Journal of Dental Herald
Comparitative Evaluation Of Antibacterial Efficacy Of Herbal Extracts And Mouth Washes
Against Subgingival Plaque Bacteria. An Invitro Study.
A. Kishore Kumar , BinduPriya. S , C. Sravani , K. AmruthaSai , S. Poornodaya , N. Ravindra Reddy1 Assistant Professor, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
2 Post Graduates, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
3 Post Graduates, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
4 Post Graduates, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
5 Post Graduates, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
6 Professor and HOD, Dept of Periodontics, C.K.S Theja Institute of Dental Science and Research, Tirupathi.
Periodontitis is a more severe destructive gum disease which may progress irreversibly in breaking down supporting periodontal structures. The
principal etiological factor in periodontal disease is dental plaque. Dental plaque is seen complex of a several hundred species of bacteria living
together forming an adherent biofilm. Bacterial flora in plaque is variable according to the site such as subgingival / supragingival .subgingival
plaque is composed of predominantly gram –ve anaerobic organisms (Tannerella forsythia, Porphyromonas gingivalis and Treponimadenticola)
are considered to be the most predominant organisms in subgingival plaque. Aim of the study was to test in vitro comparison of antibacterial activity
of herbal extract of centellaasciatica with commercially available mouth rinses against subgingival plaque bacteria. Agar well diffusion was the key
process to evaluate the antimicrobial potential of aqueous extract of the plant. MIC data showed results as bark extracts of centellaasciatica
exhibited +ve significant inhibition of bacteria even at low concentrations. Commercially available mouth washes containing synthetic and semi
synthetic active agents have several disadvantages like staining on the teeth, irritation during use, high degree of alcohol content etc. to overcome
the above disadvantages naturally occurring antibacterial herbs can be used. The herbal medicines are normally considered safer than the non
Subgingival, Supragingival, Plaque, Predominantly, Anaerobic, Porphyromonas, Gingivalis, Treponimadenticola
appropriate method of improving the condition of the inflamed
Periodontal disease is recognized as a major public health
tissues. One of the major problems associated with
problem throughout the world and is the most common cause
conventional treatment of systemic administration of
of tooth loss in adults. Periodontal disease is a general term
antibiotics is the distribution of drug throughout the body,
used to describe several pathological conditions that affect the
which is not required and it can also give rise to toxicity
supporting structures /tissues of teeth . Pain, discomfort and
problems. One method of minimizing the distribution of
cosmetic considerations are some of the factors that
therapeutic agents in the body is through the use of local drug
demonstrate severity of the problems associated with dental
delivery system. Many antibacterial are applied directly to the
diseases and hence, it is of utmost importance to minimize and
mouth for the treatment of periodontal diseases. Mouth rinses,
control dental disease . Periodontal treatment aims to cure
irrigating solutions and sustained release devices are some of
inflamed tissue, reduce the number of pathogenic bacteria and
the local delivery systems. Mouthrinsesis most frequently
eliminate the diseased pockets. Mechanical therapy,
recommended to patients whose mechanical oral hygiene
chemotherapy and systemic administration of antibiotics are
procedures are inadequate for the plaque control. The success
some of the clinical methods being utilized currently.
of any mouthrinse depends on its ability in inhibiting or killing
Conventional therapy includes scaling – removal of the
a varied number of pathogenic oral and periodontal
calculus and the plaque, curettage clearing the inflamed soft
microflora.Chlorhexidine, the most commonly used
tissue, and root planning - removal of necrotic tissues on the
compound in various mouthrinses is a proved antimicrobial
root surface.Periodontal diseases are associated with bacterial
agent. This is used in combination with a variety of compounds
infections; therefore antibacterial treatment seems to be an
under different brand names. But prolonged use of Chlorhexidine can cause several side effects including staining of teeth, gastro-intestinal problems, gingivitis, dry mouth and
Address For Correspondence:
periodontal disease some of which even help in preventing it
from occurring. There are a number of herbs that can help
Assistant Professor, Dept of PeriodonticsC.K.S Theja Institute of Dental Science and
eliminate inflammation and infection associated with
periodontal diseases. There is some evidence indicating the
beneficial effect of plant extract on gingival inflammation and plaque accumulation or subgingivalperiodontopathic
Journal of Dental Herald (January 2014, Issue:1, Vol.:1).
Collection Of Mouth Rinses
Centellaasiatica(L.) gotu kola is an ethnomedical plant used in
Three commercially available mouthrinses with different
different continents by diverse ancient cultures and tribal
chemical combinations were taken as M1, M2 and M3 and
groups. The Chinese prescribed the leaves in curing
Ciprofloxacin (CF): Standard CF antibiotic (HIMEDIA) was
leucorrhoea and toxic fever while in India, this plant is used in
dissolved in 1ml of autoclaved, deionised double distilled
the Ayurvedic system of medicine, usually described under the
water in respective concentrations used as control.
name of Mandukaparni, to treat various diseases: asthma, bronchitis, dropsy, elephantiasis, gastric catarrh, kidney
Screening For Antimicrobial Activity
troubles, leprosy, leucorrhoea, skin diseases and urethritis. It is
Antimicrobial effectiveness of various mouthwashes and plant
found to be quite effective in treatment of mouth ulcers,
extract were assessed by Agar diffusion method by cutting
healing of wounds and promote connective tissue growth.
wells from seeded agar and then filling with extract obtained
Asiaticoide and hypaphorine are the mixture of
from plant and mouthwashes. Microbial samples were reduced
pentacyclictriterpene of Centellaasiatica. It is found that
according tostandard methods and the microbial
topical and oral applications of asiaticoside improved wound
suspensionswere prepared according to tube 0/5 McFarland
healing in guinea pigs. A preliminary and follow-up study by
standard(equivalent turbidity 1/5×10 CFU/ml). This
Sastravaha et al investigated the effect of biodegradable chips
suspension was inoculated onto Wilkins charlgren agar
impregnated with Centellaasiaticaand P. granatum.
medium and MullerHinton agar mediumfor estimation of MIC as per CLSI -2012 standards. Then wells with 6 mm diameter
The objective of the study is to screen and compare the
were punchedusing sterile cork-borer on the culture spread
antibacterial activity of plant extract of Centellaasciatica and
plate.40mg/ml concentration of plant extract and 40µl of each
commercially available mouth rinses on periodontal
mouthwash were dispensed into wells and 40µl of 40mg/ml
ciprofloxacin as positive control and the negative control was (5% DMSO). Plates were then incubated at 37?C. After
Materials And Methods
incubation, the antimicrobial activities of the test materials
were determined by measuring the diameter of the zones of
The periodontal organisms (T.f and P.g) were isolated from
inhibition in millimeters with a HI-Antibiotic zone scale (HI-
deepest pockets (> 6mm) of Periodontitis patients from the
Department of Periodontics, CKS Theja College, Tirupati by removing supragingival plaque with a sterile cotton pellet and
subsequently collecting the subgingival plaque sample with a
Minimum Inhibitory Concentration (MIC) was expressed in
sterile paper point ISO-40 (densply) left in the periodontal
terms of Mean ± SD values of all three repeats of each test
pocket for 20 seconds and then the paper point was commuted
sample. ‘p’ value less than 0.005 and is considered to be very
to laboratory for further analysis in a sterile poly vinyl screw
cap vial containing 1ml of reduced transport fluid (HI-MEDIA). The test organisms were confirmed by standard
microbial, biochemical and conventional PCR analysis.
The antimicrobial efficacy of plant extract and mouth rinses
with ciprofloxacin taken as control is shown in Table-1
Comparison of antibacterial efficacy of various mouthrinses with plant extracts
Leaves of Centellaasciatica was collected locally in June 2012
and were identified and approved by department of Botany Sri
Leaves of selected plant was extensively washed under
running tap water for removal of dust particles and epiphytic
hosts normally found on the surface, followed by washing with sterilized distilled water. They were further air-dried on filter paper at room temperature and then powered with the help of sterilized pestle and motar under aseptic condition. Dry
powder was further extracted by following methodologies.
Gingivitis and periodontitis, as the multifactorial diseases are mainly derived by interaction between invasions of causative
bacteria and host immune response of varied degrees. Plaque
10ml air dry powder of the respective plant parts were mixed
induced gingivitis is the most common form of gingivitis and is
well in 100ml of sterilized distilled water and kept at room
induced by accumulation of material plaque containing more
temperature for 24hrs on an orbital shaker with 500 rpm. The
than 300 types of bacterial species. Removal of dental plaque
solution was further filtered using muslin cloth. The filtrate
that forms an important part of controlling and treating
was centrifuged at 5000 rpm for 15min. The supernatant thus
periodontal disease, which brings about both qualitative as
obtained was filtered through Whattman’s filter no.1 under
well as quantitative changes in the subgingivalmicroflora. The
strict aseptic conditions and the filterer was collected in a pre-
simple mechanical cleaning of the mouth has proven
weighed sterilized test tube. Aqueous extracts were cotton
insufficient to combat plaque formation; hence other means to
plugged and stored in refrigerator at 4°c until further use.
control microbial action must also be taken. Liquids are better able to reach areas where toothbrushes would likely get stuck.
Journal of Dental Herald (January 2014, Issue:1, Vol.:1).
Essential oil, chlorohexidine or CPC based product, using a
Drug delivery devices: Fundamentals and applications.
mouthwash will be more effective then brushing alone. Daily
New York: Marcel Dekker 1998; 491-515.
use of any of the antimicrobial mouthwashes can help fight
4. Chlorhexidine Official FDA information, side effects and
plaque buildup in the mouth and as a result fight gingivitis.
u s e s . D r u g i n f o r m a t i o n o n l i n e h t t p : /
The most common side-effect of chlorhexidine is the
formation of extrinsic yellow-brown stains on the teeth and
Plants are the natural reservoir of many
5. Lockwood B. Neutraceuticals A guide for healthcare
antimicrobial agents. In recent times, traditional medicine as
professionals. 2nd ed. London: Pharmaceutical Press
an alternative form of health care and to overcome microbial
resistance has lead the researchers to investigate the
6. Willershausen B, Gruber I and Hamm G. The influence of
antimicrobial activity of medicinal plants. So as an
herbal ingredients on plaque index and bleeding tendency
alternative natural compounds such as plant extracts are used
as natural mouth rinses typically do not contain: Alcohol,
7. Kaim JM, Gultz J, Do L and Scherer W. An in vitro
Sugar, Artificial colors, Artificial sweeteners (such as
investigation of the antimicrobial activity of an
saccharine), Stannous fluoride, a processed form of fluoride
herbalmouthrinse.J. Clin. Dent. (1998) 9: 46-48
that can stain teeth.In the present study plant extract
8. Maquart FX, Bellon G, Gillery P, Wegrowski Y, Borel JP.
Centellaasciatica was tested for its antimicrobial activity
Stimulation of collagen synthesis in fibroblast cultures by a
against periodontal pathogens by comparing three
triterpene extracted from Centellaasiatica. Connect Tissue
mouthwashes taking ciprofloxacin as control. 40µl all the three
mouthwashes showed excellent antimicrobial activities, of
9. S a s t r a v a h a G , Yo t n u e n g n i t P, B o o n c o n g P,
those M1 showed the highest zone of inhibition, taking
Sangtherapitikul P. Adjunctive periodontal treatment with
ciprofloxacin as control and the plant extract Centellaasciatica
Centellaasiatica and Punicagranatum extracts.
of 40mg/ml showed the minimal zone of inhibition of 12mm.
Apreliminarystudy. J IntAcadPeriodontol 2003; 5(4):106-
The active constituents of Centella asciatica have been found
10. Poosattarbejehmir A: focusing on periodontitis as a
to have antimicrobial effects. As perRishikesh et al the
vasculupathay : the therapeutic possibilities from the
methanol extract of Centellaasiaticashowed activity against a
perspective of a dentistry student. JPMBS 2011, 12:
wide a variety of microorganisms.Sastravaha et al in his
5Newman Mg, 11. TakaeHh, Klokkevold Pr, CaranzzaFa:
preliminary and follow-up study investigated the effect of
Clinical Periodontology. St Louis, Saunders: Eslevier;
biodegradable chips impregnated with Centellaasiaticaand P.
granatum in periodontal pockets showing that clinical signs of
12. Bernstein d, Schiff g,purice a. In vitro virucidal
inflammation are reduced.The study concluded that
effectiveness of 0.12% chlorehexidinegluconate mouth
Centellaasciatica plant extract has a significant antimicrobial
effect against periodontal bacteria.It can be used as a possible
13. Loe H, Schiott C. The effect of mouthrinses and topical
alternative for mouth rinses to overcome the side effects of
application of chlorhexidine on the development of dental
chemical agents.However further extensive research and
plaque and gingivitis in man. Journal of Periodontal
development work should be undertaken on the active
components of plant extracts and its products for their better
14. RenuSolanki Some Medicinal Plants with Antimicrobial
economic and therapeutic utilization.
Activity. International Journal Of Comprehensive Pharmacy 2010 ; 01:04.
15. Rishikesh, Md. MofizurRahmanPhytochemical screening
The antimicrobial potency of plants is believed to be due
and in vitro antimicrobial investigation of the methanolic
tannins, saponins, phenolic compounds, essential oils and
extract of centellaasiaticaleaves. IJPSR, 2012; 3(9): 3323-
flavonoids. This in vitro study gives us natural antimicrobial
agents which can help us to control periodontal infections. The effects of these extract may benefit if incorporated in toothpastes, mouth rinses and dental products to reduce
Source of Support : Nill, Conflict of Interest : None declared
plaque. Further studies are required to better evaluate the effect of these extract if used as mouth rinse. In vivo clinical testing is essential to conform in vitro results. More research into herbals in dentistry can give us simple, effective solutions to periodontal diseases.
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delivery of antimicrobial agents for the treatment of periodontal diseases. Eur J Pharm Biopharm 2000; 50: 83-99.
2. Friedman M, Golomb G. New sustained release dosage
form of chlorhexidine for dental use.J Periodontal Res 1982; 17: 323 328.
3. Steinberg D, Friedman M. Sustained release drug delivery
devices for treatment of dental diseases; In: Tyle P, Ed.
Journal of Dental Herald (January 2014, Issue:1, Vol.:1).
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