Open Access Original

Article DOI: 10.7759/cureus.53484

Miswak-Infused Glass Ionomer Cement: A

Comparative In Vitro Analysis of Antibacterial
Received 09/24/2023
Efficacy and Compressive Strength
Review began 01/09/2024
Review ended 01/31/2024 Kamala Devi 1, Jessy Paulraj 1 , Rajeshkumar Shanmugam 2, Subhabrata Maiti 3
Published 02/03/2024

© Copyright 2024 1. Pediatric Dentistry, Saveetha Dental College and Hospitals, Saveetha Institute of Medical and Technical Sciences,
Devi et al. This is an open access article Chennai, IND 2. Pharmacology, Saveetha Dental College and Hospitals, Saveetha Institute of Medical and Technical
distributed under the terms of the Creative Sciences, Chennai, IND 3. Prosthodontics, Saveetha Dental College and Hospitals, Saveetha Institute of Medical and
Commons Attribution License CC-BY 4.0., Technical Sciences, Chennai, IND
which permits unrestricted use, distribution,
and reproduction in any medium, provided
the original author and source are credited. Corresponding author: Jessy Paulraj, jessyp.sdc@saveetha.com

Abstract
Background: Glass ionomer cement (GIC) restorations are commonly used in primary dentition, due to their
aesthetic appeal, self-adhesive nature, and biocompatibility. However, the material's limited antibacterial
activity and inadequate mechanical strength highlight the necessity for modifying the material.

Aim: The study aims to evaluate and compare the antimicrobial potency and compressive strength of GIC-
incorporated Miswak extract with that of conventional GIC.

Materials and methods: After obtaining the Miswak extract, a modified GIC was formulated by combining
the extract with the conventional GIC powder and liquid components, in three different ratios (Powder:
Extract and Liquid), Group I (2:1:1), Group II (3:1:2), Group III (3:2:1), and the Group IV as control, which
consist of unmodified/conventional GIC. To evaluate and compare the antibacterial efficacy of the modified
and unmodified GIC, standard strains of Streptococcus mutans and Lactobacillus were utilized. For each
group, the minimal inhibitory concentration (MIC) assay was tested. For the evaluation of compressive
strength, cylindrical moulds were utilized in compliance with ISO 9917-1:2007 standards and tested using
the universal testing machine (Instron, ElectroPuls®, Bangalore, IND). The highest force exerted at the point
of specimen fracture was recorded to calculate the compressive strength values in MPa. The data obtained
were analyzed using the Statistical Package for the Social Sciences
(IBM SPSS Statistics for Windows, IBM Corp., Version 24.0, Armonk, NY) software. The statistical analysis
was conducted utilizing repeated measures of analysis of variance (ANOVA) to calculate the mean
MIC values and compressive strength, with pairwise comparisons assessed using Tukey's post hoc test.

Results: The results proved that the antimicrobial properties of Miswak containing GIC performed better
against S. mutans and Lactobacillus with a statistically significant difference when compared with group IV
(p<0.05), it has been found that an increase in the concentration of extract increased the antimicrobial
potency. Significant results were obtained in compressive strength where Group II (41.49±3.6) and Group III
(15.23±4.96) proved to be weaker than the control (62.69±2.58), while Group I showed no differences from
the control group (p>0.05).

Conclusion: It can be concluded that Group I was found to be better in terms of both antimicrobial
properties and compressive strength, where no significant difference in compressive strength was identified
when comparing Group I with Group IV. Thus, the overall study depicts that a lesser concentration of extract
can be the best option in terms of good antimicrobial properties without altering its strength. Hence, the
Miswak containing GIC could be a promising restorative material; further studies should include considering
intraoral variables such as masticatory stress, moisture levels and in-vivo tests of this combination.

Categories: Dentistry
Keywords: compressive strength, restorative material, modified gic, antimicrobial, miswak

Introduction
Glass ionomer cement (GIC) materials have a five-decade history in clinical applications. Over time, these
materials have undergone modifications to serve various purposes, including lining, bonding, sealing,
repairing, and restoring teeth. Although GIC is recognized for its features like fluoride release, tooth colour
matching, and adhesion to dental structures, it has drawbacks including sensitivity to moisture, limited
antimicrobial activity, prolonged wear problems, and inadequate strength, which may impede its
effectiveness [1]. Hence, in situations such as insufficient remaining tooth structure to support material or
the inability to withstand heavy occlusal loads, which affect the prognosis, GIC is strictly not recommended.
The past decades have shown that the use of these materials is expanding; however, they may hold specific
niches of clinical use. Caries is a complex and ever-changing dental condition that arises from a combination

How to cite this article

Devi K, Paulraj J, Shanmugam R, et al. (February 03, 2024) Miswak-Infused Glass Ionomer Cement: A Comparative In Vitro Analysis of
Antibacterial Efficacy and Compressive Strength. Cureus 16(2): e53484. DOI 10.7759/cureus.53484
 of factors, including the presence of biofilm, sugar consumption, and the continuous processes of
demineralization and remineralization in hard tooth tissue [2]. Dental decay affects both primary and
permanent teeth, regardless of age. The initial process involves enamel demineralization, followed by pulp
weakening, ultimately resulting in the loss of crown structure. The balance of protective and pathological
factors influences the development and progression of caries [3].

Recurrent caries is the most common reason for replacing restorations of all kinds in general dental practice.
Prevention of recurrent lesions by using fluoride-releasing restorative materials has been successful.
Because recurrent carious lesions are localized and limited, alternative treatments to restorations such as
crowns have been proposed, but this is not practical in all cases [4]. It is not advised to intentionally lose
tooth structure. Failure to establish a complete seal in the restored tooth enables the persistence of
cariogenic bacteria, leading to the recurrence of caries and ultimately resulting in restoration failure [5]. In
the case of a minimally interventional approach (ART) to treat caries, recurrent caries have been reported
more as cariogenic bacteria get trapped beneath GIC restorations due to manual removal of demineralized
tooth tissue using hand instruments. Studies have shown that ART restoration rarely fails after six years due
to the development of secondary caries. Hence, an effective way to tackle this challenge is by improving
restorative materials that can offer strong hermetic seals and exhibit superior antimicrobial capabilities.
Consequently, combining antibacterial agents with GIC might provide a therapeutic benefit. Nevertheless,
it's important to note that the inclusion of antimicrobial agents in restorative materials often has an adverse
impact on the physical and chemical properties of these materials [6]. Thus, assessing the compressive
strength alongside the antibacterial effects of modified GIC is of significant importance.

There have been various reported endeavours to enhance the antibacterial capabilities of GICs by combining
them with chlorhexidine hydrochloride, cetylpyridinium chloride, cetrimide, and benzalkonium chloride [7].
According to the literature, only chlorhexidine was widely incorporated into GIC, with all studies showing
increased antibacterial activity in vitro [8], but it was not marketed due to chemical agents that can act on
the cells of the pulp or the gingiva; hence, this study aimed to modify it with the bioactive component. The
use of natural and herbal materials in dentistry is growing in popularity. The Salvadora persica tree, which is
frequently used to make miswaks, is one of the most widely utilized materials. East Asia and West Africa are
major distribution areas for S. persica. The term "Miswak" refers to a group of wooden sticks originating from
diverse plant sources. Extracts from some of these plants have been utilized in the production of mouthwash
and toothpaste [9]. S. persica extract demonstrated substantial antibacterial effectiveness against various
oral pathogens, which encompassed Candida albicans, Streptococcus mutans, Aggregatibacter
actinomycetemcomitans, Lactobacillus acidophilus, Actinomyces naeslundii, and Porphyromonas gingivalis [10].
In the realm of oral health, S. persica is known for its antibacterial, antifungal, anticariogenic, and
antiplaque properties [11,12]. Miswak contains fluoride, phosphate, calcium and similar minerals found in
the hydroxyapatite crystal of teeth [13]. This suggests its potential to aid in dental caries restoration; hence,
this study was designed to modify GIC with Miswak extract and to assess both the antibacterial effectiveness
and physical characteristics of Miswak-modified GIC.

Materials And Methods

Extract preparation
Miswak sticks were procured from Annai Aravindh Herbals Pvt Ltd., Chennai, India. After undergoing a five-
day drying process, the Miswak sticks were utilized to prepare a mixture in a beaker, combining 100 mL of
distilled water with 6 g of finely chopped Miswak sticks. This mixture was heated using a heating mantle,
bringing it to a boil and maintaining a temperature between 60 and 70 degrees Celsius for a duration of 15
minutes. After filtration through Whatman No. 1 filter paper (Whatman Plc, Maidstone, UK), 80 ml of
filtrate was collected in a separate Erlenmeyer flask, and this filtered extract was subsequently concentrated
to a volume of 5 ml.

Specimen preparation
Type II GIC (GC Corporation) was utilized in this research study. Groups I, II, and III represent Miswak-
modified GIC groups. In these groups, Miswak-modified GIC specimens were created in three different
concentration ratios characterized by a powder GIC: Extract: Liquid GIC with Group I (2:1:1), Group II (3:1:2)
and Group III (3:2:1). Group IV serves as the control group without any modification, employing
conventional GIC. First, the conventional GIC components were mixed, and then the plant extract was added
as per the groups. The resulting cement was placed in 6 mm-diameter, 2 mm thick cylindrical moulds, and
samples were transferred to cylindrical wells. After application, disc-shaped samples were removed,
measured, and recorded. Each group comprised 12 samples, six for S. mutans testing and the remaining six
for Lactobacillus. Bacterial resistance was evaluated against S. mutans and Lactobacillus strains. Compressive
strength, as per ISO 9917-1:2007 (4.0 mm diameter, 6.0 mm height), cylindrical moulds were prepared with
12 samples in each group. After an hour, samples were soaked in deionized water for 24 hours before testing.

Preparation of bacterial species and inoculation

S. mutans and L. acidophilus strains were acquired from the Department of Microbiology. After obtaining
pure cultures, facultative strains were cultivated on Mueller-Hinton Agar (MHA), then subculture was

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 2 of 9

 separately introduced into tubes with 5 mL of sterile MHA broth. These tubes were incubated at 37°C for 24
hours, and the resulting suspension was subsequently adjusted to a 0.5 McFarland standard.

Minimal inhibitory concentration (MIC) assay

To evaluate the antimicrobial effectiveness of both modified and unmodified GIC, a total of 48 specimens,
involving 12 samples per group, were used, with each well containing 200 µl of sterilized MHA broth. Then,
50 µL of bacterial suspensions at a concentration of 5 x 10^5 CFU/ml were added to the 24 wells (S. mutans)
and the remaining for L. acidophilus. As per the groups, modified specimens containing different GIC
concentrations (2:1:1), (3:1:2), and (3:2:1) and unmodified specimens were added (Figure 1).

FIGURE 1: MIC assay in ELISA 96-well plate

A - Streptococcus mutans, B - Lactobacillus

Incubation was carried out under suitable conditions for varying time intervals (1h, 2h, 3h, 4h). The
percentage of deceased cells was measured at 540 nm using an ELISA reader during specific time intervals.

Assessment of compressive strength

The diameter of each sample was assessed using a digital micrometre before placing them vertically in a
universal testing machine (Instron, ElectroPuls®, Bangalore, IND). At a crosshead speed of 0.5 mm/min, a
compression load was applied to the specimen's long axes until fracture, and readings were recorded.

Statistical evaluation
The data were input into Microsoft Excel (Microsoft® Corp., Redmond, WA) and analyzed using the
Statistical Package for the Social Sciences (IBM SPSS Statistics for Windows, IBM Corp., Version 24.0,
Armonk, NY) software. Descriptive analysis and repeated measures of analysis of variance (ANOVA) were

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 3 of 9

 performed to calculate the mean MIC values, comparing all the groups based on several time intervals.
Tukey's post hoc test was assessed for pairwise comparison between groups. Compressive strength was
assessed with repeated measures ANOVA to compare the groups, followed by Tukey's post hoc test (P < 0.05,
95% confidence) for pairwise comparison.

Results
Antimicrobial efficacy against S. mutans
Repeated measures of ANOVA tested modified and unmodified GIC's antibacterial activity against S. mutans.
Group I, Group II, and Group III exhibited superior performance and statistically significant results
compared to Group IV (control) (Figure 2). Also, Tukey's honestly significant difference (HSD) pairwise
comparison test showed a significant difference between Group IV and the other three groups (Table 1).

FIGURE 2: Antimicrobial efficacy on Streptococcus mutans between

four groups
MIC: minimal inhibitory concentration

95% CI
Pairwise comparison Mean difference P-value
Lower Upper

Group I vs Group II 0.2983 0.027 0.032 0.001*

Group I vs Group III 0.046 0.044 0.049 0.001*

Group I vs Group IV 0.241 0.239 0.244 0.001*

Group II vs Group III 0.169 0.014 0.019 0.001*

Group II vs Group IV 0.271 0.269 0.273 0.001*

Group III vs Group IV 0.288 0.286 0.290 0.001*

TABLE 1: Pairwise comparison of antimicrobial efficacy on Streptococcus mutans between four

groups
*P value was significant at 0.05, P value was derived from multiple comparisons of Tukey's honestly significant difference (HSD) test.

Antimicrobial efficacy against Lactobacillus

For Lactobacillus, a repeated measure of ANOVA demonstrated that the modified groups exhibited notable

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 4 of 9

 antimicrobial activity, displaying superior performance compared to the control group (Figure 3). Pairwise
comparisons for Lactobacillus revealed a significant difference (p<0.05) between Group IV and the other
groups (Table 2), highlighting the pronounced antibacterial activity of Miswak-modified GIC over
conventional GIC.

FIGURE 3: Antimicrobial efficacy on Lactobacillus between four groups

MIC: minimal inhibitory concentration

95% Confidence interval

Pairwise comparison Mean difference P-value
Lower Upper

Group I vs Group II 0.030 0.029 0.031 0.001*

Group I vs Group III 0.146 0.144 0.147 0.001*

Group I vs Group IV 0.188 0.186 0.189 0.001*

Group II vs Group III 0.176 0.175 0.177 0.001*

Group II vs Group IV 0.157 0.156 0.158 0.001*

Group III vs Group IV 0.334 0.332 0.335 0.001*

TABLE 2: Pairwise comparison of antimicrobial efficacy on Lactobacillus between four groups

*P value was significant at 0.05, P value was derived from multiple comparisons of Tukey's honestly significant difference (HSD) test.

Compressive strength
The compression load was applied to the specimens, and values were recorded. A one-way ANOVA showed a
statistically significant difference between all groups with a p-value of 0.001 (Table 3). The pairwise
comparison revealed Group II and Group III differed significantly from Group IV, with Group IV (control)
showing better performance in compressive strength; however, there was no significant difference between
Group I and Group IV (control), indicating similar performance of these groups with a p value of 0.995 (p >
0.05) (Table 4).

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 5 of 9

 95% CI
Group n Mean ± SD P value
Lower Upper

Group I 12 62.51±5.54 58.99 66.03

Group II 12 41.49±3.6 39.20 43.78

0.001*
Group III 12 15.23±4.96 12.07 18.38

Group IV 12 62.69±2.58 60.44 63.73

TABLE 3: Comparison between groups for evaluation of compressive strength

*Significant at 0.05, P value was derived by one-way analysis of variance (ANOVA).

95% CI
Pairwise comparison Mean difference P-value
Lower Upper

Group I vs Group II 21.025 16.30 25.74 0.001*

Group I vs Group III 47.284 42.56 52.00 0.001*

Group I vs Group IV 0.425 4.29 5.14 0.995

Group II vs Group III 26.258 21.53 30.97 0.001*

Group II vs Group IV 20.600 15.87 25.32 0.001*

Group III vs Group IV 46.85 42.13 51.57 0.001*

TABLE 4: Pairwise comparison for evaluation of compressive strength

*significant difference at p<0.05, P value was derived from Tukey Post hoc test.

Discussion
Although dental caries treatments do not always completely eliminate all microorganisms, some remain in
residual tissue. When restorations lack effective hermetic seals, the continued presence of cariogenic
bacteria can result in recurrent caries and, consequently, the failure of the restoration [14]. To address this
challenge, one potential solution is the adoption of dental materials that exhibit properties capable of
inhibiting and eradicating bacteria, both bacteriostatic and bactericidal in nature. Introduced by Wilson and
Kent in 1972, conventional GICs are tooth-coloured, chemically bonded materials that are inherently anti-
caries and contain fluoride, a widely used compound in dentistry. It has a therapeutic effect through its
release properties. The ability of GICs to deliver fluoride continuously over an extended period of time offers
anti-caries potential that indicates a reduction in caries adjacent to the restoration [15], but again, it cannot
inhibit a broad spectrum of microorganisms. Also, cavities treated with ART (atraumatic restorative
treatment) may retain infected dentin; thus, GIC proves ineffective in halting the progression of caries,
resulting in restoration failure; hence, improving restorative materials to enhance the physical and
antibacterial attributes of conventional GIC is necessary. Plant-based phytochemicals are gaining
prominence as an alternative to commercial antimicrobial agents. They offer additional benefits, such as
traditional medicinal use and cost-effectiveness, while avoiding issues of antibacterial resistance. One
example is Miswak, an herbal antimicrobial agent extracted from the S. persica plant.

The current results of the study prove the superior antimicrobial effectiveness of Miswak-modified GIC
against S. mutans. These findings are consistent with the work done by Sofrata et al. who reported on S.
persica's antibacterial properties against oral microorganisms, including S. mutans and L. acidophilus [16]. An
in-vivo study by Kabil NS et al. revealed that the incorporation of Miswak into GIC led to superior
antibacterial properties when compared to standard GIC, aligning with our current in vitro findings [17].
Furthermore, a prospective case-control observational study involving Miswak users and non-users revealed
a significantly higher prevalence of dental caries among non-users. This study, conducted on 240
schoolchildren over two years, assessed the decayed-missing-filled (DMF) index. The lower incidence of
dental caries among Miswak users suggests its potential role in caries prevention [18]. In another study by

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 6 of 9

 Lamia Singer et al., it was observed that blending plant extracts like S. persica, Olea europaea, and Ficus
carica with GIC resulted in increased antimicrobial activity against S. mutans and Micrococcus luteus,
particularly at higher concentrations [19]. The Miswak extract possesses a range of antimicrobial and
antifungal attributes attributed to its content of chlorides, trimethylamine, fluoride, silica, saponins, sulfur,
flavonoids, and phenols. Its b-sitosterol and m-anisic acid both contribute to its antibacterial activity [19]. A
study conducted by El Tatari et al. showed that Salvadoran Persica Extract (SPE) combined with GIC showed
promising results in antimicrobial tests, especially against S. mutans [20]. Kalpavriksha AJ et al. found that
GIC with 1% CHX and Miswak extract was equally effective against S. mutans and Streptococcus sobrinus [21].
Ashouri et al. study showed that adding lyophilized Miswak to GIC improved its antimicrobial efficacy [22].
One of the studies investigated S. persica capacity to prevent collagen breakdown in demineralized dentin,
where an aqueous S. persica extract was applied to demineralized bovine root dentin samples, which under a
light microscope revealed that the extract shielded the dentin's collagen matrix from enzymatic degradation
by collagenase. This proves a potential role for S. persica in caries prevention [23]. Apart from antimicrobial
properties, this current study evaluated the compressive strength of Miswak-modified GIC.

The performance of a material in therapeutic contexts is dependent upon its capacity to bear pressure,
stress, and strain. Compressive strength is the primary measure for characterizing dental cement. Hence, it
was imperative to evaluate the compressive strength when modifying GIC. The compressive strength test
took place after a 24-hour storage period, providing an optimal timeframe for testing its mechanical
properties. A study by El Tatar showed that SPE combined with GIC at a concentration of 1% SPE maintained
the same level of compressive strength as that of the control group [20], and this is consistent with the
present study, where a lower concentration of the extract, i.e., Group I (2:1:1), unaltered the compressive
strength. Farret et al. [24], Pavithra et al. [25], and Marti et al. [26] also noted similar results, indicating that
the addition of antimicrobial agents at particular concentrations had no impact on the compressive strength
characteristics of GIC.

Hence, the clinical significance of GIC infused with Miswak extract is noteworthy, primarily due to its
potential effectiveness in combating S. mutans and Lactobacillus, pivotal contributors to caries development,
including secondary caries. By incorporating this bacteriostatic agent, it becomes possible to impede caries
progression and mitigate the risk of restoration failure. This approach could prove valuable in the treatment
of patients with deep caries, early childhood caries, rampant caries, and high caries susceptibility. A
limitation of the present study is its omission of intraoral variables such as masticatory stress, moisture
levels, and potential discrepancies by operators. Further extensive research is required in regard to the
bonding effects of GIC material's long-term stability and performance, which can promote a novel natural
bioactive restorative material.

Conclusions
The findings of the present study suggested that a lower concentration of Miswak extract has the potential
to enhance antimicrobial properties without compromising the material's strength. Therefore, Miswak
containing GIC can be a promising restorative material. Further studies are recommended to investigate
setting time, fluoride release, and in vivo behaviour.

Additional Information
Author Contributions
All authors have reviewed the final version to be published and agreed to be accountable for all aspects of the
work.

Concept and design: Jessy Paulraj, Kamala Devi, Rajeshkumar Shanmugam

Acquisition, analysis, or interpretation of data: Jessy Paulraj, Kamala Devi, Rajeshkumar Shanmugam,
Subhabrata Maiti

Drafting of the manuscript: Jessy Paulraj, Kamala Devi

Critical review of the manuscript for important intellectual content: Jessy Paulraj, Kamala Devi,
Rajeshkumar Shanmugam, Subhabrata Maiti

Supervision: Jessy Paulraj, Rajeshkumar Shanmugam, Subhabrata Maiti

Disclosures
Human subjects: All authors have confirmed that this study did not involve human participants or tissue.
Animal subjects: All authors have confirmed that this study did not involve animal subjects or tissue.
Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the
following: Payment/services info: All authors have declared that no financial support was received from
any organization for the submitted work. Financial relationships: All authors have declared that they have

2024 Devi et al. Cureus 16(2): e53484. DOI 10.7759/cureus.53484 7 of 9

 no financial relationships at present or within the previous three years with any organizations that might
have an interest in the submitted work. Other relationships: All authors have declared that there are no
other relationships or activities that could appear to have influenced the submitted work.

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