Study design
We conducted a randomized, double-blind, placebo-controlled clinical trial to evaluate the efficacy of vitamin and mineral supplementation in patients with IBD in remission. Participants were randomly allocated in a 1:1 ratio to either supplementation with a tablet containing vitamin B1, vitamin B6, vitamin B12, and a magnesium oxide tablet, or a matching placebo, daily for 4 weeks to investigate its effect on fatigue, disease activity, and HRQoL in patients with IBD.
Setting
This intervention was a single-center study. Eligible participants were recruited from an outpatient gastroenterology clinic in Tehran, Iran, from June 2023 to February 2024. A trained nutritionist was responsible for administering all questionnaires and for monitoring participants throughout the study period.
Participants
A total of 98 adult patients with IBD in remission were enrolled based on the endoscopic remission and specialist diagnosis. Inclusion criteria were age between 18 and 65 years, a confirmed diagnosis of IBD based on specialist evaluation, medical history, clinical symptoms, laboratory findings from stool and blood samples, and radiology investigations, including colonoscopy and endoscopy. Participants were required to have no evidence of pregnancy, breastfeeding, or anemia, and to have a body mass index (BMI) within the normal to overweight range (18–29.9 kg/m2) due to the effect of obesity on fatigue. Obese patients were excluded30.
Randomization and masking
Participants were randomly allocated in a 1:1 ratio to either supplementation of a tablet containing vitamin B1, vitamin B6, vitamin B12, and a magnesium oxide tablet or a placebo daily for 4 weeks. Block randomization was performed by an independent statistician at Iran University of Medical Sciences using a computerized random number generator. A blinded study coordinator labeled and packaged the supplements and placebo. Both participants and investigators were blinded to group allocation.
Procedures
Participants received two oral tablets daily for 4 weeks. The intervention group received a tablet containing vitamins B1 (Thiamine), B6 (Pyridoxine), B12 (Cyanocobalamin) (manufactured by Mahban Darou Co), and one tablet of magnesium oxide (manufactured by Jalinus Co). The dosing of the supplement was 100 mg vitamin B1, 100 mg vitamin B6, and 200 µg vitamin B12. Additionally, patients received a 250 mg magnesium oxide tablet daily. The placebo group received two tablets containing cornstarch, identical in appearance to the supplements. Each participant received two bottles containing an equal number of tablets. Compliance was assessed by pill count at the end of the intervention period, and adherence to study supplementation was > 80% of the tablets. No adverse events were observed during the study.
Measurements
Demographic data were gathered using a questionnaire designed by the principal investigator. HRQoL was evaluated using the Inflammatory Bowel Disease Questionnaire (IBDQ-9), which measures gastrointestinal, systemic, emotional, and social impacts of IBD17. The IBDQ-9 is a shortened, validated quality-of-life instrument specific to IBD. It contains nine items rated on a 7-point Likert scale (1 = worst function, 7 = best function). Total scores range from 9 to 63, with higher scores indicating better HRQoL. It is has been validated in Iranian population31.
Disease activity in patients with UC was assessed using the Patient-reported Simple Clinical Colitis Activity Index (P-SCCAI). This questionnaire has been validated to assess symptoms, severity, and activity of UC. The questions included in this questionnaire cover topics such as the frequency of bowel movements, the presence of blood in stool, overall health status, urgency to defecate, and non-gastrointestinal symptoms experienced during the past week32. The sum gives a total score (typically 0–19 in traditional scoring), with higher scores indicating more active disease. Values above predefined thresholds (for example, ≥ 5 in some studies) are used to indicate active disease33.
For patients with Crohn’s disease, disease activity was measured using the Simplified Crohn’s Disease Activity Index (SCDAI), which is a validated questionnaire used to assess the symptoms, severity, and activity of CD. The questionnaire includes questions regarding the frequency of loose or watery stools, abdominal pain, general health status, use of antidiarrheal medications, extra-intestinal manifestations, the presence of abdominal masses, hematocrit levels, and body weight. The scores assigned to the responses are summed, and based on the total score, disease activity is assessed. The score range is from 0 to 600. A score below 150 indicates a phase of disease improvement, 150 to 219 indicates mild disease, 220 to 450 indicates moderate disease, and a score above 450 indicates severe disease34.
The Inflammatory Bowel Disease Fatigue Scale (IBD-F) questionnaire was used to assess the severity and impact of fatigue on patients with IBD. The fatigue score is measured using a Likert scale from 0 to 4 for each question. The questionnaire has three sections, and we used Sects. 1 and 2 in this study. Section 1 evaluates the severity of fatigue, while Sect. 2 examines the impact of fatigue on these patients. The total score in Sect. 1 ranges from 0 to 20, and a score of 0 indicates no fatigue, rendering further response unnecessary. The second section includes a score range of 0 to 120 (questions 3, 4, 9, 12, 13, and 14 may be omitted)35.
Variables
The primary endpoint was improvement in fatigue score after 4 weeks of supplementation compared with placebo. Secondary endpoints were improvements in HRQoL, disease activity for UC and CD, and physical activity level evaluated with the IPAQ. Independent variables consisted of the intervention group (supplementation vs. placebo) and intake of micronutrients, including vitamins B1, B6, B12, and magnesium. Covariates and potential confounders included age, sex, body mass index (BMI), duration of disease, baseline physical activity, and concurrent medication use.
Sample size
Sample size was determined a priori using a pilot study of 30 participants (15 per group) to obtain a realistic estimate of the variability of the primary outcome and to inform the expected effect size. The primary endpoint was fatigue (IBD-F) analyzed as a continuous outcome, and the primary treatment effect was defined as the between-group difference in mean change from baseline to week 4 (i.e., the time × group interaction, equivalent to comparing individual change scores between arms). Assuming equal allocation, the required sample size per arm was calculated using the standard two-sample formula for detecting a difference in mean change:
\(\:n\text{\hspace{0.25em}\hspace{0.05em}}=\text{\hspace{0.25em}\hspace{0.05em}}\frac{2\text{\hspace{0.17em}}({z}_{1-\alpha\:/2}+{z}_{1-\beta\:}{)}^{2}\text{\hspace{0.17em}}{\sigma\:}_{{\Delta\:}}^{2}}{{\delta\:}^{2}},\) where \(\:\delta\:\)is the anticipated between-group difference in mean change, \(\:{\sigma\:}_{{\Delta\:}}\)is the standard deviation of the change scores, \(\:\alpha\:\)is the two-sided type I error, and \(\:1-\beta\:\)is power. Based on the pilot data, we planned for a clinically meaningful effect of \(\:\delta\:=5\) points and used \(\:{\sigma\:}_{{\Delta\:}}=7\), with α = 0.05 (two-sided) and 80% power, yielding a requirement of 32 participants per group (64 total). Allowing for 10% attrition/missing post-intervention data, the final target sample size was inflated to 36 participants per group (72 total). As a conservative sensitivity check, the SD of change was also derived from baseline/post SDs and their correlation using \(\:{\sigma\:}_{{\Delta\:}}=\sqrt{{\sigma\:}_{0}^{2}+{\sigma\:}_{1}^{2}-2r{\sigma\:}_{0}{\sigma\:}_{1}}\); with \(\:{\sigma\:}_{0}={\sigma\:}_{1}=9\) and \(\:r=0.60\), \(\:{\sigma\:}_{{\Delta\:}}\approx\:8.05\), resulting in 42 participants per group (84 total) and 47 per group (94 total) after 10% attrition.
Statistical analysis
All statistical analyses were performed according to a prespecified analysis plan with a two-sided significance level of 0.05. Continuous variables were summarized as mean ± standard deviation (SD) when approximately normally distributed, or as median [interquartile range (IQR)] when distributions were skewed. Categorical variables were reported as number (percentage). Baseline comparability between the intervention and placebo groups was assessed using the independent t-test or Mann–Whitney U test for continuous variables, as appropriate, and Fisher’s exact test for categorical variables. Within-group pre–post changes were evaluated using the Wilcoxon signed-rank test, and unadjusted between-group differences in change were examined using the Mann–Whitney U test.
The primary endpoint was the change in fatigue from baseline to week 4, assessed using the IBD-F Sects. 1 and 2. Secondary endpoints included changes in health-related quality of life (IBDQ-9), disease activity in ulcerative colitis (P-SCCAI), disease activity in Crohn’s disease (SCDAI), and physical activity (IPAQ). The primary adjusted analyses were conducted using linear mixed-effects models with a random intercept for participant to account for within-subject correlation over time. Time was modeled as a binary indicator (0 = baseline, 1 = week 4), and fixed effects included study group (intervention vs. placebo), time, and the time × group interaction, with the interaction term representing the primary treatment effect (i.e., the between-group difference in change over time). Because baseline medication category differed significantly between treatment arms, medication category (drug type) was included as an adjustment covariate in all mixed-effects models. To enhance robustness to model misspecification and potential heteroscedasticity, robust (cluster-robust) standard errors (CR2) with Satterthwaite degrees of freedom were used to obtain 95% confidence intervals and p-values. Effect estimates are reported as β coefficients with 95% confidence intervals (CIs). Analyses were performed using all available observations, and participants with missing post-intervention values for a given outcome contributed baseline data but were not included in outcome-specific change calculations for which follow-up measurements were unavailable.
Ethics information
The trial was conducted in accordance with the Declaration of Helsinki, and all participants signed written informed consent at baseline. This randomized controlled trial (RCT) was registered in the Iranian Registry of Clinical Trials (IRCT20090822002365N28) on 29 April 2023 with ethics approval no. IR.IUMS.REC.1401.951 from the ethics committee of the Iran University of Medical Sciences.