INTRODUCTION:

Dental caries and periodontal disease are the two major dental diseases caused by dental plaque. The young supragingival plaque consists of mainly gram negative and gram positive cocci and rods.^[1] Mechanical plaque control measures are usually inadequate in removing the dental plaque. Hence, chemical control measures like antibiotics and mouthwashes are recommended for the treatment process. The side effects of antibiotics, particularly antibiotic resistance, recommends the need for alternate solutions.

Allicin, obtained from the freshly crushed garlic cloves has been extensively investigated since ancient times for its antibacterial, antifungal and antiviral properties. Studies have shown that allicin is the potential inhibitory effect on the growth of various gram positive and gram negative bacteria such as Micrococcus, Enterobacter, Escherichia, Klebsiella, Lactobacilli, Pseudomonas, Salmonella, Shigella, Proteus, Helicobacter pylori, Staphylococcus, and Streptococcus species^[2-5]. Houshmand B et al (2013)^[6]conducted invitro study and report garlic extract is effective against oral pathogens such as Streptococcus mutans, Streptococcus sanguis, Streptococcus salivarius, Pseudomonas aeruginosa, and Lactobacillus spp.

This study was aimed to assess the antibacterial effect of garlic extract against human dental plaque pH. The hypothesis of this study is that the use of mouthwashes incorporated with allicin may resist the decrease in the plaque pH and thus prevent dental caries.

METHODOLOGY:

A double blinded randomized control study was conducted after obtaining ethical clearance from the Institutional Human Ethical Committee of Sathyabama Dental College and Hospital, Chennai. Participants were informed about the study and a written informed consent was obtained from each participant before the start of the study. Individuals who had a DMFT (Decayed Missing and Filled Tooth) score of 0, Community Periodontal Index (CPI) score of 0 and who signed the informed consent were alone included for the study. Individuals who had history of any systemic illness and who are under medication were excluded from the study.

The garlic mouthwash was prepared by diluting 1000ppm allicin liquid to 100ppm.^[4]The addition of water will stabilize the allicin and will extend its half life to about 30-40 days, allows allicin to be absorbed into the water, which would become highly antibiotic.^[7] The chlorhexidine rinse was prepared by diluting a commercially available chlorhexidine (Hexidine, ICPA Health Products Ltd) 50ppm mouthwash in 1:1 ratio with distilled water. The combination mouthwash was prepared with the allicin and chlorhexidine solution being equally mixed in 1:1 ratio. Both the solutions are known to possess antibacterial property and were mixed in equal proportions to assess whether they exhibited “synchronous” benefits.

90 participants took part in the study. The subjects who participated in the study were refrained from brushing and other oral hygiene measures. The study participants were then divided randomly into 3 groups by the chief investigator each group comprising of 30 participants: Group 1: allicin extract, Group 2: chlorhexidine mouthwash and Group 3: combination of both. The supra gingival plaque was collected from the maxillary posterior teeth and lingual surfaces of mandibular posterior teeth using spoon excavator.^[7] The plaque collected was diluted in 20ml of distilled water and baseline pH was estimated using a digital pH meter.

The participants in all the groups were then asked to rinse their mouth with glucose mouth rinse prepare by mixing 10grams of glucose with 100ml of distilled water. This glucose acts as a component of cariogenic challenge.^[7] The study participants and investigators who carried out the observation were then blinded. The participants were asked to swish and spit by using the mouthwashes. The plaque was collected and then mixed with 20ml of distilled water and the pH was esti mated. This procedure was repeated at regular five minute interval for half an hour.

The data collected was entered in Windows Excel 2010 and statistical analysis was done using version 20 SPSS software. ANOVA with Repeated Measures was used to compare the mean the three groups. Where ever there were significant shifts in mean in ANOVA result, the pairwise comparison was conducted between the groups using a Bonferroni post hoc test to justify which group significantly differed from the other. Level of significance was set as 0.05.

RESULT:

The table 1 shows a significant difference between the mean values of plaque pH within each group from baseline to 30 minutes. The post hoc test in table 2 shows a significant increase from baseline itself in all the groups. From table 3, its observed that there is a significant difference between the study groups from 5 minutes to 30 minutes. The pairwise comparison in table 4 shows a significant difference in all groups from 5 minutes in all groups. From graph 1 it is found that the mean dental plaque pH value of the combination group (allicin + chlorhexidine) forms a gradual increase in the linear pattern. The mean value of the combination group was found to be higher than other study groups.

Table 01: Comparison within the group using Repeated measures ANOVA.

Allicin Group			Chlorhexidine Group		Combination group
	Mean	S. D	Mean	S. D	Mean	S. D
Baseline	6.17	0.21	6.2	0.25	6.17	0.21
5	6.15	0.23	6.32	0.25	6.43	0.22
10	6.33	0.2	6.53	0.24	6.62	0.2
15	6.53	0.19	6.72	0.18	6.83	0.17
20	6.71	0.18	6.87	0.18	7.04	0.17s
25	6.93	0.13	7.08	0.16	7.26	0.15
30	7.01	0.26	7.17	0.26	7.47	0.08
F value	420.4		431.54		1454.3
p value	0.0001		0.0001		0.0001

Table 02: Post hoc comparison within the group at different time intervals.

	Group 1		Group 2		Group 3
	Mean difference	p value	Mean difference	p value	Mean difference	p value
Baseline vs 5 min	0.021	0.252	-0.147	0.0001*	-0.259	0.0001*
Baseline vs 10 min	-0.158	0.0001*	-0.353	0.0001*	-0.446	0.0001*
Baseline vs 15 min	-0.356	0.0001*	-0.544	0.0001*	-0.659	0.0001*
Baseline vs 20 min	-0.539	0.0001*	-0.698	0.0001*	-0.867	0.0001*
Baseline vs 25 min	-0.757	0.0001*	-0.906	0.0001*	-1.09	0.0001*
baseline vs 30 min	-0.842	0.0001*	-1.001	0.0001*	-1.3	0.0001*
5 min vs 10 min	-0.179	0.0001*	-0.207	0.0001*	-0.187	0.0001*
5 min vs 15 min	-0.377	0.0001*	-0.398	0.0001*	-0.4	0.0001*
5 min vs 20 min	-0.56	0.0001*	-0.551	0.0001*	-0.608	0.0001*
5 min vs 25 min	-0.778	0.0001*	-0.759	0.0001*	-0.831	0.0001*
5 min vs 30 min	-0.863	0.0001*	-0.854	0.0001*	-1.041	0.0001*
10 min vs 15 min	-0.198	0.0001*	-0.191	0.0001*	-0.213	0.0001*
10 min vs 20 min	-0.381	0.0001*	-0.344	0.0001*	-0.421	0.0001*
10 min vs 25 min	-0.599	0.0001*	-0.552	0.0001*	-0.644	0.0001*
10 min vs 30 min	-0.684	0.0001*	-0.648	0.0001*	-0.854	0.0001*
15 min vs 20 min	-0.183	0.0001*	-0.153	0.0001*	-0.208	0.0001*
15 min vs 25 min	-0.401	0.0001*	-0.361	0.0001*	-0.431	0.0001*
15 min vs 30 min	-0.487	0.0001*	-0.457	0.0001*	-0.641	0.0001*
20 min vs 25 min	-0.218	0.0001*	-0.208	0.0001*	-0.223	0.0001*
20 min vs 30 min	-0.303	0.0001*	-0.303	0.0001*	-0.433	0.0001*
25 min vs 30 min	-0.086	0.009*	-0.096	0.01*	-0.21	0.0001*

Table 03: Overall comparison between the group (group 1 vs group 2 vs group 3) at various time intervals using one way ANOVA

	F	Sig.
Baseline	0.056	0.946
min5	62.085	0.0001*
min10	92.8	0.0001*
min15	91.359	0.0001*
min20	117.012	0.0001*
min25	69.948	0.0001*
min30	96.631	0.0001*

Table 04: Post hoc comparison between the group at different time intervals

Time interval	Comparison group	Mean difference	p value
Baseline	Allicin group vs chlorhexidine group	0	1
	Allicin group vs combination group	0.01111	0.773
	Chlorhexidine group vs combination group	0.01111	0.773
5 min	Allicin group vs chlorhexidine group	-.37444*	0.0001*
	Allicin group vs combination group	-.28000*	0.0001*
	Chlorhexidine group vs combination group	.09444*	0.007*
10 min	Allicin group vs chlorhexidine group	-.38667*	0.0001*
	Allicin group vs combination group	-.28778*	0.0001*
	Chlorhexidine group vs combination group	.09889*	0.001*
15 min	Allicin group vs chlorhexidine group	-.34222*	0.0001*
	Allicin group vs combination group	-.30333*	0.0001*
	Chlorhexidine group vs combination group	0.03889	0.162
20 min	Allicin group vs chlorhexidine group	-.36667*	0.0001*
	Allicin group vs combination group	-.32778*	0.0001*
	Chlorhexidine group vs combination group	0.03889	0.141
25 min	Allicin group vs chlorhexidine group	-.24444*	0.0001*
	Allicin group vs combination group	-.33333*	0.0001*
	Chlorhexidine group vs combination group	-.08889*	0.003*
30 min	Allicin group vs chlorhexidine group	-.15889*	0.0001*
	Allicin group vs combination group	-.45778*	0.0001*
	Chlorhexidine group vs combination group	-.29889*	0.0001*

Graph 1: Mean comparison between the study groups

DISCUSSION:

Chlorhexidine mouthwash is a commonly used chemical substance for controlling the formation of dental plaque. Many studies have been done on herbal extracts to prevent plaque accumulation on the tooth surface as an alternative to chlorhexidine as Chlorhexidine has found to have various side effects. Moreover, many studies conducted using chlorhexidine have shown that its effect on reducing oral microflora is dose related. The bacterial counts returns to baseline levels within a few weeks depending on the dose.^[8,9]

Garlic, a member of the Allium family have gained focus among researchers because of its medicinal property and availability of the extract for use.^[10,14] Saxena G et al (2018)^[12]and . Kshirsagar MM et al (2018)^[13]conducted invitro studies and reports that Garlic extract has an anticariogenic effect against caries causing bacteria.

The present study analyzed the changes in dental plaque after using mouth washes containing allicin, chlorhexidine and combination of both. It was found allicin had an effect on dental plaque pH. But the pH increased more when it was used in combination with Chlorhexidine. Similar finding was reported by Kudva S et al (2012).^[7] The spicy flavor of garlic stimulates the saliva, which contains a higher concentration of bicarbonate ions. This bicarbonate ions increase the buffering capacity of saliva that neutralizes the acid substrates and increases the dental plaque pH.

CONCLUSION:

To conclude, garlic mouth wash is effective against dental plaque pH. However, the results were best obtained when used in combination with chlorhexidine.

ACKNOWLEDGEMENT:

The authors would like to acknowledge Dr. Mahesh Jagedeson, Department of Public Health Dentistry, Karpaga Vinayaga Institute of Dental Science, Chennai, for statistical assistance.

CONFLICT OF INTEREST:

Nil.

REFERENCES:

1. Hardie JM. Oral microbiology: Current concepts in the microbiology of dental caries and periodontal disease. Br Dent J 1992; 172:271‑8.

2. Fani MM, Kohanteb J, Dayaghi M. Inhibitory activity of garlic (Allium sativum) extract on multidrug-resistant Streptococcus mutans. J Indian Soc Pedodont Prev Dent. 2007; 25: 164 – 168.

3. Tsao SM, Yin MC. In vitro antimicrobial activity of four diallyl sulphides occurring naturally in garlic and Chinese leek oil. J Med Microbiol. 2001; 50: 646 – 649.

4. Groppo FC, Ramancciato JC, Simoes RP, Florio FM, Strtatoratto A. Antimicrobial activity of garlic, tea tree oil, and chlorhexidine against oral microorganisms. Int Dent J. 2002; 52: 443 – 447.

5. Zardast M, Namakin K, Kaho JE, Hashemi SS. Assessment of antibacterial effect of garlic in patients infected with Helicobacter pylori using urease breath test. Avicenna Journal of Phytomedicine. 2016 Sep;6(5):495.

6. Houshmand B, Mahjour F, Dianat O. Antibacterial effect of different concentrations of garlic (Allium sativum) extract on dental plaque bacteria. Indian Journal of Dental Research. 2013 Jan 1; 24(1):71.

7. Kudva S, Prabhakar S, Pai V, Tegginamani A. Effects of garlic extract on salivary pH: A clinical study. Arch Orofac Sci. 2012 Jan 1;7(1):1-8.

8. Ribeiro LG, Hashizume LN, Maltz M. The effect of different formulations of chlorhexidine in reducing levels of mutans streptococci in the oral cavity: A systematic review of the literature. Journal of Dentistry. 2007 May 1;35(5):359-70.

9. Hassan SM, Mobarak EH, Fawzi EM. The efficacy of different regimens of chlorhexidine as an antimicrobial agent for a group of Egyptians. J Egypt Public Health Assoc. 2008 Feb;83(5-6):435-50.

10. Den YY, Chiu HC, Wang YB. Effects of Garlic Extract on Acid Production and Growth of Streptococcus mutans. Journal of Food and Drug Analysis. 2009 Feb 1;17(1):59-63.

11. Lee CK, Schmitz B. Determination of pH, total acid, and total ethanol in oral health products: Oxidation of ethanol and recommendations to mitigate its association with dental caries. Journal of dentistry, Oral Medicine and Dental Education. 2009;3(1):1-4.

12. Saxena G, Sadawarte K, Kaore NM. Antibacterial activity of aqueous extract of garlic (Allium sativum) on standard strains. Journal of Evolution of Medical and Dental Sciences. 2018 May 7;7(19):2320-2.

13. Kshirsagar MM, Dodamani AS, Karibasappa GN, Vishwakarma PK, Vathar JB, Sonawane KR, Jadhav HC, Khobragade VR. Antibacterial activity of garlic extract on cariogenic bacteria: An in vitro study. Ayu. 2018 Jul;39(3):165.

14. Johnson M, Olaleye ON, Kolawole OS. Antimicrobial and antioxidant properties of aqueous garlic (Allium sativum) extract against Staphylococcus aureus and Pseudomonas aeruginosa. Microbiology Research Journal International. 2016 Mar 29:1-11.

Received on 15.04.2020 Modified on 29.05.2020

Research J. Pharm. and Tech. 2021; 14(2):977-980.

DOI: 10.5958/0974-360X.2021.00174.8