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ORIGINAL ARTICLE
Year : 2017  |  Volume : 35  |  Issue : 1  |  Page : 41-46
 

Effect of green tea mouth rinse on Streptococcus mutans in plaque and saliva in children: An in vivo study


1 Department of Pedodontics and Preventive Dentistry, Government Dental College, Jaipur, Rajasthan, India
2 Department of Pedodontics and Preventive Dentistry, Jaipur Dental College, Jaipur, Rajasthan, India
3 Department of Dentistry, Goyal Child and Dental Clinic, Jaipur, Rajasthan, India
4 Department of General Dentistry, S.P. Medical College, Bikaner, Rajasthan, of, India

Date of Web Publication31-Jan-2017

Correspondence Address:
Anil Kumar Goyal
Department of Pedodontics and Preventive Dentistry, Government Dental College, Jaipur, Rajasthan
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0970-4388.199227

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   Abstract 

Aims: This study was conducted to evaluate and compare the antimicrobial efficacy of green tea catechin as a mouth wash on colony count of Streptococcus mutans in children. Materials and Methods: A sample size of thirty children was selected out of screened 290 children by simple random sampling between the age group of 7 and 12 years. The study was conducted over a period of 2 weeks. After 24 h of oral prophylaxis, the baseline samples were collected and each group was subjected to mouth rinsing with green tea mouth wash for 2 weeks and further plaque and saliva samples were collected at 1- and 2-week intervals from baseline. Microbiological analysis of plaque and saliva samples was done by Dentocult SM strip kit (Orion Diagnostica, Finland), and the results were statistically analyzed and tabulated. Results: Statistically, there was highly significant reduction in S. mutans count in plaque as well as in saliva for after 1- and 2-week intervals from baseline. Conclusion: Hence, finally, our study showed that green tea catechin is effective as a mouth wash against S. mutans and having better action in plaque as compared to saliva. It can be used as an adjunct to commercially available mouthwashes.


Keywords: Antimicrobial efficacy, green tea catechin, mouth wash, Streptococcus mutans plaque, saliva


How to cite this article:
Goyal AK, Bhat M, Sharma M, Garg M, Khairwa A, Garg R. Effect of green tea mouth rinse on Streptococcus mutans in plaque and saliva in children: An in vivo study. J Indian Soc Pedod Prev Dent 2017;35:41-6

How to cite this URL:
Goyal AK, Bhat M, Sharma M, Garg M, Khairwa A, Garg R. Effect of green tea mouth rinse on Streptococcus mutans in plaque and saliva in children: An in vivo study. J Indian Soc Pedod Prev Dent [serial online] 2017 [cited 2017 Feb 28];35:41-6. Available from: http://www.jisppd.com/text.asp?2017/35/1/41/199227



   Introduction Top


Healthy diet and lifestyle are the best weapons to fight disease. Hence, diet and lifestyle of an individual have been studied extensively.[1]

Tea after water is the most popularly consumed beverage worldwide, with a per capita consumption of 120 ml/day.[1] Tea has had a significant role in human history, from its legendary discovery in the year 2737 BC by the Emperor Shen Nung when leaves fell into a pot of boiling water to the present day when tea is a universally enjoyed beverage. Green tea has been used in China as a healthy, medicinal drink for approximately 5000 years. Green tea was the drink of choice among the leaders and the wealthy. However, following the fall of the Mongolian Empire back in 1368 AD, the whole population of China began to experience the wonderful consumption of tea. The medicinal uses of tea were first reported by Chinese scholars in a text written by Pen T'Sao circa 24-221 AD, and the physiological effects of tea continue to challenge scientists.[2]

The flavonoids present in green tea include catechins (flavan-2-ols), such as epicatechin (EC), epicatechin-3-gallate (ECG), epigallocatechin (EGC), and epigallocatechin-3-gallate (EGCG). Green tea also contains carotenoids, tocopherols, ascorbic acid (Vitamin C), minerals, such as Cr, Mn, Se, or Zn, and certain phytochemical compounds.[1],[3]

Numerous studies in a variety of experimental animal models have demonstrated that catechin possesses antioxidant, antimutagenic, antidiabetic, anti-inflammatory, antibacterial and antiviral, and above all, cancer-preventive properties. It also contributes to control the body weight, rise the bone density, as well as stimulate the immune system. Its potential for decreasing the risk of skin cancer is under investigation.[1],[3],[4]

Increasing scientific and consumer interest in health benefits of tea has led to the inclusion of tea extracts in oral nutritional supplements and topical preparations. Green tea contains a number of bioactive chemicals; it is particularly rich in flavonoids, including catechins and their derivatives. The most abundant polyphenolic compound EGCG is thought to contribute to the beneficial effects of tea.[4]

Green tea (Camellia sinensis) contains polyphenolic catechins reported to exhibit significant antimicrobial properties. Green tea, at a concentration of 40 mg/mL, brewed at 90°C at 5, 20, and 40 min, was determined to be moderately effective against Streptococcus mutans.[5]

Therefore, keeping in mind these facts about green tea, this study was conducted to evaluate the antimicrobial efficacy of green tea catechin as a mouth wash on colony count S. mutans in children.


   Materials and Methods Top


This study was conducted in S.S. Public Higher Secondary School, Triveni Nagar, Jaipur, to evaluate and compare the antimicrobial efficacy of conventionally prepared green tea mouth wash. The materials and equipment used for the study [Figure 1] are as follows.
Figure 1: Armamentarium

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  • Mouth mirror, tweezers, and explorer
  • Sterile gloves
  • Ultrasonic scaler
  • Green tea extract (50%)
  • Green tea catechin mouth wash (0.25% w/v)
  • Dentocult SM Strip Mutans Kit of Orion Diagnostica, Finland, containing:


    • Square tip plaque strip (10)
    • Round tip salivary strip (10)
    • Paraffin tablets (10)
    • Bacitracin disc (50)
    • Patient labels (10)
    • Selective culture vials (10)
    • Manufacturer model chart (1)
    • Incubator (BOD Incubator – Super Deluxe) [Table 1].


Table 1: Selective culture media

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Preparation of green tea catechin mouth wash

Green tea extract (50%) powder was prepared by Lakshyam Herbs Pvt. Ltd., Noida, Uttar Pradesh, which was sent to Guru Nanak Institute of Pharmacy, Hyderabad, for preparation of catechin mouth wash (0.25%w/v).

Method

First, the 0.25% w/v catechin solution was prepared by dissolving 250 mg of catechin in 25 ml of distilled water in a 100 ml beaker.

In another beaker, 25 ml of water was mixed and dissolved completely with sodium chloride, sodium benzoate, and sodium bicarbonate.

The second solution is transferred in 0.25% catechin solution and mixed with amaranth solution which was then filtered and collected in a 100 ml measuring cylinder, followed by addition of distilled water to make the volume 100 ml.

The solution was transferred to 100 ml plastic bottles which were sealed under sterile conditions for dispensing to children [Figure 2] and [Table 2].
Figure 2: Green tea catechin mouth wash

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Table 2: Ingredients of mouth wash

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Selection criteria

A sample size of thirty children was selected out of screened 290 children by simple random sampling between the age group of 7–12 years. The study was conducted over a period of 2 weeks.

The subjects volunteered to participate after verbal and written information. Ethical clearance was taken from Ethical Clearance Committee, and informed consent was taken from the parents of all the children. The selected sampling was done taking into consideration the following inclusion and exclusion criteria.

Inclusion criteria

  • Systemically healthy patients
  • No fixed or removable orthodontic appliances or removable prosthesis
  • No use of green tea catechin as oral rinse earlier
  • No history of oral prophylaxis done for at least 3 months before the study
  • Mixed dentition period
  • Decay-Missing-Filled/decay-missing-filled score >4.


Exclusion criteria

  • History of fluoride treatment in the past 2 weeks
  • Eating before 1–2 h before sample collection
  • Use of antimicrobial mouth rinse before several hours of sample collection
  • History of antibiotic therapy in the subjects within, previous 3 months.


After selection of the children according to inclusion and exclusion criteria, oral prophylaxis of all the subjects was done using an ultrasonic scaler. Then, the subjects were instructed to abstain from any oral hygiene measure for next 24 h.

Plaque sample collection

After 24 h of oral prophylaxis, baseline samples were collected from subjects. Culture vials were taken at room temperature 1 h before the sample collection. Bacitracin discs were placed in the culture vials 15 min before the sample collection and vials were shacked thoroughly.

Plaque samples were collected with a sterile probe tip from four specific sites including [Figure 3]:[6]
Figure 3: Plaque sample collection

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  • Buccal surface of right maxillary first molar
  • Labial surface of maxillary incisor
  • Lingual surface of mandibular incisor
  • Lingual surface of the left mandibular first molar.


Collected plaque samples were evenly and thoroughly distributed on square tip plaque strip (Dentocult SM Strip Mutans Kit of Orion Diagnostica, Finland) using sterile ear buds. The same procedure was repeated after 1 week and 2 weeks.

Saliva sample collection

Before collection of salivary samples, the subjects were asked to chew paraffin tablets for 1 min and then spit the remaining saliva. Saliva samples were collected by pressing the round tip salivary strips against the dorsal surface of tongue and removed with gently closed lips [Figure 4]. The same sample collection procedure was repeated after 1 and 2 weeks.
Figure 4: Saliva sample collection

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After collection of both saliva and plaque samples, the strips were placed in the labeled culture vials and incubated at 37°C for 48 h.

After incubation, the colony counts of S. mutans were interpreted using model chart provided by manufacturer and score were given from 0 to 3 after comparing the incubated strips with model chart [Figure 5] and [Figure 6].
Figure 5: Dentocult SM Strip mutans Model chart

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Figure 6: Slide showing Streptococcus mutans CFU/ml in plaque and saliva at (a) baseline (Class-3), (b) 1 week (Class-1) and (c) 2 weeks (Class-0) intervals

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After collecting baseline samples, the subjects were given the green tea catechin mouth rinses and were asked to use 10 ml of mouth wash for 3 min twice in a day to rinse under supervision for 2 weeks.

The subjects were then asked to start maintaining their regular oral hygiene measures.

Statistical analysis

The values recorded and tabulated separately and compared. The results were statistically analyzed using Student's paired t-test.


   Results Top


Mean change and standard deviation of S. mutans count in plaque followed by mouth rinsing with green tea catechin mouth wash subjects were baseline value 2.67 ± 0.47 that was reduced after rinsing with green tea catechin mouth wash to 1.80 ± 0.74 and 1.20 ± 0.54 after 1 week and 2 weeks, respectively [Table 3] and [Graph 1 [Additional file 1]].
Table 3: Mean ± standard deviation of Streptococcus mutans in green tea catechin subjects from baseline to various intervals in plaque

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After 1 week, the mean change from baseline value was 0.87 ± 0.62, and after 2 weeks, it was 1.47 ± 0.50 which is statistically highly significant in relation to P < 0.001 for both 1- and 2-week intervals from baseline [Table 4] and [Graph 2 [Additional file 2]].
Table 4: Mean change ± standard deviation of Streptococcus mutans in green tea catechin subjects from baseline to various intervals in plaque

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In subjects, baseline value of S. mutans in saliva was 2.14 ± 0.72 that was reduced after rinsing with green tea catechin mouth wash to 1.47 ± 0.72 and 1.00 ± 0.73 after 1 week and 2 weeks, respectively [Table 5] and [Graph 3 [Additional file 3]].
Table 5: Mean ± standard deviation of Streptococcus mutans in green tea catechin subjects from baseline to various intervals in saliva

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After 1 week, the mean change from baseline value was 0.67 ± 0.47, and after 2 weeks, it was 1.13 ± 0.49, which is statistically highly significant in relation to P < 0.001 for both 1- and 2-week intervals from baseline [Table 6] and [Graph 4 [Additional file 4]].
Table 6: Mean change ± standard deviation of Streptococcus mutans in green tea catechin subjects from baseline to various intervals in saliva

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   Discussion Top


Green tea which has been considered to have medicinal benefits and is a healthful beverage since ancient times consisting mainly of polyphenols (catechins) which constitute up to 30% of the dry weight. The major catechins in green tea are (+)-catechin (C), (-)-EC, (+)-gallocatechin, (-)-ECG, (-)-EGC, and (-)-EGCG.[6]

Numerous health benefits of green tea and its constituents have been reported. Green tea is a powerful antioxidant and has anti-inflammatory properties.[7]

In subjects, baseline value in plaque was 2.67 ± 0.47 that was reduced after rinsing with green tea catechin mouth wash to 1.80 ± 0.74 and 1.20 ± 0.54 after 1 week and 2 weeks, respectively [Table 3] and [Graph 1].

Otake et al.[8] showed that a mixture of simple catechins extracted from green tea (consisting mainly of EGCG, EGC, and its epimer gallocatechin, EC and ECG), at 100 mg/L (i.e., less than “cup of tea” concentration), caused very substantial inhibition of adherence of S. mutans to saliva-coated hydroxyapatite.

Several workers have demonstrated that the enzymatic activity of glucosyl transferase from S. mutans and Streptococcus sobrinus is inhibited by tea catechins. Otake et al.[8] and Hattori et al.[9] both found that EGCG and ECG were more active in inhibition of glucosyl transferase than other catechins, but the former group reported greater potency (EGCG at 167 mg/L caused 91% inhibition) than that found by the latter (EGCG at 450 mg/L caused 50% inhibition).

Mean change and standard deviation of S. mutans count in plaque followed by mouth rinsing with green tea catechin mouth wash from baseline to various intervals in subjects are shown in [Table 4] and [Graph 2].

After 1 week, the mean change from baseline value was 0.87 ± 0.62, and after 2 weeks, it was 1.47 ± 0.50 which is statistically highly significant in relation to P < 0.001 for both 1- and 2-week intervals from baseline.

EGCG and EC have been reported to disrupt reconstituted bacterial membranes in a model system.[10] Tea and its components show broad-spectrum antibacterial activity.[11]

A study done by Awadalla et al.[12] 2009 to assess the possible protective properties of green tea on oral health, using the following measurements: S. mutans count in saliva and plaque, salivary and plaque pH values, gingival bleeding index (GBI). The results of this study showed that there was a statistically significant difference among subjects pre- and post-rinsing with 2% green tea for 5 min concerning S. mutans count in saliva and plaque, salivary and plaque pH values, and GBI. This study supports the effectiveness of local application of green tea as antibacterial and anticarcinogenic material as it decreases the acidity of the saliva and plaque.

Tehrani et al. compared green tea with sodium fluoride mouth rinse on salivary S. mutans. Green tea mouth rinse resulted in significant reduction of colony number of salivary S. mutans and Lactobacillus, which is comparable with sodium fluoride mouth rinse.[13]

Neturi et al. compared green tea with chlorhexidine (CHX) and plain water on plaque and found that green tea and CHX were equally effective against S. mutans.[14]

In some other study, green tea and honey mouth rinse were assessed on salivary S. mutans. A statistically significant reduction in the average number of S. mutans at baseline and postintervention in the children who were assigned to the honey (P = 0.001) and green tea (P = 0.001) groups was found.[15]

In a recent study against salivary S. mutans, green tea mouth rinse was found to be significantly better than CHX mouth rinse (P = 0.005).[16]


   Conclusion Top


Hence, finally, our study showed that green tea catechin is effective as a mouth wash against S. mutans and having better action in plaque as compared to saliva.

Green tea catechin can be used as an adjunct to commercially available mouthwashes because:

  • Taste of green tea is well accepted by the children
  • Effective in reducing the number of S. mutans in plaque as well as in saliva
  • Cost-effective when compared to commercially available mouthwashes.


Since it is just a beginning, further studies can be conducted to evaluate systemic effects, long-term trials, shelf life, and other oral microorganism antibacterial efficacy with a larger sample size.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
   References Top

1.
McKay DL, Blumberg JB. The role of tea in human health: An update. J Am Coll Nutr 2002;21:1-13.  Back to cited text no. 1
    
2.
Balentine DA. Special issue: Tea and health. Crit Rev Food Sci Nutr 1997;37:691-2.  Back to cited text no. 2
    
3.
Wiseman SA, Balentine DA, Frei B. Antioxidants in tea. Crit Rev Food Sci Nutr 1997;37:705-18.  Back to cited text no. 3
    
4.
Higdon JV, Frei B. Tea catechins and polyphenols: Health effects, metabolism, and antioxidant functions. Crit Rev Food Sci Nutr 2003;43:89-143.  Back to cited text no. 4
    
5.
Becker R, Hirsh S, Hu E, Jamil A, Mathew S, Newcomb K, et al. Inhibitory Effects of Camellia sinensis (Green Tea) on Streptococcus mutans. Available from: http://www.depts.drew.edu/govschl/njgss2009/journal/TeamPapers/team3.pdf. [Last accessed on 2016 Aug 08].  Back to cited text no. 5
    
6.
Asokan S, Rathan J, Muthu MS, Rathna PV, Emmadi P, Raghuraman, et al. Effect of oil pulling on Streptococcus mutans count in plaque and saliva using Dentocult SM strip mutans test: A randomized, controlled, triple-blind study. J Indian Soc Pedod Prev Dent 2008;26:12-7.  Back to cited text no. 6
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7.
Cabrera C, Artacho R, Giménez R. Beneficial effects of green tea – A review. J Am Coll Nutr 2006;25:79-99.  Back to cited text no. 7
    
8.
Otake S, Makimura M, Kuroki T, Nishihara Y, Hirasawa M. Anticaries effects of polyphenolic compounds from Japanese green tea. Caries Res 1991;25:438-43.  Back to cited text no. 8
    
9.
Hattori M, Kusumoto IT, Namba T, Ishigami T, Hara Y. Effect of tea polyphenols on glucan synthesis by glucosyltransferase from Streptococcus mutans. Chem Pharm Bull (Tokyo) 1990;38:717-20.  Back to cited text no. 9
    
10.
Ikigai H, Nakae T, Hara Y, Shimamura T. Bactericidal catechins damage the lipid bilayer. Biochim Biophys Acta 1993;1147:132-6.  Back to cited text no. 10
    
11.
Yam TS, Shah S, Hamilton-Miller JM. Microbiological activity of whole and fractionated crude extracts of tea (Camellia sinensis), and of tea components. FEMS Microbiol Lett 1997;152:169-74.  Back to cited text no. 11
    
12.
Awadalla HI, Ragab MH, Bassuoni MW, Fayed MT, Abbas MO. A pilot study of the role of green tea use on oral health. Int J Dent Hyg 2011;9:110-6.  Back to cited text no. 12
    
13.
Tehrani MH, Asghari G, Hajiahmadi M. Comparing Streptococcus mutans and Lactobacillus colony count changes following green tea mouth rinse or sodium fluoride mouth rinse use in children (Randomized double-blind controlled clinical trial). Dent Res J (Isfahan) 2011;8 Suppl 1:S58-63.  Back to cited text no. 13
    
14.
Neturi RS, Srinivas R, Simha VB, Sree SY, Shekhar CT, Kumar SP. Effects of green tea on Streptococcus mutans counts – A randomised control trail. J Clin Diagn Res 2014;8:ZC128-30.  Back to cited text no. 14
    
15.
Abdelmegid F, Al-Agamy M, Alwohaibi A, Ka'abi H, Salama F. Effect of honey and green tea solutions on Streptococcus mutans. J Clin Pediatr Dent 2015;39:435-41.  Back to cited text no. 15
    
16.
Thomas A, Thakur SR, Shetty SB. Anti-microbial efficacy of green tea and chlorhexidine mouth rinses against Streptococcus mutans, Lactobacilli spp. and Candida albicans in children with severe early childhood caries: A randomized clinical study. J Indian Soc Pedod Prev Dent 2016;34:65-70.  Back to cited text no. 16
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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
 
 
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  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]



 

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