Effectiveness of vitamin C and D supplementation as an adjunct in the management of gingivitis in school-going children: a randomized controlled trial

Abstract

Background

Gingival diseases in Pakistan rank among the nation’s foremost oral health issues. Vitamins C and D may influence gingival health through collagen metabolism and immune regulation.

Objective

The study aimed to assess whether vitamin C and D supplementation can reduce bleeding on probing (BOP), measured as the percentage of bleeding sites (BOP%).

Materials and methods

A total of 136 school-going children, aged 8–16 years old, were included in this randomized controlled trial (RCT). Eligible participants presenting to the University of Lahore Dental Hospital were recruited. Oral health status was determined using the BOP examination. Participants were divided into two groups: Group A (standard care) and Group B (standard care plus vitamin C and D supplements). Children in both groups A and B were treated with ultrasonic scaling and given structured oral hygiene instructions. Children in group B (intervention group) were given an 8-week plan of vitamin C and D supplementation. Participants were recalled at 4 and 8 weeks to reassess.

Results

Among 150 enrolled children, 14 were lost to follow-up; 136 were included in analysis (Group A n = 68, Group B n = 68). Baseline BOP% was significantly higher in Group B (p < 0.001). Thus follow-up comparisons were adjusted for baseline. Group B had lower BOP% than Group A at 4 weeks (p < 0.001) and remained lower at 8 weeks (p < 0.001). Both groups demonstrated a significant reduction in BOP% over time.

Conclusion

Vitamin C and D supplementation improved gingival health among school-going children.

Supplementary Information

The online version contains supplementary material available at https://doi.org/10.1186/s12889-026-27539-2.

Introduction

Oral health spans a wide spectrum of conditions, including dental caries (tooth decay), periodontal diseases, oral cancers, oral manifestations of systemic infections, trauma, congenital diseases, and infections of the oral cavity [1]. Many of these oral health issues are largely preventable and respond well to treatment if addressed in their early stages. The Global Burden of Disease (GBD) study reported that approximately 3.5 billion people worldwide are impacted by oral diseases. [2] Periodontal diseases, of which gingival inflammation is an early sign, stand among the foremost causes of tooth loss around the world, adversely impacting oral function, aesthetics, and overall quality of life [3, 4]. Pathogenic microorganisms, along with various host-derived factors, drive the inflammatory processes that lead to inflammation and tissue breakdown [5]. Although mechanical interventions such as scaling and root planning remain the primary therapeutic approach, persistently high incidence of gingivitis in resource-challenged settings suggests that current strategies may not be sufficient. Micronutrient deficiencies are frequently observed in developing countries such as Pakistan, which can worsen tissue breakdown by weakening immune defenses and hindering tissue regeneration [6, 7].

Vitamin C, known for its antioxidant role, is essential for collagen formation and preservation of the periodontal ligament’s (PDL) structural stability [8]. When vitamin C levels dip below optimal thresholds (0.9–1.2 mg/dL), wound healing is compromised, leaving tissues more prone to infection, inflammation, and increased levels of bleeding [9]. Vitamin D has traditionally been linked to bone health, but it has also gained attention as a key immunomodulator capable of shaping both innate and adaptive immune mechanisms [10]. Consequently, inadequate vitamin D levels may heighten susceptibility to PDL deterioration [11, 12]. Pakistani population suffers from widespread malnutrition due to limited availability of nutrient-rich foods; thus, the collective burden of oral diseases and the significance of nutritional interventions in lower-income countries are amplified [13, 14].

It is well established that severe vitamin C deficiency directly results in scurvy, which manifests as gingival bleeding and worsens the effects of existing inflammation [15]. Bleeding on probing (BOP) has also been shown to improve with vitamin C supplementation [16]. Similarly, a direct correlation has been noted between low serum vitamin D and increased oral soft tissue disease indicators such as BOP [17, 18].

Despite these critical links, empirical data on the oral health-related effects of vitamin C and D supplementation in the pediatric population remains sparse, signifying a critical research gap. Addressing this gap could inform cost-effective, non-invasive strategies that complement conventional treatments, ultimately improving oral and systemic health outcomes. Therefore, this randomized controlled trial (RCT) aimed to evaluate the effectiveness of adjunctive vitamin C and D supplementation in reducing gingival inflammation among school-going children with gingivitis.

Problem statement

Gingivitis is common in children aged 8–16 years and is primarily managed through mechanical plaque control. However, the additional benefit of vitamin C and D supplementation alongside nonsurgical periodontal therapy in this age group remains unclear.

Study objectives

• To compare changes in clinical gingival inflammation after nonsurgical periodontal therapy with and without adjunctive vitamin C and D supplementation among children with gingivitis.

• To assess whether adjunctive vitamin C and D supplementation provides additional improvement in gingival bleeding outcomes compared with nonsurgical periodontal therapy alone.

Materials and methods

The single-center, parallel group RCT was initiated upon obtaining Institutional Review Board (IRB) approval from the Ethics committee of the University Institute of Public Health, The University of Lahore, Ref No. IRB-UOL-FAHS/835II/2019, on 17-01-2019. It was also retrospectively entered into the UK’s Clinical Study Registry in 2025. The Study was conducted in the University of Lahore Dental Hospital, where school-going children aged 8 to 16 years old were recruited. Children younger than this age group were considered too dependent on parental involvement for oral hygiene-related behaviors. This age group is also at high risk for plaque and hormone-induced gingival inflammation, making it an ideal group to detect the adjunct benefits of vitamins. As BOP% was being measured, all ages showed appropriate, fully erupted permanent teeth for examination; primary teeth and partially erupted permanent teeth were excluded to reduce measurement variability caused by exfoliating primary or erupting permanent teeth. Simple random sampling was used to select patients from the available list. Sample size was calculated using G*Power software (version 3.1.9.4) for a two-independent-means t-test, with an effect size of 0.52, based on a similar intervention study for gingivitis parameters [19], with α = 0.05 and power = 0.85. The required sample size was 136 participants (68 per group). Initially, a sample size of 150 participants was calculated after accounting for a 10% dropout rate. At the end of the study, a total of 136 participants completed the trial, divided into two groups, A and B, at a 1:1 ratio. Figure 1 shows the participant flow in a CONSORT diagram and assessment schedule.

Fig. 1.

CONSORT flow diagram with intervention and assessment schedule

Systemically healthy school-going children aged 8–16 years old from Lahore were included in the study. Guardians of all participants, as well as the participants themselves, gave consent to follow-up examinations. Children with diabetes, bleeding disorders, immunodeficiency, oral conditions needing urgent treatment, and those who had undergone scaling in the past 3 months were excluded. Screening for relevant medical and dental history was performed to ensure participants met the inclusion criteria. Selection for valid study participants was conducted at the University of Lahore Dental Hospital.

Participants were randomly assigned to group B (intervention group + standard care) and group A (standard care). Allocation was concealed using sequentially numbered, opaque, sealed envelopes. Both groups were assessed by counting the number of BOP sites and calculating the percentage (BOP%). The primary outcome was BOP%, checked at baseline, 4, and 8 weeks; this was done by trained dental professionals. Probing of 6 sites in all erupted permanent teeth was performed, and BOP% was calculated for use in analysis (sites with bleeding/sites examined) x 100. Both groups received ultrasonic scaling and oral hygiene instructions (OHI) as standard care. Additionally, group B was prescribed micronutrient supplementation (600 IU/day vitamin D3 (cholecalciferol) and 25 mg/day vitamin C) for 8 weeks. This dosage of Vitamin D is consistent with age based recommended intakes. The vitamin C dosage served as a conservative adjunct supplemental dose intended to improve feasibility and tolerability in the complete study population as age dependent dosing was not calculated. [20, 21]. The participants were recalled at 4 and 8 weeks to reassess BOP scores. Adherence to vitamin supplements among participants was assessed using parent and participant self-reports.

The study duration was 1 year, allowing time for ethical approval, participant recruitment, scheduling, and completion of follow-up assessments. Upon completion of the study duration, follow-ups for 136 participants were recorded. To minimize bias, the baseline and follow-up BOP examinations were performed by examiners who were blinded to group allocation. Vitamin C and D supplement dispensing was handled separately from assessment of BOP. Because no placebo was used, participants were not blinded to supplementation status. No adverse events related to supplementation were reported on follow-up.

Data was analyzed using IBM SPSS Statistics for Windows, Version 24.0 (IBM Corp., Armonk, NY, USA). Median values were determined. Oral hygiene habits were compared between the two groups. Because BOP% distributions deviated from normality, we used non-parametric tests, Mann Whitney U for between-group comparisons at each time point and the Friedman test for within-group change over time. The Friedman test was used to assess outcomes within a group over time, and Mann-Whitney U tests were used to assess intergroup relationships. Quade Rank ANCOVA was done to compare the 4- and 8-week follow-up while adjusting for baseline differences in BOP%. For Quade rank ANCOVA, baseline BOP% was used as the covariate and the follow-up BOP% (week 4 and week 8) was rank transformed, adjusted group differences were then tested using an ANCOVA model on the ranked outcome (Quade method), yielding adjusted mean ranks. Given that BOP% is a continuous variable and that dichotomization would result in loss of information and reduced statistical power, a multiple linear Regression model was fitted to identify predictors of BOP%, using 8-week BOP% as the dependent variable and gender, brushing habits, weekly flossing habits, annual dental visit frequency, and study group (A or B) as independent variables. As a sensitivity analysis appropriate for proportional outcomes, we also conducted fractional logistic regression using week-8 bleeding sites as events and total sites examined as trials, adjusting for the same covariates (Supplementary Table 1).

Results

Out of 136 subjects, 88 (65%) were male, and 48 (35%) were female. Table 1 compares participant characteristics related to oral hygiene as judged at the initial appointment. There was no significant difference between groups A and B regarding these practices.

Table 1.

Oral hygiene behaviors by group

Oral hygiene behavior	Category	Group A n (%)	Group B n (%)	p-value
Weekly flossing habits	0 days	55 (80.9)	59 (86.8)	0.326
	1–3 days	4 (5.9)	4 (5.9)
	4–6 days	7 (10.3)	4 (5.9)
	7 days	2 (2.9)	1 (1.5)
Annual dental visit frequency	Only in case of pain	54 (79.4)	53 (77.9)	0.810
	1–2 times	11 (16.2)	11 (16.2)
	More than 2 times	3 (4.4)	4 (5.9)
Brushing habits	Not daily	2 (2.9)	4 (5.9)	0.636
	Once daily	34 (50.0)	28 (41.2)
	Twice daily	32 (47.1)	36 (52.9)

P-values were obtained from the Mann–Whitney U test (two-tailed) comparing the ordinal distribution of each behavior between Group A and Group B

For all study participants, regardless of group, the changes in BOP% over the study duration were judged as highly significant. This change across all participants was judged statistically significant (p < 0.001 using the Friedman test). As with all study participants, Table 2 shows that, for both groups separately, the BOP percentages across all 3 time periods are statistically significant (p < 0.001). Table 3 shows the intergroup comparison of BOP percentage at Baseline, 4 Weeks, and 8 Weeks in groups A and B. At all 3 time points, the scores were statistically significant (p < 0.001, using the Mann-Whitney U test). At Baseline, Group B had a higher baseline BOP% (Mean rank: 99.54 compared to 37.46). Although participants were randomized, an imbalance in BOP% was observed. This may have occurred by chance in this moderate sized trial without use of stratified or block randomization and was therefore adjusted for in the analysis.

Table 2.

BOP% timeline for both groups

Group	Time	Median±IQR	P-Value
A	Baseline	22.45±1.60	<0.001**
	4 Weeks	13.40±2.35
	8 Weeks	8.55±1.57
B	Baseline	24.95±1.95	<0.001**
	4 Weeks	11.80±1.90
	8 Weeks	7.55±1.50

**p < 0.001 (Friedman Test)

Table 3.

Intergroup comparison of BOP%

Measure	Group	Mean Rank	P-value
Baseline BOP%	Group A (n=68)	37.46	<0.001**
	Group B (n=68)	99.54
4 Week BOP %	Group A (n=68)	88.60	<0.001**
	Group B (n=68)	48.40
8 Week BOP %	Group A (n=68)	85.85	<0.001**
	Group B (n=68)	51.15

**p < 0.001 (two-tailed Mann–Whitney U). Quade rank ANCOVA was also done (Table 3) to account for the differences at Baseline

At both 4 and 8 Weeks the BOP sites percentage is significantly lower for Group B than Group A (Mean rank 48.40 compared to 88.60 at 4 weeks and 51.15 compared to 85.85 at 8 weeks).

Multiple linear regression analysis (ANCOVA framework) with Week-8 BOP% as the dependent variable, adjusting for gender, flossing habits, brushing habits, and dental visit frequency. The intervention group remained a significant predictor of lower Week-8 BOP% (B = − 0.92, p < 0.001) (Table 4).

Table 4.

Multiple linear regression predicting BOP% at 8 weeks

Predictor	B	SE	β	p-value
Gender (ref - Male)	0.059	0.155	0.030	0.703
Weekly flossing habits (per category increase)	-0.179	0.108	-0.130	0.099
Annual dental visit frequency (per category increase)	0.080	0.138	0.045	0.565
Group (ref – Group A)	-0.924	0.151	-0.475	<0.001
Brushing habits (per category increase)	-0.036	0.131	-0.022	0.783

Dependent variable: BOP% at 8 weeks

B unstandardized coefficient, SE Standard error, β standardized coefficient

In the binomial logistic model (fractional logistic regression), Group B had lower odds of bleeding at week 8 compared with Group A (adjusted OR 0.877, 95% CI 0.839–0.916; p < 0.001; (Supplementary Table 1).

Table 5 shows a Quade rank analysis of covariance (as the data did not meet normality assumptions), which was conducted to compare the adjusted ranks of BOP% between groups while controlling for baseline values that showed a significant difference (p < 0.001). At 4 weeks, a significant difference was observed between the control and intervention groups (F{1,133}=20.81, p < 0.001, partial η²=0.135), with the intervention group showing lower ranked BOP%. At 8 weeks, the difference remained significant (F{1,133}=15.07, p < 0.001, partial η²=0.102), indicating sustained improvement in the intervention group relative to the control group. These findings suggest that the reduction in BOP% among group B was significantly better, even after adjusting for baseline differences.

Table 5.

Quade rank ANCOVA for adjusted BOP% (ranks) at 4 and 8 weeks

Time Point	Group	Adjusted Mean Rank	Std. Error	95% CI	F (df=1,133)	p-value	Partial Eta²
Week 4	Group A (n=68)	90.28	5.60	79.21–101.35	20.81	<0.001	0.135
	Group B (n=68)	46.72	5.60	35.65–57.79
Week 8	Group A (n=68)	87.84	5.84	76.29–99.39	15.07	<0.001	0.102
	Group B (n=68)	49.16	5.84	37.61–60.71

Values represent adjusted mean ranks of BOP% after controlling for baseline BOP% (Quade Rank ANCOVA)

Figure 2 shows the key findings from this study, a graphical representation of improvement in BOP% over the course of the study for both groups.

Fig. 2.

The progression of BOP% for both groups over time from baseline to the end of the trial

Discussion

This study hypothesized that vitamin C and D supplementation could provide a significant adjunct to the management of Gingival inflammation. The findings indicated that micronutrient support improved inflammation resolution among children, aligning with a large body of existing literature that has linked vitamin deficiency to worse periodontal outcomes.

Gingivitis is one of the most common dental health issues in school-going children, especially in low- and middle-income countries like Pakistan. Most epidemiological surveys conducted across different areas of Pakistan report high plaque scores, gingival inflammation, and calculus deposition among children due to unsatisfactory oral hygiene practices and a lack of awareness among parents and caregivers [22, 23]. Countries with developing socioeconomic conditions show a general lack of access to preventive dental services and poor health literacy, acting as the main cause for the early onset and continuation of gingivitis among children [24].

A key factor that contributes to gingivitis and periodontitis in extreme cases among children is the general lack of formal oral hygiene education and awareness at schools and at home. Inquiries carried out in Lahore, Karachi, and Peshawar, Pakistan reveal that a considerable number of children in school-going age show irregular brushing, less understanding of proper brushing methods, and infrequent use of floss or other adjunctive hygiene aids [25, 26]. Many children rely on traditional brushing techniques at irregular intervals, without guidance, resulting in inconsistent plaque control and eventually periodontal problems. These problems are further amplified by dietary habits such as high-carbohydrate intake and low-nutrient-rich foods. These habits promote plaque formation and can reduce micronutrient intake, leading to deficiencies, as reported in the literature [27].

Limited dietary diversity, low fruit and vegetable intake, and restricted sunlight exposure translate into a very high prevalence of vitamin C and D deficiency among children living in urban and semi-urban settings [28, 29]. Nutritional surveys indicate that inadequate vitamin D levels are common in Pakistan, while a significant proportion of the population suffers from suboptimal vitamin C intake, both of which are crucial for maintaining gingival health [30]. Vitamin C deficiency impairs collagen synthesis and weakens the epithelium, thereby increasing the susceptibility of gingival tissues to plaque-induced inflammation and bleeding. Vitamin D deficiency compromises immunomodulation, enhancing the risk of gingival infections, and may also contribute to exaggerated inflammatory responses in the periodontal tissues.

Our findings show that supplementation with vitamins C and D can significantly improve periodontal status as determined by BOP when added to standard care. These findings are consistent with clinical evidence supporting the role for vitamin C in reducing gingival inflammation. In a double-blind RCT, an ascorbic acid derivative dentifrice was associated with significant improvement in gingival inflammation among patients with gingivitis [31]. Such improvements are particularly meaningful in developing countries, where micronutrient deficiencies and limited awareness of oral hygiene coexist and mutually promote gingival inflammation. Evidence also supports the adjunctive role of vitamin C in periodontal therapy, with a review reporting improvement in bleeding indices in gingivitis across clinical studies. This data complements our findings using BOP% as an inflammation endpoint [32].

The increased reduction in BOP within the intervention group is consistent with evidence that vitamin C lowers oxidative stress and maintains periodontal connective tissue matrix stability [33], while vitamin D and C enhance innate immunity, increase antimicrobial peptide expression, and diminish gingival inflammation [34]. The immunomodulatory role of vitamin D has also been supported in interventional work. A randomized dose–response trial reported a dose-dependent anti-inflammatory effect of vitamin D supplementation on gingivitis outcomes, aligning with the improved inflammation resolution observed in our intervention group [35]. Supporting evidence also demonstrates an inverse association between serum 25-hydroxyvitamin D concentrations and chronic gingivitis measured by BOP. This also aligns with the proposed immunomodulatory role of vitamin D in gingival inflammation [36].

These findings emphasize the integration of nutrition-focused interventions along with routine oral hygiene education in school dental programs, as gingivitis is a reversible condition. Integrating targeted micronutrient supplementation into school-based health programs and enhancing parental education are viable, cost-effective strategies to combat pediatric gingivitis. Evidence also suggests the effect may be strongest in deficient populations. A systematic review and meta-analysis reported larger reductions in bleeding tendency when baseline vitamin C status is low. This may partially explain why supplementation can be more impactful in settings where micronutrient deficiencies are common [37]. In regions with limited dental infrastructure, early nutritional intervention can serve as a powerful adjunct to traditional care, effectively reducing the oral disease burden in populations with similar nutritional deficiencies [38].

Limitations

• Single-center recruitment limits generalizability.

• A follow-up period of 8 weeks may be considered short, and studies with longer follow-up might be required.

• A marked baseline imbalance in BOP% was present despite randomization, this baseline difference in BOP% was handled statistically however this may reflect imbalance and is a limitation.

• As the intervention group was given both vitamin C and D simultaneously, there are no separate groups for each vitamin. Thus, the specific individual contribution of vitamin C and vitamin D cannot be determined.

• The dosage of vitamin C and D given to the intervention group was fixed, and we did not evaluate alternative dosing regimens nor was dietary behavior considered or controlled for to not have it confound with supplement effect. So, whether higher or lower doses would produce greater or smaller reductions in BOP% based on individual habits and experience could not be assessed.

• The vitamin C dose (25 mg/day) given was below the NIH recommended dietary allowance for a large portion of participants (≥ 9 years). Thus, the effect of providing an age appropriate or higher dose is not determined.

• Since no placebo was used participants in the supplementation group were likely aware of receiving vitamins and may have modified oral hygiene behaviors thus performance bias may be present.

• Adherence to the supplement regimen and hygiene practices was self-reported and not confirmed more objectively, which can lead to bias.

• This trial was registered retrospectively. Such registration may lead to selective outcome reporting bias thus the findings should be viewed through this lens.

• Although the selected age range is relevant to plaque and hormone associated gingival inflammation, pubertal status varies considerably within our subjects age range. It is plausible that hormonal changes stemming from puberty may be influencing gingival bleeding independently of supplementation and pubertal stage was not measured or adjusted for. Biochemical confirmation of serum vitamin levels at baseline or follow-up was not conducted, thus study cannot determine if already present deficiency was being corrected or not.

Future recommendations

• Longer follow-up period.

• Multi-center recruitment.

• Biochemical confirmation at baseline and follow-up to determine accurate status of vitamin levels among participants.

• Separate vitamin C and D supplement groups.

Conclusion

This study highlights the importance of micronutrient supplementation of vitamin C and D in addition to non-surgical periodontal therapy. This leads to a significant reduction in gingival inflammation in school-aged children compared with standard therapy alone. Vitamin C and D supplementation are effective adjunct strategies for managing gingival inflammation in the pediatric population, especially in areas with micronutrient deficiencies.

Authors’ contributions

Arooj ul Hassan (AH) conceived the study, designed the research methodology, data acquisition, and developed the manuscript. Asif Hanif (AsH) contributed to study design, data analysis, and statistical interpretation and supervised the project. Muhammad Hassan (MH) coordinated data collection, participant recruitment, and reviewed the manuscript and clinical assessments. Obaid Bajwa (OB) assisted with data acquisition, patient recruitment, and manuscript drafting. Muhammad Anas (MA) contributed to the interpretation of findings, drafting of the manuscript, and preparation of tables and figures. Muhammad Sohail Zafar (MSZ) provided critical revision of the manuscript, methodological input, and final approval of the version to be published.

Funding

This research did not receive funding.

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author.

Declarations

Ethics approval and consent to participate

Written informed consent was obtained from the parents or legal guardians of all participating children before enrollment in the study. Age appropriate written assent was also obtained from the children themselves. The study was conducted in accordance with the ethical principles of the Declaration of Helsinki (2013 revision).

Ethical Approval was taken from the IRB of the University Institute of Public Health, The University of Lahore. Ref No. IRB-UOL-FAHS/835II/2019, on 17-01-2019.

Consent for publication

Not Applicable.

Competing interests

The authors declare no competing interests.

Clinical Trial Number

ISRCTN, ISRCTN18356310, 05/08/2025