Introduction

Dental aesthetics, particularly tooth color, is fundamental in modern perceptions of health and attractiveness. Discoloration of enamel due to extrinsic factors, such as diet, smoking, and poor oral hygiene, or intrinsic factors, like aging and trauma, remains a primary reason for cosmetic dental consultations [1]. To address this concern, the demand for accessible and effective whitening agents has surged, especially among consu­mers seeking over-the-counter options, such as toothpaste and mouthwash [2].

Tooth color significantly influences the aesthetic harmony of smile, and patients increasingly pursue dental treatments to achieve whiter teeth [3]. While professional
bleaching yields expressive results, it can sometimes cause side effects, including morphological changes and altera­tions in enamel properties, such as microhardness and surface roughness [4]. As an alternative, the dental industry offers whitening toothpastes, which are considered more cost-effective and user-friendly method for achieving whiter teeth, promising results within a few weeks of continuous use [5]. These products are widely available over-the-counter and do not require dental appointment [6].

Whitening toothpastes typically contain abrasive components capable of removing extrinsic stains that contribute to tooth discoloration [7]. More recently, activated charcoal has become a popular ingredient in whitening toothpastes. Activated charcoal is a carbon-rich material known for its whitening effect, often attributed to its abrasive action in removing surface stains and plaque [8]. However, there is debate and insufficient scientific evidence regarding activated charcoal’s effectiveness in changing the tooth color, or its safe application on the enamel surface [8].

Besides abrasives and activated charcoal, some whitening toothpastes contain low concentrations of hydrogen peroxide, typically ranging from 1.5% to 6% [9]. The whitening mechanism of hydrogen peroxide involves releasing reactive oxygen particles, which diffuse through the enamel and oxidize chromogens [10]. Furthermore, it was suggested that the whitening potential of many toothpastes, even those containing hydrogen peroxide, is primarily due to their abrasive features [11].

Enamel microhardness is a critical property reflecting the integrity and resistance of the tooth surface to mechanical and chemical challenges. Preserving micro­hardness is essential for maintaining oral health and structural integrity of dental tissues, as it directly relates to the tooth’s ability to resist masticatory forces and other mechanical challenges [11]. Whitening agents, including abrasive components found in toothpastes and oxidative agents, such as hydrogen peroxide, have the potential to alter the enamel surface’s properties, including microhardness [12]. For instance, hydrogen peroxide, an oxidative agent, can potentially induce demineralization due to its pH, or affect the organic matrix of enamel, thus compromising its structural integrity and microhardness [13].

Objectives

The objective of this study was to comparatively evaluate the effect of a whitening toothpaste containing 2% hydrogen peroxide versus one containing activated charcoal on enamel color and surface microhardness. The null hypothesis was that there is no statistically significant difference between these two agents in terms of their effects on enamel color change and surface microhardness.

Material and methods

Material

For this study, three commercial toothpastes were used: Colgate Optic White Advanced® (2% hydrogen peroxide), Colgate Luminous White Activated Charcoal® (activated charcoal-based), and Sensodyne Repair & Protect (non-whitening fluoride toothpaste) as a control. The specifications of materials used in the study are detailed in Table 1.

Sample size calculation

The research used 60 specimens with a sample size of 20 in each group. According to prior study by Sultan [10], a sample size of 20 within every group and a significance a level of 0.05 (two-tailed) at a 95% confidence interval with 80% power to identify a variation among averages of 18.84, were considered. The findings will be regarded as statistically significant if a p-value is below 0.05 (two-tailed) in 80% (the power) of studies. The variation in averages in the rest of the 20% of studies will be labeled as not statistically significant. StatMate 2.0 software from GraphPad was used for all analyses.

Teeth selection

Sixty sound human premolars were selected after obtaining permission from the faculty of Dental Medicine’s Ethics Committee of the Al-Azhar University, Cairo, Egypt, using reference authorization code of 1199/1143. Teeth were extracted due to periodontal disease in diabetic patients. Inclusion criteria were: (1) intact teeth; (2) non-caries teeth; (3) premolars with complete root formation; and (4) free from any developmental and formative anomalies. Exclusion criteria were: (1) carious teeth; (2) presence of previous restorations, fractures, or cracks; (3) prior endodontic treatments; and (4) presence of non-carious lesions (attrition, fluorosis).

Sample preparation

The selected teeth were cleaned of soft tissue debris and stored in 0.1% thymol solution at 4°C until further use. The crowns were separated from the roots 2 mm below the cementoenamel junction using a diamond disc under water cooling. Enamel slabs measuring 6 mm × 6 mm × 3 mm were obtained from the buccal surface (Figure 1). Enamel surfaces were sequentially polished with silicon carbide papers (#600, #1200, and #2000 grit) and diamond suspension (1 µm) to standardize surface smoothness. The specimens were ultrasonically cleaned for 10 minutes and stored in distilled water at 37°C before testing.

Grouping

The specimens were randomly assigned into the following three groups (n = 20 per group):

- group HP: samples brushed with toothpaste containing 2% hydrogen peroxide (Colgate Optic White Advanced®);

- group AC: specimens brushed with activated charcoal-based toothpaste (Colgate Luminous White Activated Charcoal®);

- group C: samples brushed with a non-whitening fluoride toothpaste (Sensodyne Repair & Protect) as a control.

Toothbrushing protocol

All groups underwent simulated toothbrushing using a mechanical brushing machine (MEV-4-10XY, Odeme Dental Research, Brazil), with a load of 200 g and 30,000 brushing cycles (150 cycles/min), simulating approximately 2 years of clinical brushing, as validated in a previous in vitro study using 30,000 brushing cycles [14]. A toothpaste slurry (1 : 3 ratio of toothpaste to distilled water by weight) was prepared freshly before each brushing session [14].

Color measurement

Color was measured at baseline and after brushing regimen using a VITA Easyshade Compact spectrophotometer (VITA Zahnfabrik, Germany), calibrated before each measurement. Three readings were taken per spe­cimen under standardized lighting conditions and averaged to ensure reliable color measurement [15]. The CIE L*a*b* color system was used to calculate color change (ΔE) using the following formula [16]:

ΔE = √ [(ΔL)² + (Δa*)² + (Δb*)²].

Microhardness testing

Knoop microhardness measurements were performed before and after 14-day intervention using a microhardness tester (HMV-2, Shimadzu Corporation, Japan). A load of 25 g was applied for five seconds per indentation, with five indentations made per spe­cimen at nonoverlapping sites, and mean value was recorded [17].

Statistical analysis

Data were analyzed using SPSS version 25.0 (IBM Corporation, Armonk, NY, USA). Normality was assessed using Shapiro-Wilk test. Intragroup comparisons were performed using paired t-tests, and intergroup comparisons were analyzed using one-way ANOVA, followed by Tukey’s post hoc test. Results were considered statistically significant if p-value was less than 0.05 (p < 0.05).

Results

Color change (ΔE)

The mean color change values (ΔE) for each group are presented in Table 2 and Figure 2. All groups demonstrated statistically significant color improvement after 14 days of treatment. Group HP (2% hydrogen pero­xide toothpaste) showed the highest mean color change (ΔE = 6.72 ± 1.84), significantly greater than both activated charcoal group (ΔE = 5.86 ± 3.66) and control group (ΔE = 4.12 ± 2.03) (p < 0.05). The differences between group AC and group C were also statistically significant (p < 0.05).

Microhardness

The results of the enamel microhardness tests before and after brushing protocol are summarized in Table 3 and Figure 3. In group HP (2% hydrogen peroxide toothpaste), there was a statistically significant reduction in microhardness values from baseline (287.45 ± 32.18) to after brushing (251.23 ± 28.94) (p = 0.012). In contrast, both activated charcoal (AC) and control group (C) revealed no statistically significant changes in microhardness values (p = 0.634 and p = 0.758, respectively).

Discussion

Polyphenolic chemicals’ adsorption upon the enamel surface and their interactions with pellicle proteins are the primary causes of extrinsic discoloration [18], with food and drink containing the majority of these organic chromogens. Because it produces quick and noticeable effects without weakening the tooth structure, teeth whitening has grown in popularity and demand in recent years. However, there may be negative consequences from this approach, such as morphological alterations in microstructure, microhardness, and surface roughness of the enamel [19].

The dental industry has provided an alternative to lessen or even eliminate these risks with a variety of whitening toothpaste formulations. These are regarded as a class of easy and less costly homemade remedies for people who want whiter teeth, with promised results after two to four weeks of consistent use [20]. Abrasive ingredients that can aid in the removal of external stains are present in some of these toothpastes, while hydrogen peroxide is present in small amounts in others [21]. In order to obtain whiter teeth, the utilization of whitening toothpastes that include activated charcoal has grown in popularity [8].

Furthermore, small variations in surface enamel microstructure can be detected using a microhardness assessment, which are associated with minerals being lost (demineralization) or regained (reminera­lization) [22]. To determine if whitening toothpaste chemicals are harmful to enamel, surface microhardness testing is frequently employed [22-24]. It is a straightforward, quick, easy, nondestructive, and reliable test for evaluating demineralization and reminera­lization alterations affecting the whitened teeth. It assesses the material’s resistance to plastic deformation using a standard source. Additionally, it reduces experimental variance by allowing the same sample to be examined multiple times [25].

The present study compared the effects of two over-the-counter whitening toothpastes, i.e., one containing 2% hydrogen peroxide and the other containing activated charcoal, on enamel color and microhardness over a 14-day simulated brushing protocol. Significant differences were observed in both whitening efficacy and enamel microhardness, with important clinical implications.

Regarding color change among the groups, the hydrogen peroxide toothpaste exhibited the highest color change, significantly greater than that of both the activated charcoal and control groups. This finding may be attributed to the fact that the small amount of hydrogen peroxide (2%) can whiten teeth by releasing reactive oxy­gen molecules. The long chain, dark-colored complex chromophore molecules are separated into smaller ones by these low molecular weight molecules as they diffuse across interprismatic gaps, producing the intended color shift that results in effective whitening [10, 25].

This result is consistent with prior research [26-28], showing that a more pronounced and tolerable color shift can be achieved with lower peroxide concentrations. On the other hand, this finding disagree with Favaro et al.’s [29], who found that low hydrogen pero­xide concentrations in bleaching treatments may restrict their clinical efficacy.

The activated charcoal group showed a moderate color change (although lower than hydrogen peroxide), and it was still statistically significant compared to the control group. This may be explained by the fact that activated charcoal is known for its high adsorptive capa­city, allowing it to bind to surface stains and pigments on the teeth. Its whitening effect is primarily due to mechanical removal of extrinsic stains rather than chemical alteration of intrinsic tooth color [3]. Moreover, because of its great surface area and high porosity, activated charcoal effectively and gradually cleans the teeth [30].

Systematic review [12] has indicated that activated charcoal-based toothpastes have a lower whitening effect compared to other agents, such as hydrogen peroxide. Also, Vaz et al. [30] reported that the whitening impact of an activated charcoal toothpaste was statistically lower than that of a toothpaste containing hydrogen peroxide. However, other studies contradict this finding. For instance, Ribeiro et al. [31] described limited or nonsigni­ficant whitening with charcoal-containing products, attri­buting its action mainly to surface stain removal without deeper chromophore oxidation. This discrepancy may be due to differences in methodology; our study was in vitro in nature, while Ribeiro et al.’s [31] was a clinical trial.

The microhardness results revealed a significant reduction in enamel hardness in the hydrogen pero­xide group. This may be because pH of the hydrogen peroxide-based toothpaste is lower than that of other toothpastes. A lower pH causes enamel softening by allowing hydroxyapatite to dissolve [32]. Long-term exposure to acidic materials on the tooth surface can result in dental erosion, a chemical reaction leading to permanent loss of the tooth’s hard tissue [33]. Subsequent brushing with abrasive dentifrices can exacerbate alte­rations to the enamel surface because they may eliminate the weakened enamel and exacerbate the erosion and wear occurrences [34].

Additionally, it was noted that the significant decrease in microhardness after using hydrogen peroxide was attributed to the loss of mineral content caused by demineralization resulting from oxidation-reduction processes of bleaching chemicals [35]. Another explanation for the reduction in enamel microhardness within the hydrogen peroxide group may be alteration in its microstructure (i.e., porosity and surface area), which was most likely caused by the organic enamel deterioration [36].

This aligns with previous findings by Eric et al. [36], who demonstrated that hydrogen peroxide agents can reduce enamel microhardness due to demineralization effects. Although hydrogen peroxide effectively whitens, it may compromise the enamel integrity, particularly in formulations lacking remineralizing agents. On the other hand, the present findings are inconsistent with those of Borges et al. [37], who found that hydrogen peroxide agents did not significantly impact the enamel microhardness.

In contrast, the activated charcoal group maintained stable microhardness levels, while the control group exhibited a slight but nonsignificant increase in micro­hardness. This may be due to the presence of a high concentration of fluoride content (1450 ppm) in the exa­mined charcoal-based and control toothpastes, which restores the microhardness of the enamel surface. The presence of fluoride in the formulation of toothpaste has the ability to enhance remineralization of the enamel surface [38].

This supports Koc Vural et al. study [39], who observed that there was no significant change in microhardness from charcoal dentifrices, while the microhardness of enamel treated with regular fluoridated toothpaste increased. Additionally, Oliveira et al. [40] demonstrated that following a week of treatment, the fluoride-only toothpaste considerably raised the level of microhardness of demineralized samples.

Therefore, our results reject the null hypothesis, as a statistically significant difference was found between these two agents in terms of their effects on enamel color change and surface microhardness.

The study’s limitations include its in vitro nature, which does not fully replicate the dynamic environment of oral cavity as well as its use of only one concentration of hydrogen peroxide and single commercial brand of toothpaste. Also, the study did not evaluate long-term effects beyond the 14-day simulated brushing protocol applied, which may not capture cumulative enamel alterations or whitening sustainability over time. Future studies incorporating in vivo analysis, broader product comparison, and extended evaluation periods are recommended to validate and expand upon our findings.

Conclusions

The results suggest that hydrogen peroxide toothpaste is more effective for achieving noticeable whitening but has an adverse effect on the enamel microhardness. On the contrary, charcoal-based toothpaste offers a safer profile in terms of enamel microhardness but with reduced whitening efficacy. These findings highlight the necessity for personalized dental care, where both cosmetic objectives and enamel preservation should influence product selection.

Disclosures

Author contributions: Conceptualization: N.A.H.A., N.M.S.; Methodology: N.A.H.A., N.M.S.; Investigation and data collection: M.E.F.; Formal analysis: N.A.H.A., H.A.E., A.A.E.; Data curation: M.E.F., H.A.E.; Writing of original draft: O.N.E.; Writing – review and editing: A.G.H.; Supervision: N.A.H.A., A.G.H.; Project administration: N.A.H.A. All authors have read and agreed to the published version of the manuscript.

Funding: This research received no external funding.

Institutional Review Board statement: Not applicable.

Informed consent statement: Not applicable.

Data availability statement: All the data generated or analyzed in this study are included in this manuscript. The data generated in this study may be requested from the corresponding author.

Acknowledgments: None.

Conflicts of interest: The authors declare no conflicts of interest.

AI use statement: Not applicable.

References