feacalis is obtained from Department of Microbiology, Saveetha Dental College and maintained in nutrient agar slope at 4°C.

Methodology:

The extracts were prepared in the following concentrations in sterile water. 2.5mg/ml, 5mg/ml and 10mg/ml. 100µl of extract of different concentrations were loaded on sterile filter paper discs measuring 6mm in diameter, so that the concentration of the extract on each disc was 250µg , 500 µg and 1000 µg respectively. The discs were dried and kept aseptically

Screening of antibacterial activity [Disc diffusion technique]:

Broth culture of the bacterial strains compared to Mac Farland’s standard ^14,150.5 were prepared. Lawn culture of the test organisms were made on the Muller Hinton agar [MHA-Hi media M1084] plates using sterile cotton swab and the plates were dried for 15 minutes. Filter paper discs loaded with different concentrations of the extract were placed on the respective plates. The plates were incubated at 37°C overnight and the zone of inhibition of growth was measured in millimeters. Standard antibiotic discs of amoxicillin (30mcg/disc) and Penicillin G (30mcg/disc) were used as positive control. All the tests were done in triplicate to minimize the test error.

Determination of minimum inhibitory concentration:

Macro broth dilution or tube dilution method was done to determine the Minimum inhibitory concentration (MIC) of the extracts^14,15. A series of two fold dilution of each extract ranging from 8mg/ml to 0.125mg/ml was made in Muller Hinton broth as specified by National Committee for Clinical Laboratory Standards (NCCLS, 1998).100μl of standard inoculum of the bacterial strains matched to 0.5 Mc Farland’s standard were seeded into each dilution. Two control tubes were maintained for each test batch. These included antibiotic control (tube containing extract and growth media without inoculum) and organism control (tube containing the growth medium and the inoculum). The tubes were incubated at 37°C for 24 hours and checked for turbidity. MIC was determined as the highest dilution (that is, lowest concentration) of the extract that showed no visible growth.

Minimum Bactericidal Concentration (MBC):

The MBCs were determined by selecting tubes that showed no growth during MIC determination; a loop full from each tube was sub cultured onto Muller Hinton agar plates and incubated for further 24 hours at 37^oC. The least concentration, at which no growth was observed, was noted as the MBC

RESULT AND DISCUSSION:

The antibacterial activity of the extracts at different concentrations was screened by disc diffusion technique and the zone of inhibition was measured in mm diameter. The results are given in the table 1. The minimum inhibitory concentration [MIC] and minimum bactericidal concentration [MBC] were also determined for the extracts and the results are given in table 2.The extract showed a zone of inhibition of 23 mm diameter at concentration1000 ug and the MIC and MBC values were found to be 2mg/ml.

Oral health influences the general quality of life and poor oral health is linked to chronic conditions and systemic diseases. The association between oral diseases and the oral microbiota is well established. The present study was to evaluate the antibacterial activity of ethanolic root extract of Glycyrrhiza glabra against caries causing organisms. The results obtained from our study shows that ethnolic extract has got a very good antibacterial activity against the selected oral pathogens.

Table1:Anti-bacterial activity of ethanolic root extract of Glycyrrhiza glabra

Extract	Conc. [µg]	Zone of inhibition [In mm diameter]
Ethanolic root extract of Glycyrrhiza glabra	250	14
	500	17
	1000	23
Penicillin G	30mcg/disc	28
Amoxycillin	30mcg/disc	30

CONCLUSION:

The anti-bacterial activities could be enhanced if active components are purified and adequate dosage determined for proper administration. Although there may be benefits to using herbal medicines in the practice of dentistry, we really do not know much about them. With additional research, there definitely will be a niche for herbal treatments in dentistry

REFERENCES:

1 Portenier I, Waltimo TMT , Haapasalo M. Enterococcus faecalis -the root canal survivor and ‘star’ in post treatment disease. Endodontic Topics. 2003; 6: 135-59.

2 Sheetal verma and S.P. Singh., Current and future status of herbal medicines. Veterinary world, Vol. 1 (11): 347 – 350.

3 Enzo A. Palombo. Traditional Medicinal Plant Extracts and Natural Products with Activity against Oral Bacteria: Potential Application in the Prevention and Treatment of Oral Diseases.

Review article. Evidence – Based complementary and Alternative Medicine Vol. 2011, Article ID 680354,15 pages

4 Olukoga A, Donaldson D. Historical perspectives on health. The history of liquorice: the plant, its extract, cultivation, and commercialization and etymology. J R Soc Health 1998; 118: 300-304

5 A. AAl-Qarawi, H.A. Abdel Rahman, B.H. Ali, Liquorice (Glycyrrhiza glabra and the adrenal kidney -pituitary axis in rats. Food and chemical toxicology, Vol 40: issue 10 pages 1525-1527

6 Fan Y. G.; Shi Z. Q.; He B. L.; Extraction separation and application for glycyrrhizic and glycyrrhetic acid. Natural product Research and Development. 1996, 8, 93‐99

7 Yang L.; Liu Y. L.; Lin S. Q.; The determination of flavonoid in 6 kinds of licorice root by HPLC .Acta Pharmaceutics Sinica. 1990, 25,840-848

8 Manoj M. Nitalikar, Kailas C. Munde, Balaji V. Dhore, Sajid N. Shikalgar Antibacterial Activities of Glycyrrhiza glabra Root Extract ,International Journal of PharmTech Research Vol.2, No.1, pp 899‐901

9 Tharkar P.R., Tatiya A.U., Shinde P.R ., Surana S.J., Patil U.K., Antifungal Activity of Glycyrrhiza glabra Linn. and Emblica officinalis Gaertn. by Direct Bioautography Method International Journal of PharmTech Research Vol.2, No.2, pp 1547-1549, April-June 2010

10 Morteza-Semnani K, Saeedi M, Shahnavaz B, Comparison of antioxidant activity of extract from roots of licorice (Glycyrrhiza glabra L.) to commercial antioxidants in 2% hydroquinone cream. J Cosmet. Sci. 2003 Nov-Dec; 54(6):551-8.

11 Anderson DM, Smith WG. The antitussive activity of glycyrrhetinic acid and its derivatives. J.Pharm. Pharmacol. Jul 1961; 13:396‐404.

12 Kang HE, Lee MG, Hwang SJ, Kim SC, Lee CH, et al. Liquiritigenin, a flavonoid aglycone from licorice, has a choleretic effect and the ability to induce hepatic transporters and phase‐II enzymes. Am J Physiol Gastrointest Liver Physiol. Feb 2009; 296(2):372‐81

13 Okimasu E, Moromizato Y, Watanabe S, et al. Inhibition of phospholipase A2 and platelet aggregation by glycyrrhizin, an anti-inflammation drug. Acta Med Okayama 1983; 37:385‐391.

14 Collins,CH and Lyne, P.M 1976.Microbiological methods, London, Butterworths and Co.288p

15 Betty A. Forbes., Daniel F. Sahm., Alice S. Weissfeld. Bailey and Scott’s Diagnostic Microbiology 11^th edition Mosby page 229 – 257.

Received on 12.05.2014 Modified on 25.05.2014

Research J. Pharm. and Tech. 7(11): Nov. 2014 Page 1237-1239

Extract	Concentrations of the extracts
	8mg/ml	4mg/ml	2mg/ml	1mg/ml	0.5mg/ml	0.25mg/ml	0.125mg/ml
	NG	NG	NG	Turbidity	Turbidity	Turbidity	Turbidity