Effectiveness of Vitamin C in managing gingival hyperpigmentation: A systematic review and meta-analysis

Abstract

Background:

Gingival hyperpigmentation is a significant esthetic concern, particularly in individuals with high smile lines. Traditional surgical methods for depigmentation often face the challenge of recurrence. Vitamin C, due to its antioxidant properties and role in melanin inhibition, has emerged as a promising adjunctive agent for gingival depigmentation.

Aim:

This systematic review and meta-analysis aimed to evaluate the effectiveness of Vitamin C in managing gingival hyperpigmentation, either as a standalone treatment or in combination with other modalities.

Materials and Methods:

The protocol of this systematic review was registered in PROSPERO (CRD42024613787), and it was in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analysis 2020 guidelines. A comprehensive literature search was conducted across multiple databases, including PubMed, Scopus, Cochrane Library, and Google Scholar, for studies published between 2018 and 2024. The review included randomized controlled trials (RCTs) and clinical trials comparing Vitamin C interventions to conventional depigmentation techniques, and a meta-analysis was conducted using the standardised mean difference (SMD) as the effect measure.

Results:

Eight studies were included for qualitative synthesis and three RCTs for quantitative analysis. The meta-analysis revealed a statistically significant improvement in depigmentation scores at 1-month follow-up (SMD: 1.93; 95% confidence interval: 1.37–2.48). However, no significant differences were observed at baseline or at the 3-month follow-up, where high heterogeneity was present.

Conclusion:

Vitamin C demonstrates promising short-term efficacy in the treatment of gingival hyperpigmentation, especially when used as an adjunct to conventional therapy. However, variability in protocols and long-term outcomes highlights the need for further standardized trials.

INTRODUCTION

Gingival health and esthetics are crucial elements of a pleasing smile. However, when excessively or prominently visible during smiling, gingival pigmentation may lead to psychological distress and self-consciousness in affected individuals.[1] Although gingival pigmentation is not pathologic, the presence of darkened gums commonly referred to as “black gums” can pose a cosmetic concern, especially in individuals with a high smile line. The etiology of gingival pigmentation may be either physiological or pathological, and it is observed across all ethnicities and geographic regions.[2] Melanin pigmentation of the gingiva arises from melanin synthesized by melanocytes.[3] It can be either active or passive and is often influenced by external factors such as tobacco use.[4] Although pigmentation levels can vary with age and ethnicity, there is no notable difference between sexes. A variety of gingival depigmentation (GD) techniques have been developed, including scalpel scraping, bur abrasion, cryotherapy, electrosurgery, and laser therapy.[5] These procedures work by eliminating the pigmented epithelial layer and a part of the underlying connective tissue, allowing for the regrowth of gingival epithelium free from pigmentation. The most common shortcoming of all these procedures is re-pigmentation, which can occur as early as 33 days. Thus, the depigmentation procedures are challenging for the clinician. To mitigate these limitations and enhance the stability of treatment outcomes, Vitamin C has been evaluated as an adjunctive modality in the management of gingival hyperpigmentation.

Vitamin C, also known as ascorbic acid, is a water-soluble antioxidant that is crucial for the synthesis of collagen. It plays an important role in maintaining the health of bones, teeth, ligaments, and gingival tissues. It also modulates immune function and has been shown to reduce hyperpigmentation by inhibiting the tyrosinase enzyme through interaction with copper ions at its active site.[6] This mechanism results in decreased melanin synthesis and gingival lightening.[7] Recent literature highlights Vitamin C not only as an effective depigmenting agent but also as a potential adjunct to reduce relapse rates following GDprocedures, suggesting promising implications for its adjunctive use in clinical practice.[8]

This systematic review and meta-analysis aimed to assess the efficacy of Vitamin C as an adjunctive treatment for managing gingival hyperpigmentation. By analyzing data from multiple studies, particularly randomized controlled trials (RCTs), the review seeks to provide a detailed summary of the existing evidence and explore the potential role of Vitamin C in supporting GD therapy.

MATERIALS AND METHODS

This review was carried out in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines[9] following the methodologies established in earlier systematic reviews.[10] The study protocol was also registered with the International Prospective Register of Systematic Reviews (PROSPERO ID: CRD42024613787).

The research question was formulated based on the PICOS framework, which considers the Population, Intervention, Comparison, Outcome, and Study design.[11]

Question: Does the role of Vitamin C as an adjunct in patients with gingival hyperpigmentation result in a comparable or improved degree of pigmentation, as assessed by the Depigmentation Index, when compared to a conventional surgical depigmentation procedure?

Inclusion criteria

P (Population): Patients with gingival hyperpigmentation undergoing surgical depigmentation procedures.

I (Intervention): Adjunctive use of Vitamin C along with surgical depigmentation.

C (Comparison): Surgical depigmentation alone or without a Vitamin C adjunct.

O (Outcome): Assessment of depigmentation using the Dummett- Oral Pigmentation Index (DOPI).

Exclusion criteria

1. In vitro studies

2. Non-English publications

3. Animal studies

4. Case reports

5. Case series

6. Reviews

7. Conference abstracts.

Outcome measures

Primary outcome: Degree of gingival pigmentation assessed using the DOPI.

Search strategy and information sources

Following the publication of a systematic review in 2020, an extensive literature search was conducted to identify additional relevant studies published since that review.

Five electronic databases – PubMed, Scopus, Cochrane Library, ScienceDirect, and LILACS, along with Google Scholar, were systematically searched. A hand search of relevant journals and references was also performed to retrieve any additional studies. For unpublished data or ongoing research, attempts were made to contact the corresponding authors directly.

Electronic searches were performed in PubMed, Google Scholar, and through manual reference checking of relevant articles. The search terms included “hyperpigmentation,” “gingival pigmentation,” “vitamin C,” “RCT,” and “clinical trials.” Only studies published between January 2018 and December 2024 were considered for inclusion.

Screening and selection of papers

Four independent reviewers (RV, SR, VB, and PB) screened titles and abstracts according to predetermined inclusion and exclusion criteria. Full-text versions of studies that met the preliminary criteria were assessed for final inclusion. Any disagreements were addressed and resolved through discussion with the involvement of an additional reviewer when necessary.

Characteristics of included studies

A total of eight studies were included in this systematic review, comprising one case report, two case series, and five RCTs. These studies explored the use of Vitamin C, either alone or in combination with additional methods like microneedling, for treating physiological gingival hyperpigmentation. The intervention across studies primarily involved intramucosal or intradermal administration of Vitamin C, and outcomes were generally assessed using the DOPI [Table 1].

Table 1.

Characteristics of included studies evaluating Vitamin C for gingival depigmentation

Author (year)	Type of study	Population	Intervention	Control group	Outcome
Mostafa D (2022)[16]	Case report	Gingival hyperpigmentation	Vitamin C	-	Gingival depigmentation
Mostafa D Sr. (2023)[15]	Case series	Gingival hyperpigmentation	Vitamin C	-	Gingival depigmentation
Sandhu A (2023)[14]	Case series	Gingival hyperpigmentation	Vitamin C	-	Gingival depigmentation
El-Mofty M (2021)[23]	RCT	Gingival hyperpigmentation	Vitamin C	-	Gingival depigmentation
Swarna Meenakshi P (2024)[19]	RCT	Gingival hyperpigmentation	Vitamin C intradermal injection	Diode laser	Gingival depigmentation
Yussif NM (2019)[24]	RCT	Gingival hyperpigmentation	Local Vitamin C injection	Scalpel	Gingival depigmentation
Chaudhary DS (2023)[20]	RCT	Gingival hyperpigmentation	Microneedling + Vitamin C	Scalpel	Gingival depigmentation
El-Mofty M(2023)[21]	RCT	Gingival hyperpigmentation	Vitamin C injection	Scalpel	Gingival depigmentation

“–” indicates no control group used. RCT – Randomized controlled trial

Although all eight studies were reviewed for qualitative synthesis, the final meta-analysis included only three RCTs because they provided standardized and comparable quantitative data such as mean values, standard deviations, and sample sizes. Case reports and case series lacked control groups and sufficient statistical data, rendering them unsuitable for inclusion in the meta-analysis. Thus, the meta-analysis was restricted to high-quality RCTs that allowed for meaningful statistical comparison of the effectiveness of Vitamin C versus conventional depigmentation techniques [Table 2].

Table 2.

Comparison of Dummett Oral Pigmentation Index scores between Vitamin C and control group at baseline, 1 month and 3 months

Study (author, year)	Control group (n, mean±SD)	P
Baseline
Swarna Meenakshi P (2024)[19]	8, 2.61±0.17	0.66
Esmat SA (2023)[21]	13, 2.26±0.51	0.434
Chaudhary DS (2023)[20]	15, 2.60±0.51	0.076
1 month
Swarna Meenakshi P (2024)[19]	8, 1.07±0.31	0.03
Esmat SA (2023)[21]	13, 0.27±0.36	<0.0001
Chaudhary DS (2023)[20]	22, 0.53±0.52 (1 week)	0.754
3 months
Swarna Meenakshi P (2024)[19]	8, 0.87±0.17	0.04
Esmat SA (2023)[21]	13, 0.23±0.35	<0.0001
Chaudhary DS (2023)[20]	15, 0.85±0.13	–

n – Number of participants; Mean±SD – Mean value±standard deviation; P – Probability value for comparison between the study groups; P<0.05 was considered statistically significant

Characteristics of excluded studies

During the screening process, several studies were excluded after full-text evaluation for failing to meet the predefined eligibility criteria. The most common reasons for exclusion were limitations in study design, such as in vivo experimental models, case series, and case reports, as well as the absence of comparator groups or incomplete outcome reporting.[12,13,14,15,16]

This step was critical in preserving the methodological integrity of the review and ensuring that only robust and clinically relevant studies contributed to the final analysis [Table 3].

Table 3.

Excluded studies and reasons for exclusion

Author (year)	Reason for exclusion
Yussif NM (2017)[12]	In vivo study
MurthyMB (2012)[13]	Case series
Mostafa D (2022)[16]	Case report
Mostafa D Sr. (2023)[15]	Case series
Sandhu A (2023)[14]	Case series

Quality assessment and risk of bias

The methodological quality of the included RCTs was assessed using the Cochrane Risk of Bias tool.[17] Two reviewers independently evaluated the studies across seven domains: random sequence generation, allocation concealment, blinding of participants and personnel, blinding of outcome assessment, incomplete outcome data, selective reporting, and other sources of bias. Each domain was categorized as “low risk,” “high risk,” or “unclear risk” based on the information provided in the respective studies. Any discrepancies between reviewers were resolved through discussion and consensus.

Most domains across the included trials demonstrated a low risk of bias. However, selective reporting was judged as unclear in all included studies, and incomplete outcome data were considered unclear in one study due to insufficient reporting. Overall, the included randomized controlled trials were considered to have a low-to-moderate risk of bias [Figure 1].

Figure 1.

Risk of bias assessment of Included Randomized Controlled Trials

The methodological quality of the present systematic review was evaluated using the AMSTAR-2 (A Measurement Tool to Assess Systematic Reviews) instrument, which examines 16 domains related to methodological rigor and transparency. The review satisfied the majority of critical domains, including a comprehensive search strategy, duplicate study selection, data extraction, appropriate risk of bias assessment, and suitable statistical methods for data synthesis[18] [Table 4].

Table 4.

A Measurement Tool to Assess Systematic Reviews, version 2, assessment of the methodological quality of the present systematic reviews

AMSTAR-2 domain	Assessment	Remarks
Research question and inclusion criteria included PICO components	Yes	Clearly defined Population (gingival hyperpigmentation), Intervention (Vitamin C), Comparator (scalpel/none), and Outcome (gingival depigmentation)
Protocol registered before commencement of review	Yes	Protocol prospectively registered before study initiation
Justification for inclusion of study designs	Yes	Inclusion of RCTs, case series, and case reports is justified due to the limited available evidence
Comprehensive literature search strategy	Yes	Multiple databases searched (PubMed, Scopus, Embase, Google Scholar) with manual search of references
Study selection performed in duplicate	Yes	Two independent reviewers screened titles/abstracts and full texts
Data extraction performed in duplicate	Yes	Data independently extracted and cross-verified
List of excluded studies and justification provided	Yes	Reasons for exclusion documented in the PRISMA flowchart
Description of included studies in adequate detail	Yes	Study characteristics are summarised in tabular format
RoB assessment of individual studies	Yes	Conducted using the Cochrane RoB tool for RCTs; descriptive appraisal for nonrandomised studies
Reporting of funding sources of included studies	Partial Yes	Funding information not available for all included studies
Appropriate methods used for the statistical combination of results	Yes	Fixed- or random-effects models selected based on heterogeneity
Assessment of the impact of RoB on results	Yes	RoB was considered during interpretation and discussion
Assessment of publication bias	No	Not performed due to inclusion of fewer than 10 studies
Explanation and discussion of heterogeneity	Yes	Statistical heterogeneity was assessed using I2 statistics and discussed
Reporting of conflicts of interest	Yes	The authors declared no conflict of interest

AMSTAR-2 – A Measurement Tool to Assess Systematic Reviews, version 2; RoB – Risk of bias; RCTs – Randomized controlled trial; PICO – Population, Intervention, Comparison, Outcome

The overall confidence in the results of this review was rated as moderate. Although most methodological standards were met, the inability to assess publication bias due to the limited number of included studies slightly reduced the overall rating. Nevertheless, the findings of this review are considered reliable within the constraints of the available evidence.

Search outcomes and evaluation

The results were interpreted following PRISMA guidelines. The review addresses the strengths and limitations of the included studies, their clinical relevance, and suggests directions for future research. Although many studies have investigated treatments for gingival hyperpigmentation, only three RCTs fulfilled the criteria for inclusion in the meta-analysis. These studies featured similar control groups and outcome measures, specifically the DOPI index, with follow-up periods of three months.[19,20,21] Heterogeneity analyses were performed at both baseline (pretreatment) and posttreatment time points, with the latter assessing the treatment effect in comparison with the control group. The effect size was expressed as a standardized mean difference (SMD) along with a 95% confidence interval (CI). Mean values and standard deviations for both treated and control groups were used to calculate the SMD for each study. Weights were assigned to studies using the inverse variance method. The degree of heterogeneity was evaluated using the I² and τ² statistics. All statistical analyses were performed using the meta package in R software (version 4.4.0, R Foundation for Statistical Computing, Vienna, Austria), with a significance threshold set at 5%.

RESULTS

Study selection

A total of 2102 records were identified through database searches and manual reference screening. After duplicate removal and title and abstract screening, 52 articles underwent full-text assessment for eligibility. Of these, three studies satisfied the inclusion criteria and were ultimately included in the qualitative and quantitative synthesis [Figure 2].

Figure 2.

PRISMA 2020 Flow Diagram of Study Selection

At baseline, the τ² value was calculated to be 0.03 (95% CI: 0.0–5.95). Since the confidence interval includes zero as its lower bound, this indicates minimal heterogeneity among the included studies at baseline, consistent with Higgins and Thompson’s guidelines. The τ value was 0.175, implying that the estimated standard deviation of the true effect sizes is 0.175. An I² = 4.2% and an H = 1.02 further suggest that only about 4% of the observed variation is due to differences in true effect sizes. The prediction interval ranged −3.91 to 4.05, indicating that future studies might report either positive or negative treatment effects based on current findings. The Q statistic, used to assess heterogeneity, was 2.09 with 2 degrees of freedom, yielding a P = 0.35. This implies that heterogeneity at baseline was not statistically significant [Figure 3].

Figure 3.

Forest plot comparing baseline Depigmentation Index scores between Vitamin C and control groups. CI: Confidence interval; df: Degrees of freedom; τ² (tau-squared): Between-study variance in a random-effects model; χ² (Chi-squared): test for heterogeneity; 12: percentage of total variation across studies due to heterogeneity; Z: Test statistic for overall effect; P: Probability value. P < 0.05 was considered statistically significant

At 1 month, the overall SMD based on the random effects model was 1.93 (95% CI: 1.37, 2.48), and the interval does not include zero, suggesting the treatment effect, which was statistically significant (P < 0.001). Further, the τ² value was 0 (95% CI: 0–5.67), suggesting very low heterogeneity across studies. The value of I² was 0%, and H was 1.0, indicating hardly any variation in the data due to the true effect size differences. The prediction interval ranged from −1.67-5.52, suggesting the possibility of a negative treatment effect in future studies. However, the interval is skewed on the positive side, suggesting a greater likelihood of positive effects in the studies. The Q-value obtained was 1.44 (degree of freedom, df: 2) with a corresponding P = 0.487, suggesting that the heterogeneity was not statistically significant [Figure 4].

Figure 4.

Forest plot comparing Depigmentation Index scores at 1 month between Vitamin C and control groups

At the 3-month follow-up, the overall standardized mean difference (SMD) using a random effects model was 1.31 (95% CI: −0.47–3.09). Since the confidence interval includes zero, the treatment effect was not statistically significant (P = 0.151). The τ² value was 2.19 (95% CI: 0.42–94.11), which excludes zero, indicating the presence of heterogeneity between studies. The estimated standard deviation of the true effect sizes (τ) was 1.48. The I² statistic was 90.9% (95% CI: 76.2%–96.5%), indicating that approximately 91% of the observed variation is attributable to genuine differences in effect sizes across studies. The heterogeneity is regarded as “very large.” The prediction interval ranged from −20.79-23.41, suggesting the possibility of negative treatment effects in future studies at 3 months of evaluation. The positive and negative treatment effects are equally likely at 3 months. The Q-value obtained was 21.95 (df: 2) with a P < 0.001, indicating significantly large heterogeneity in true effect sizes of the included study [Figure 5].

Figure 5.

Forest plot Comparing Depigmentation Index scores at 3 months between Vitamin C and control groups

The meta-analysis revealed a statistically significant positive effect of Vitamin C at 1-month follow-up (SMD: 1.93; 95% CI: 1.37–2.48) with negligible heterogeneity. However, no notable effect was detected either at baseline or at the 3-month follow-up, with the latter exhibiting considerable heterogeneity.

DISCUSSION

The potential of Vitamin C in preventing the recurrence of pigmentation after GD has drawn growing interest. Research indicates that Vitamin C may contribute not only to the initial depigmentation process but also to the long-term stability of results. For instance, Sheel et al. observed that the postoperative application of topical Vitamin C was associated with the absence of pigmentation recurrence over a 9-month follow-up period.[8] This is consistent with the findings of Sandhu et al., who noted in their comparative case series that the patient receiving Vitamin C postoperatively showed sustained gingival lightness with no signs of erythema or pigment relapse.[14] Further clinical evidence is anticipated from an ongoing randomized study exploring the role of Vitamin C in preventing the reappearance of pigmentation postdepigmentation therapy.[22]

The current systematic review consolidates the clinical evidence regarding the use of Vitamin C as an adjunct in reducing melanin pigmentation, particularly in the gingiva. While short-term outcomes, especially at the 1-month follow-up are encouraging, current literature remains limited regarding the evaluation of Vitamin C as a standalone treatment modality. To uphold methodological rigor, this review excluded in vitro studies, animal experiments, case reports, and case series, recognizing their limited clinical applicability.

Among the eight studies included in the qualitative synthesis, three were nonrandomized designs (one case report and two case series) by Mostafa et al. and Sandhu et al., which demonstrated positive clinical outcomes following Vitamin C treatment.[14,15,16] However, due to the absence of control groups, small sample sizes and lack of standardized outcome measures, these studies were excluded from meta-analysis.

The remaining five RCTs provided more robust data. These included studies by Swarna et al., Chaudhary et al., Esmat et al., El-Mofty et al., and Yussif et al.[19,20,21,22,23,24] Each study assessed GD using Vitamin C delivered either as intramucosal injections or via microneedling. However, only three of these RCTs were eligible for inclusion in the meta-analysis due to complete reporting of quantitative outcome data.

At baseline, the current analysis found no significant difference in pigmentation levels (I² = 4.2%), a finding consistent with Chaudhary et al., who noted no immediate reduction in pigmentation immediately after Vitamin C injection[20] and Mostafa et al., who also observed that pigmentation reduction was not evident until after multiple sessions or weeks of follow-up.[15]

The meta-analysis revealed a statistically significant positive effect of Vitamin C at 1-month follow-up (SMD: 1.93; 95% CI: 1.37–2.48) with negligible heterogeneity. However, no significant effect was observed at baseline or at 3 months, with the latter showing substantial heterogeneity.

The statistically significant depigmentation effect of Vitamin C at 1-month follow-up in the current meta-analysis is in agreement with the findings of Aroca et al., who observed a notable reduction in gingival melanin pigmentation after 4 weeks of applying 2% ascorbic acid solution.[25] Similarly, Al-Hajri et al. reported a significant improvement in pigmentation scores following subepithelial Vitamin C injections, with marked results evident within 1 month of treatment.[26]

However, the current review found a nonsignificant treatment effect at 3 months with high heterogeneity (I² =90.9%). This contrasts with the study by Aroca et al., which demonstrated sustained depigmentation over a 3-month period.[25] The discrepancy may be due to differences in treatment protocols. Aroca et al. applied topical 2% ascorbic acid twice daily for 4 weeks, whereas the studies included in the current review predominantly used single or fewer injectable sessions. In addition, Aroca’s study excluded smokers and included patients with mild baseline pigmentation, while the included studies had more variability in pigmentation severity and did not consistently control for confounding factors such as smoking or oral hygiene status. These variations likely influenced both the initial response and the recurrence patterns observed at longer follow-ups.

The lack of a significant effect at 3 months (SMD: 1.31; 95% CI: −0.47–3.09), coupled with high heterogeneity, is consistent with the results reported by Haque et al. who observed variable long-term outcomes following GD with injectable Vitamin C.[27] Similar to the present review, their study observed initial improvement followed by varying degrees of repigmentation by the 3^rd month, particularly in patients with higher baseline pigmentation scores. However, unlike the studies included in this review, Haque et al. administered Vitamin C weekly for a longer duration (6 weeks vs. 3–4 weeks in most included trials), and their cohort included a higher proportion of smokers and individuals with melanin-rich ethnic backgrounds. These differences in application frequency, participant demographics, and baseline melanin levels may have contributed to the wider range of treatment responses and recurrence rates observed in their study, which partially explains the heterogeneity noted in the present analysis.

These results suggest that Vitamin C may be effective not only for initial pigment reduction but also for maintaining the depigmentation effect over time, potentially reducing recurrence rates. These results align with the promising short-term effects observed in this review, though the variability in long-term results (particularly at the 3-month mark) warrants further investigation to confirm the optimal application protocols and sustained effects of Vitamin C in clinical practice.

Limitations

This review is limited by the small number of RCTs eligible for meta-analysis, which may affect the generalizability of the findings. Variability in intervention protocols, dosage, and follow-up durations across studies introduced heterogeneity that may have influenced outcomes. In addition, incomplete reporting of outcome measures and lack of control for confounding factors such as smoking and oral hygiene further limit the strength of the conclusions. The exclusion of non-randomised studies, while methodologically appropriate, may have omitted relevant clinical observations. Recent RCTs by Haque et al.[27] also explored Vitamin C-based depigmentation approaches, but were excluded due to variations in methodology and outcome reporting.

CONCLUSION

Vitamin C has demonstrated efficacy as a depigmenting agent in the short-term management of gingival hyperpigmentation. The antioxidant properties of Vitamin C, along with its ability to inhibit melanin synthesis, make it a promising minimally invasive alternative to conventional surgical interventions. Furthermore, techniques that combine vitamin C with microneedling have demonstrated superior clinical outcomes, suggesting a potential synergistic effect between the two modalities.

However, despite these promising findings, the long-term sustainability of depigmentation following vitamin C treatment requires further investigation. The variability in outcomes at the 3-month follow-up highlights the need for standardized protocols regarding dosage, application methods, and treatment frequency.

Future RCTs with larger sample sizes and extended follow-up periods are essential to establish the long-term efficacy and optimal application strategies for Vitamin C in GD. Until then, topical application of Vitamin C can be considered a promising adjunctive treatment, particularly for patients seeking nonsurgical options.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.