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ORIGINAL ARTICLE
Year : 2019  |  Volume : 37  |  Issue : 4  |  Page : 365-371
 

Effect of sesame oil, ozonated sesame oil, and chlorhexidine mouthwash on oral health status of adolescents: A randomized controlled pilot trial


Department of Pedodontics and Preventive Dentistry, KSR Institute of Dental Science and Research, Tiruchengode, Tamil Nadu, India

Date of Web Publication7-Nov-2019

Correspondence Address:
Dr. Asokan Sharath
Department of Pedodontics and Preventive Dentistry, KSR Institute of Dental Science and Research, Tiruchengode - 637 215, Tamil Nadu
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/JISPPD.JISPPD_244_19

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   Abstract 


Context: Oil pulling procedure involves swishing of oil in the mouth for various oral health benefits. Aim: The aim of the study was to evaluate the effectiveness of sesame oil (SO), ozonated SO (OSO), and chlorhexidine (CHX) mouthwash on the oral health status of adolescents. Study Settings and Design: Parallel multi-arm double-blinded randomized trial was done in a Government higher secondary school. Materials and Methods: A total of 75 adolescents aged 12–14 years with decay-missing-filled index ≤3 were randomly assigned to three groups (n = 25): Group I (SO), Group II (OSO), and Group III (CHX mouthwash). Baseline (T1) Debris Index (DI-S), Calculus Index (CI-S), Oral Hygiene Index-Simplified (OHI-S), Plaque Index (PI), and salivary Streptococcus mutans count were recorded. All the groups were subjected to intervention with the respective mouth rinses for 15 days. The index scores and the salivary S. mutans count were reassessed after 15 days (T2) and 1 month (T3), and the results were statistically analyzed. Statistical Analysis: The statistical analysis was done using IBM SPSS Statistics for Windows. The statistical significance was set at P ≤ 0.05. Kolmogorov–Smirnov and Shapiro–Wilk test were used to test the normality of the data. The Friedman test and Wilcoxon-signed rank test were carried out for intragroup comparison. Kruskal–Wallis and Mann–Whitney U-test were employed to analyze inter-group comparison. Results: All the groups showed statistically significant reduction in DI-S, CI-S, OHI-S, PI, and S. mutans count after 15 days. Conclusion: Oil pulling therapy using SO and OSO showed a significant improvement in oral hygiene.


Keywords: Oil pulling, ozonated oil, sesame oil


How to cite this article:
Vadhana V C, Sharath A, Geethapriya P R, Vijayasankari V. Effect of sesame oil, ozonated sesame oil, and chlorhexidine mouthwash on oral health status of adolescents: A randomized controlled pilot trial. J Indian Soc Pedod Prev Dent 2019;37:365-71

How to cite this URL:
Vadhana V C, Sharath A, Geethapriya P R, Vijayasankari V. Effect of sesame oil, ozonated sesame oil, and chlorhexidine mouthwash on oral health status of adolescents: A randomized controlled pilot trial. J Indian Soc Pedod Prev Dent [serial online] 2019 [cited 2019 Nov 14];37:365-71. Available from: http://www.jisppd.com/text.asp?2019/37/4/365/270479





   Introduction Top


Dental caries is a disease of multifactorial etiology. The caries process is directly linked to the ability of microorganisms to colonize onto the tooth surface and form dental plaque. Dental plaque is a structurally and functionally organized biofilm which has a diverse microbial composition that remains relatively stable over time.[1] Streptococcus mutans is the predominant microorganism found in the dental plaque associated with caries. The development of a preventive regimen that targets the microbial risk factor is the most comprehensive and successful approach toward the prevention of caries.[2]

The most reliable methods of oral hygiene maintenance are mechanical methods of tooth cleaning using toothbrushes in adjuvant with chemotherapeutic agents.[3] Among them, chlorhexidine (CHX) is considered as the “gold standard,” as it has a broad spectrum of activity. However, its use is discouraged because of its unpleasant taste and undesirable side effects such as tooth staining and altered taste sensation.[4]

One of the traditional Indian folk remedies that has been practiced recently is oil pulling which was familiarized by Dr. F. Karach. The swishing of oil activates enzymes and draws toxins out of the blood.[5] Their antioxidant effects damage the cell wall of microorganisms and destroy them. The emulsification process which occurs due to agitation of oil in the mouth leads to the formation of a soapy layer, which can alter the adhesion of the bacteria on the tooth surface, remove superficial worn out squamous cells, and improve oral hygiene.[6] As it inhibits bacterial adhesion, it also prevents plaque coaggregation.[7]

At present, there are a number of indigenous natural medicinal products which deserve due recognition for their contribution to improving oral health. Sesame oil (SO) has the following advantages over the standard and commercially available mouthwashes. It causes no staining, no lingering after taste, and no allergic reactions. It is five to six times more cost-effective than the commercially available mouthwashes, and moreover, readily available in the household.[7] Considering these benefits, oil pulling therapy with SO could be promoted as a measure for the prevention of oral disease.[8]

Currently, ozone therapy is gaining popularity as a modern noninvasive method of treatment. It is a powerful oxidizing agent with a high antimicrobial power against oral pathogens. Ozone (O3), when in contact with organic fluids, causes the formation of reactive oxygen molecules (O2) which influence the cellular metabolism, tissue repair, and antimicrobial effect. In addition, ozone therapy can be systemically or locally applied, cost-effective and has restricted intolerance or contraindication with minimal side effects.[9] Verma et al.[10] analyzed the effect of commercially available ozonated oil on gingivitis and found it to be positive.

Through extensive literature search, it was revealed that there is a lack of adequate studies testing the efficacy of freshly prepared ozonated SO (OSO). Hence, the rationale of this study was to evaluate the effectiveness of SO and OSO on oral health status by assessing the changes in the Debris Index (DI-S), Calculus Index (CI-S), Oral Hygiene Index-Simplified (OHI-S), Plaque Index (PI), and S. mutans count.


   Materials and Methods Top


Preparation of ozonated sesame oil

Ozonized SO was prepared by passing ozone gas through commercially available SO (Idhayam Oil, VVV Sons India) using ozone generator (Ozone Engineers). The output was titrated to 2 g/h for about 2 min to adjust the concentration of ozone to 0.01 ppm. Since half-life of ozone is only 20 min and it was freshly prepared every day just before the usage.

In vitro trial

Preceding the clinical trial, the antimicrobial effect of SO, OSO, and CHX mouthwash was evaluated by agar well-diffusion method. S. mutans (MTCC-890) was lawn cultured in Mueller-Hinton agar following which 6 mm sterile cork broker was used to create wells in agar plates. Each plate had two wells, one for mouth rinse and the other for the antibiotic. Each of the wells was loaded with 50 μl of respective mouth rinse. Triplicates of plates were prepared for each group and incubated at 37°C for 24–48 h to check for their zone of inhibition.

In vivo trial

The protocol of this parallel multi-arm randomized trial was analyzed and approved by the institutional review board and ethical committee. Permission to conduct the study, and the list of schools was obtained from the Assistant Elementary Educational Officer. A Government school was randomly selected by lottery method. The purpose of the study was explained to the school authorities, and their approval was obtained. Written consent in regional language (Tamil) from the parents and verbal assent from adolescents was obtained. Clinical examination was done in school premises by two calibrated investigators. The adolescents were seated on chair and were subjected to dental examination under natural sunlight. A total of 130 adolescents aged 12–14 years were initially screened. Personal details such as medical history, dental history including sugar intake, frequency of brushing, use of topical or systemic antibiotics, mouthwashes, and toothpaste were obtained from the parents using a validated questionnaire. Adolescents aged 12–14 years with decay-missing-filled index <3 and who had not underwent any dental treatment in the past 1 month, no history of systemic and congenital heart diseases, drug allergy, use of topical, or systemic antibiotics for the past 1 month, and use of mouthwashes for the past 3 months were recruited.

The sample size was estimated to be 20 per group based on the study done by Saravanan et al.[11] The power of test was set at 80% and alpha error 5%. To safeguard the optimal level of precision and anticipated attrition of the sample (25%), five samples were added to each group. Accounting this, the sample size was estimated to be 25 adolescents per group.

Seventy-five adolescents were randomly divided into three groups with 25 adolescents in each group using a table of random numbers. Sequence generation and allocation were done by two fellow postgraduates who were not involved in the study (allocation ratio 1:1:1). The enrolled adolescents were allocated to Group I, SO; Group II, OSO; and Group III, CHX mouthwash. In this study, the principal investigator was not blinded of the allocation, as OSO was prepared by the investigator. The secondary investigator, the laboratory technicians, and the statistician were blinded in the study.

Oral prophylaxis was performed for all adolescents 5 days prior to the start of the study. They were instructed to carry on with their routine oral hygiene practices. Baseline (T1) DI-S, CI-S, OHI-S, and PI were recorded for all adolescents. The debris score and calculus score were recorded, and the values were interpreted using OHI-S as described by Greene and Vermilion in 1964. The plaque score was recorded based on the criteria given by Silness and Loe in 1964 after applying the disclosing solution (AlphaPlac, Two Tone Disclosing Solution, The Bombay Burmah Trading Corporation Ltd., Mumbai, Maharashtra, India) on the selected index teeth in both the jaws facially and lingually. The indices were recorded by two independent investigators at the school premises under natural sunlight. The interexaminer reliability was done by Cohen's kappa coefficient and was found to be 0.86.

For the estimation of baseline salivary S. mutans colony count, all the adolescents were instructed not to eat or drink 30 min prior to the saliva sample collection. Stimulated salivary samples were collected in sterile urine cups by chewing paraffin wax. Saliva was collected by spitting method until 2 ml of saliva was collected and transferred to the laboratory for the total colony count of S. mutans. Spread plate method was used for evaluating the S. mutans count.

Brain heart infusion agar was used for culturing S. mutans. Saliva samples of about 1 ml were serially diluted up to 104 dilution and were spread on whole of the agar plates, and the plates were incubated at 37°C at 24 h. The colonies were identified by their morphology and confirmed using Gram staining and catalase test. The S. mutans present in the sample was counted using colony counter, and the number of colonies was noted.

Intervention period

Adolescents were asked not to eat anything 2 h prior to the rinsing procedure. The study groups were allowed to rinse with SO and OSO on all weekdays for 15 days (T2) in the school premises. Around 10 ml of respective oil was taken in the mouth, sipped, sucked, and pulled between the teeth for 10 min until the viscous oil turned thin and milky white. The participants were informed not to swallow the oil, as it might contain bacteria and toxins. The control group was asked to rinse with 10 ml of diluted 0.12% CHX mouthwash for a minute. The procedure was continued for 15 days, and the DI-S, CI-S, OHI-S, and PI were recorded, and salivary samples were collected for the evaluation of S. mutans count.

Adolescents were instructed to discontinue their respective mouth rinses after 15 days of intervention and asked to continue their regular oral hygiene measures. Data collection was repeated after 30 days (T3). [Figure 1] shows CONSORT flowchart of the study design.
Figure 1: CONSORT flowchart

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Statistical analysis

The statistical analysis was done using IBM SPSS Statistics for Windows (Version 22.0, IBM Corp., Armonk, New York, USA). The statistical significance was set at P ≤ 0.05. Kolmogorov–Smirnov and Shapiro–Wilk test were used to test the normality of the data. Since the data did not satisfy the assumptions of equality and normal distribution, Friedman test and Wilcoxon-signed rank test were carried out for intragroup comparison. Kruskal–Wallis and Mann–Whitney U-test were employed to analyze the change in the values for inter-group comparison.


   Results Top


[Table 1] shows the results of antibacterial activity of three mouth rinses against S. mutans. No zone of inhibition was obtained for SO. The addition of ozone to SO produced a zone of inhibition. The maximum zone of inhibition was obtained for CHX mouthwash.
Table 1: Antibacterial efficacy of three groups against Streptococcus mutans

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[Table 2] shows intragroup comparison of DI-S score, CI-S score, OHI-S score, PI score, and salivary S. mutans count at different time intervals. Significant difference in DI-S score, CI-S score, OHI-S score, PI score, and S. mutans count was found in all the three groups at different periods studied. Post hoc analysis showed a significant reduction in DI-S score, CI-S score, OHI-S score, and PI score in all the three groups between T1 and T2 and T1 and T3. S. mutans count showed a significant reduction between T1 and T2 in all three groups. Group I and II showed a significant reduction in scores between T1 and T3.
Table 2: Intragroup comparison of all variables at each time intervals in all groups

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[Table 3] shows inter-group comparison of DI-S score, CI-S score, OHI-S score, PI score, and salivary S. mutans count between the groups at different time intervals. There was no significant difference in the DI-S score, CI-S score, OHI-S score, and salivary S. mutans count between the groups in any of the three-time periods studied. There was a statistically significant difference in the plaque scores between the groups at T2 and T3. Significant reduction in PI scores was seen Group II and III at T2. At T3 significant reduction was seen in Group III.
Table 3: Inter-group comparison of all variables at different time intervals

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   Discussion Top


The oral cavity serves as a focal point of entry for pathogens into the systemic circulation. Dental caries has been found to significantly affect the systemic health and quality of life of individuals.[12] Lack of oral hygiene allows an increase in virulent microbial colonization of the oral biofilm.[13] Therefore, mechanical and chemical means of controlling the quantity and quality of the oral biofilm are important in preventing caries. The use of chemical mouthwash might cause an allergic reaction in few individuals, and prolonged usage causes loss of taste sensation and staining of teeth.[14] Various forms of alternative or traditional medicinal treatments such as Ayurveda have started to gain popularity, one among that is oil pulling. In the present study, SO was used since it is a regular constituent of South Indian cooking and is known to have numerous health benefits. A study by Anand et al.[15] had shown significant reduction in bacterial count after SO pulling and reduced susceptibility of host to dental caries.

Recently, ozone is being discussed as a possible alternative oral antiseptic agent. In dentistry, it is used either as gaseous form, ozonated water, or as ozonated oil. When it is dissolved in an oil base, it chemically reacts with oil and forms long complex molecules.[16] Hence, in this study, OSO was selected over ozonated water, because oil was found to have a prolonged retention in the oral cavity. Considering the lack of adequate studies on freshly prepared ozonated oil, the present study was planned to formulate freshly prepared OSO to evaluate the antibacterial effect of SO and OSO against S. mutans and to compare the effect of SO, OSO with CHX mouthwash on oral health status of adolescents.

Antimicrobial effect

In the present study, maximum zone of inhibition was obtained for CHX followed by OSO. On the other hand, there was no zone of inhibition obtained for SO. This might be because, when ozone gas is bubbled through plant or vegetable extracts, they react with oxygen to form ozonoids which can induce the disruption of microbial cell wall.[17] This was in accordance with the results of Asokan et al.[6] who evaluated the antibacterial activity of SO and lignans (sesamin and sesamolin) isolated from SO against S. mutans, Streptococcus mitis, and S. viridans.

Oral health status

The results of the present study revealed that there was a statistically significant reduction in the debris score, calculus score, and OHI-S scores in all the groups when the baseline score was compared with 15 days and 30 days. Although there was a reduction in scores of all the three groups at 15 days, the reduction was not statistically significant between the groups. Only the CHX group showed a reduction in the scores at the end of 30 days. This was in accordance with the study done by Wang et al.[18] who concluded that CHX containing mouthwash reduced the debris and calculus scores after 201 days which could be due to the inhibition of buildup of debris and calculus formation. In contrary, SO group and OSO group showed slight increase in scores at the end of 30 days. This might be because of discontinuation of the rinses which had no positive effects on the oral hygiene status. Hence, the effect of oil pulling could be enhanced only on routine usage.

Plaque index

The present study revealed there was a statistically significant reduction in plaque scores in all three groups at the two time periods studied. Similar results were obtained by Asokan et al.[19] who concluded that oil pulling therapy significantly reduced the PI scores after 10 days of use. The reduction in plaque might be due to the viscosity of oil or saponification or soap making process due to alkali hydrolysis of fat.[19] Verma et al.[10] compared the effects of ozonated oil and CHX gel, after gingival massage for 3 weeks, and concluded that there was a significant reduction in PI in both the groups.

At the end of 15 days, there was a significant difference in plaque scores between the three groups. CHX and OSO were better in reducing plaque compared to SO. This result was in contradiction to the randomized controlled trial by Dani et al.[20] who assessed the antiplaque effect and plaque-induced gingivitis of oil pulling using SO and found that SO was found to be as effective as CHX. In our study, at the end of 30 days, CHX group showed a minimal increase in plaque scores which was statistically significant. This might be due to the substantivity of CHX, which adhere to the tissues such as oral mucosa and teeth. This helps to maintain a potent sustained release, which, in turn, reduced the bacterial count and prevented the accumulation of dental plaque.[21]

Salivary Streptococcus mutans count

In our study, there was a statistically significant reduction in the salivary S. mutans count in all the three groups when the baseline scores were compared with 15 days. Our findings of SO were in resemblance to the results concluded by Asokan et al.[22] who stated that oil pulling therapy with SO reduced S. mutans count. This might be because of the several probable reasons: (i) the viscous nature of the oil probably inhibits bacterial adhesion, (ii) SO which is a vegetable fat when acted on by the salivary bicarbonates, the soap making process is initiated which are good cleansing agents, (iii) when dispersed in water, it forms minute droplets which increase the surface area of the oil, thereby enhancing its cleansing action, (iv) the lignans sesamin or sesamolin present in the SO could probably protect the oral cavity and reduce the microbial count. Ximenes et al.[23] also stated the similar finding for CHX group, this might be because of the substantial antimicrobial action which causes lysis of bacterial cell wall and precipitates the cytoplasmic content of bacterial cell. In addition, it abolishes the bacterial phosphoenolpyruvate and inhibits its metabolic activity. Celiberti et al.[24] concluded that ozonated water had a better effect than CHX mouthwash in reduction of S. mutans count. This proved the better antibacterial property of ozonated water, which could be the result of oxidation potential of ozonated water that destructs the cell wall and cytoplasmic membranes of bacteria.

At the end of 30 days, there was a statistically significant reduction in the count only in SO and OSO group. This might be because CHX can be bacteriostatic or bactericidal depending on its dosage. Since low concentration of CHX (0.12%) was used in our study, the bacteriostatic action might have diminished on discontinuation of its usage.

Limitations

  • Larger sample of children with different caries status should be included to get a more precise evidence on the antibacterial properties of OSO
  • Long-term effects of ozone on the oral health status should be assessed
  • Some children did not like the unpleasant odor of OSO.



   Conclusion Top


  • In vitro evaluation of mouth rinses against S. mutans showed antibacterial activity for OSO and CHX mouthwash
  • All the three groups showed a significant reduction in DI-S, CI-S, PI, and S. mutans count in saliva after 15 days.


Thus, oil pulling using OSO can be used as an adjunct to regular oral hygiene measures. This can also be considered as an alternative to CHX mouthwash.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

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Marsh PD. Dental plaque as a biofilm and a microbial community – Implications for health and disease. BMC Oral Health 2006;6 Suppl 1:S14.  Back to cited text no. 1
    
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Krzyściak W, Jurczak A, Kościelniak D, Bystrowska B, Skalniak A. The virulence of Streptococcus mutans and the ability to form biofilms. Eur J Clin Microbiol Infect Dis 2014;33:499-515.  Back to cited text no. 2
    
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Mythri H. Oil pulling: A traditional method on the edge of evidence. Dent Hypotheses 2017;8:7-60.  Back to cited text no. 3
    
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Traina AA. Biological Effects of Ozone in Water on Dermal Wounds Healing in Rats Thesis. São Paulo: Faculty of Dentistry, University of Sao Paulo; 2008.  Back to cited text no. 9
    
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Verma SK, Deo B, Ranjan M, Prakash O, Shekhar M, Verma S. Comparison of efficacy of ozonated oil and chlorhexidine gel in the management of gingivitis. Int J Curr Res 2016;8:76-9.  Back to cited text no. 10
    
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Saravanan D, Ramkumar S, Vineetha K. Effect of oil pulling with sesame oil on plaque-induced gingivitis: A microbiological study. J Dent Orofacial Res 2013;3:175-80.  Back to cited text no. 11
    
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Segata N, Haake SK, Mannon P, Lemon KP, Waldron L, Gevers D, et al. Composition of the adult digestive tract bacterial microbiome based on seven mouth surfaces, tonsils, throat and stool samples. Genome Biol 2012;13:R42.  Back to cited text no. 12
    
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Jenkinson HF. Beyond the oral microbiome. Environ Microbiol 2011;13:3077-87.  Back to cited text no. 13
    
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Anand TD, Pothiraj C, Gopinath RM, Kayalvizhi M. Effect of oil-pulling on dental caries causing bacteria. Afr J Microbiol Res 2008;2:63-6.  Back to cited text no. 15
    
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Cruz O, Menendez S, Martinez ME, Clavera T. Application of ozonized oil in the treatment of alveolitis. In: Ozone in Medicine. 2nd ed. Cuba: International Symposium on Ozone Applications, Maret; 1997. p. 24-6.  Back to cited text no. 16
    
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Shapiro S. The inhibitory action of fatty acids on oral bacteria. Oral Microbiol Immunol 1996;11:350-5.  Back to cited text no. 17
    
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Wang G, Li W, Liu Y, Chen X, Huang J, Zhao Y, et al. Efficacy of dental health education and a novel mouthwash on periodontal health of navy personnel on a long ocean-going training mission. Int J Clin Exp Med 2016;9:53-60.  Back to cited text no. 18
    
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Asokan S, Emmadi P, Chamundeswari R. Effect of oil pulling on plaque induced gingivitis: A randomized, controlled, triple-blind study. Indian J Dent Res 2009;20:47-51.  Back to cited text no. 19
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