DISSERTATION – SYNOPSIS

DR. SNEHA S VIJAYASHEKAR

POST GRADUATE STUDENT

DEPARTMENT OF ORAL PATHOLOGY AND MICROBIOLOGY

BATCH 2013-16

FAROOQIA DENTAL COLLEGE AND HOSPITAL,

UMAR KHAYYAM ROAD, TILAKNAGAR,

MYSORE, KARNATAKA- 570021

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

4th T BLOCK, JAYANAGAR, BANGALORE

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES

4th T BLOCK, JAYANAGAR, BANGALORE

ANNEXURE II

PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION

1. NAME OF THE CANDIDATE AND

ADDRESS

DR. SNEHA S VIJAYASHEKAR,

POST GRADUATE STUDENT,

DEPARTMENT OF ORAL PATHOLOGY AND

MICROBIOLOGY,

FAROOQIA DENTAL COLLEGE AND HOSPITAL,

UMAR KHAYYAM ROAD, MYSORE 570021

2. NAME OF THE INSTITUTION FAROOQIA DENTAL COLLEGE AND HOSPITAL,

UMAR KHAYYAM ROAD, MYSORE 570021

3. COURSE OF STUDY AND SUBJECT MASTER OF DENTAL SURGERY IN ORAL

PATHOLOGY AND MICROBIOLOGY

4. DATE OF ADMISSION OF COURSE 26 JULY 2013

5.

TITLE OF THE TOPIC:

EVALUATION OF ANTIMICROBIAL EFFICACY OF PSIDIUM GUAJAVA, ACACIA

ARABICA, PUNICA GRANATUM ON STREPTOCOCCUS MUTANS, LACTOBACILLI,

PORPHYROMONAS GINGIVALIS, ENTEROCOCCUS FEACALIS AND CANDIDA

ALBICANS.

6. BRIEF RESUME OF THE INTENDED STUDY:

6.1 Need for the study:

Dental biofilms constitute an ecosystem of microorganisms. The major microorganisms includes

Streptococcus mutans, Lactobacilli, Porphyromonas gingivalis, Enterococcus feacalis and Candida

albicans which are responsible for dental caries, endodontic and periodontal diseases. Controlling the

levels of these microorganisms in the oral biofilm will aid in the prevention of these diseases of the oral

cavity.

Numerous antimicrobial agents have been used to eliminate these microorganisms from the oral cavity of

which chlorhexidine is the most commonly used mouth rinse and root canal irrigant. The incorporation of

broad spectrum antimicrobial mouth rinses as adjuncts to oral hygiene regimens has assumed greater

importance with the recognition that most individuals are unable to maintain adequate oral hygiene.

Excessive use of these agents can result in microbial resistance, imbalance in oral flora and numerous

adverse effects. These problems necessitate further research for alternate novel antimicrobial agents that

are safe and potent against specific oral pathogens. Phytochemicals derived from medicinal plants have

proven to have high antimicrobial, anti-inflammatory, anti-oxidant and biocompatible properties. Few

amongs them are,

Punica granatum (pomegranate): This plant has been used to treat various ailments in traditional

medicine. It has been noted for its antimicrobial and antioxidant potency.

Psidium guajava (guava): This tropical tree has been used as an effective and reliable aid in cleaning

teeth in folk medicine. It also has anti-diarrhoeal, anti-inflammatory, anti-oxidant properties etc.

Acacia arabica (Indian gum arabica): The gum of this tree has been used for thousands of years for

various clinical implications. This gum has the potential to inhibit early plaque formation and gingival

inflammation.

The purpose of this study is to evaluate and compare the antimicrobial efficacy of guava leaves,

pomegranate fruit juice and Acacia arabica gum on S. mutans, Lactobacillus, E. faecalis, P. gingivalis and

C. albicans.

6.2 Review of literature:

In a study by Saraya S et al (2008), the anticariogenic activity of guava leaves extracts against S. mutans

by agar diffusion method was studied. The results showed significant bactericidal activity against S.

mutans.1

Pradeep AR et al (2010) compared Acacia arabica gel and chlorhexidine gel and evaluated their

antiplaque effects. Patients on Acacia arabica showed significant clinical improvement in gingival and

plaque scores compared to other patients.2

In a study conducted by Jebashree HS et al (2011) the plant P. guajava was initially evaluated for the

anticariogenic activity against S. mutans by the disc diffusion method. The ethyl acetate extract of this

plant showed high antibacterial activity against S. mutans. The antifungal activity was tested against C.

albicans and the results revealed that P. guajava extract showed siginificant antifungal activity against C.

albicans.3

Alsaimary IE (2012) isolated S mutans, E coli, S aureus, Klebsiella, Proteus and Pseudomonas from 60

samples of dental caries and evaluated the antimicrobial acivity of pomegranate juice against these

organisms. The pomegranate juice was found to be effective against S. mutans in comparison to the other

microorganisms included in there study.4

A study conducted by Kote et al (2011) using pomegranate juice on the dental plaque microorganisms S.

mutans and Lactobacillus showed significant reduction in the level of dental plaque microorganisms after

rinsing with pomegranate juice.5

6.3 Aim of the study:

To evaluate and compare the antimicrobial efficacy of guava leaves, pomegranate fruit juice and Acacia

arabica gum on S. mutans, Lactobacillus, E. faecalis, P. gingivalis and C. albicans.

6.4 Objectives of the study:

1) To evaluate the antimicrobial efficacy of P. guajava against S. mutans, Lactobacillus, E. faecalis,

P. gingivalis and C. albicans.

2) To evaluate the antimicrobial efficacy of P. granatum against S. mutans, Lactobacillus, E.

faecalis, P. gingivalis and C. albicans.

3) To evaluate the antimicrobial efficacy of Acacia arabica against S. mutans, Lactobacillus, E.

faecalis, P. gingivalis and C. albicans.

4) To evaluate the minimal inhibitory concentration of the extracts of P.gaujava, P.granatum, Acacia

arabica on S. mutans, Lactobacillus, E. faecalis, P. gingivalis and C. albicans.

5) To compare the antimicrobial efficacy of P.guajava, P.granatum, Acacia arabica on S. mutans,

Lactobacillus, E. faecalis, P. gingivalis and C. albicans.

7. MATERIALS AND METHODS

7.1 Source of data:

Source of samples: Gauva leaf powder and Acacia arabica gum will be obtained from the NKCA

Pharmacy Ltd., Mysore. Commercially available fresh pomegranate will be obtained.

Standard ATCC microbial cultures S.mutans (ATCC25175), Lactobacillus fermentum (ATCC9338),

P.gingivalis (ATCC33277), E.faecalis (ATCC29212), C.albicans (ATCC10231) will be procured from

Mercury Instruments and Chemicals, Mysore.

Study set up: Department of Oral Pathology and Microbiology, Farooqia Dental College and hospital.

Study Design: An in-vitro study will be conducted to evaluate the antimicrobial efficacy of these

medicinal plants on S.mutans, Lactobacillus,E.faecalis,P.gingivalis and C.albicans.

7.2 METHOD OF COLLECTION OF DATA:

Sample size- Each of the 3 plant extracts will be tested against each of the 5 microorganisms in sets of 6

cultures, giving a total number of 90 culture tests.

MATERIALS

1 Guava leaves extract

2 Acacia Arabica gum extract

3 Pomegranate juice extract

4 0.2% Chlorhexidine solution

5 Distilled water

6 S. mutans, Lactobacillus, E. faecalis, P. gingivalis, C. albicans

7 Brain Heart Infusion broth

8 Brain heart infusion agar

9 Potato Dextrose broth

10 Potato Dextrose Agar (acidified)

11 MRS broth

12 MRS agar

13 Quarter strength ringer’s solution

14 Cork borer

15 Sterilized swabs

16 Incubator

17 Autoclave

18 Vernier caliper

19 Glass micropipettes

20 Petriplates

21 McFarland scale

22 Methanol

23 Soxhlet apparatus

7.3 METHODOLOGY

Guava leaf extract and Acacia arabica gum extract will be prepared with methanol and distilled water as

solvents. Pomegranate juice extract will be prepared from the procured pomegranates.

Microbial cultures will be maintained on brain heart infusion (BHI) broth and bacterial growth will be

checked by changes in turbidity at 24 hrs by using the McFarland scale.

Well agar diffusion method will be performed to test the antimicrobial efficacy of the above extracts. BHI

(brain heart infusion) agar and PDA (potato dextrose agar) plates will be prepared and cultures (200μl)

will be inoculated by spreading on to agar plates by using sterile swab brush across the media. Four round

wells, 4mm deep and 8mm diameter shall be punched using sterile cork borer and it will be marked as A,

B, C, and D.

Group A – Gauva leaf extract

Group B – Acacia Arabica gum extract

Group C – Pomegranate juice

Group D – Chlorhexidine

Guava leaf extract, pomegranate juice, acacia arabica gum extract shall each be added to the respective

wells and the bacterial plates will be incubated for 24hrs at 370C in an incubator. C.albican plates will be

incubated at 280C for 72 hrs. After the incubation period, the plates will be removed and zones of

inhibition will be measured in millimeters using vernier calliper and statistically analysed.

The minimum inhibitory concentration (MIC) will be determined for all the 3 plant extracts against S.

mutans, Lactobacillus, E. faecalis, P. gingivalis and C. albicans as per the standard procedure followed

by National Committee for Clinical Laboratory Standards(NCCL).

Duration of study: 6 months - 1 year.

8.

7.4 STATISTICAL ANALYSIS:

The following statistical procedures will be used to analyze the data.

1. Descriptive statistics

2. Chi- square test

3. Contingency coefficient analysis

4. T test

5. ANOVA

Statistical analysis will be done using ‘ Statistical Package for Social Sciences (SPSS)’ software using

Windows V16.

7.5 Does the study require any investigations or interventions to be conducted on patients or other

humans or animals? If so, please describe briefly.

No.

7.6 Has ethical clearance been obtained from your institution in case of 7.5?

No

LIST OF REFERENCES:

1. Saraya S, Kanta J, Sarisuta N et al. Development of guava extract chewable tablets for anticariogenic

activity against Streptococcus mutans. J of Pharma Sci. 2008;35(1-4):18-23.

2. Pradeep AR, Agarwal E et al. Clinical and microbiologic effects of commercially available gel and

powder containing Acacia arabica on gingivitis. Aust Dent J. 2010;57:312-8.

3. Jebashree HS, Kingsley SJ, Sathish ES, Devapriya D. Antimicrobial activity of few medicinal plants

against clinically isolated human cariogenic pathogens- An In Vitro Study. International Scholarly

Research Network. 2011;1-6.

4. Alsaimary IE. Efficacy of some antibacterial agents against Streptococcus mutans associated with

tooth decay. Afri J of Biotech. 2012;11(88):15457-9.

5. Kote S, Nagesh L. Effect .of pomegranate juice on dental plaque microorganisms (Streptococci and

Lactobacilli). Anc Sci Life. 2011;31(2):49-51.