Does propolis affect the quality of life and complications in subjects with irritable bowel syndrome (diagnosed with Rome IV criteria)? A study protocol of the randomized, double-blinded, placebo-controlled clinical trial

Considering to the tight relationship between inflammation, oxidative stress, dysbiosis of the gut microbiota, and alteration in the intestinal permeability with IBS, it seems that propolis supplementation would contribute to improve the health of the gastrointestinal system in patients with IBS. Most clinical trials reported the safety and efficacy of propolis supplementation on glycemic control in patients with type 2 diabetes mellitus [22]. To the best of our knowledge, there is no clinical trial investigating the effect of propolis in patients with IBS.

The current clinical trial aims to evaluate the effect of propolis supplementation on the IBS Severity Index (IBSSI). Also, secondary objectives of this trial include the changes in weight, body mass index (BMI), waist circumference (WC), IBS quality of life (IBS-QOL), and anxiety with propolis supplementation compared to placebo. For controlling of confounder factors, changes in dietary intakes, physical activity levels, and anxiety will be assessed in the current trial.

The current study is a prospective, single-center, randomized, double-blind, placebo-controlled clinical trial evaluating the efficacy of propolis supplementation in subjects with IBS. This clinical trial will be performed at the gastrointestinal clinic, a referral center for gastrointestinal diseases in Ahvaz, the southwest of Iran. All volunteers will provide written informed consent before participation in this randomized clinical trial.

The current trial is registered at the Iranian Registry of Clinical (ID: IRCT20190708044154N1). The study protocol was drafted and developed in accordance with the Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT) 2013 checklist (Additional file 1). The protocol flow chart and timeline of the study are shown in Fig. 1 and Table 1, respectively. A protocol deviation is accidental and unintentional changes to the study protocol which have no significant impact on the study’s risk or benefit, subject’s rights, safety or welfare, and the integrity of the data [23]. The protocol deviations may result from the action of the patients or researchers. Examples of the protocol deviation for this study include:

Any amendments of this study protocol which have related to safety and benefit of participants will need to be agreed upon by Department of Clinical Nutrition in Tabriz University of Medical Sciences and approved by the Medical Ethics Committee of Tabriz University of Medical Sciences, Tabriz, Iran, before the implementation of the study. Any change in the study protocol will be sent to Trial Journal (www.​trialsjournal.​biomedcentral.​com).

This clinical trial will be performed at the gastrointestinal clinic, a referral center for gastrointestinal diseases in Ahvaz, the southwest of Iran. All participants will be informed of this study through advertisements on the clinic’s television.

Volunteers will be recruited from outpatients referred to the gastrointestinal clinic in Ahvaz, Iran, through advertisement on the clinic’s television. Patients who meet the international Rome IV criteria for diagnosing IBS will be assessed for eligibility criteria, after differentiating IBS from structural bowel disorders by an expert gastroenterologist (Dr. PM) at this clinic. The international Rome IV diagnostic Questionnaire is the well-known and latest diagnostic instrument in the field of IBS in clinical researches. According to Rome IV criteria, individual who had recurrent abdominal pain or discomfort at least 1 day/week in the last 3 months with symptom onset at least 6 months before a diagnosis has IBS if she/he had at least two of the following criteria: symptom improvement with defecation, onset simultaneous with a change in the frequency, or/and onset simultaneous with a change in the form or appearance [3].

The inclusion criteria will consist of patients who have an age between 18 and 65 years, who have an IBS-C or IBS-M (much more common forms of IBS) according to the BSFS, and who fill out a written informed consent to participation in this trial.

The exclusion criteria will consist of patients who are a pregnant or breastfeeding, who have/had a malignancy or other chronic digestive diseases such as IBD, who have regular use of drugs that modify GI movements (such as metoclopramide, cisapride, narcotics, and diphenoxylate), who have regular use of laxatives or antibiotics, who have regular use of the prebiotic or probiotic supplement, who have regular use of antidepressant drugs, who have a history of major surgery in the gastrointestinal tract (Billroth’s operation, having an ostomy and any resection of any part of the digestive tract), who underwent psychotherapy, who have a restricted diet (such as vegetarian diets), and who have/had an allergy to bee products.

Participants who are unwilling to continue collaborating on the study, have the sensitivity to propolis supplement, become pregnant, alter the patient’s disease to other gastrointestinal diseases, or have poor compliance with the assigned treatment (less than 80%) will be withdrawn from follow-up.

Stratified permuted block randomization will be used to balance gender and IBS types between two groups over time. Four strata is constructed according to sex and IBs types as follows:

Within each stratum mentioned above, eligible patients will be randomly allocated in a ratio of 1:1 to receive propolis or placebo with a block size of 4. A block size of 4 is constructed six different arrangements (block) to assign patients to the trial groups as follows equally:

The sequence of the blocks mentioned above will be prepared for each stratum using a random numbers table. The study pharmacist will provide the placebo tablet similar to propolis tablet in color, odor, taste, shape, size, and weight, and they will be held in drug containers. All containers will be the same in terms of shape, color, odor, size, and weight and will be kept inside consecutively numbered, opaque, sealed envelopes, which are completely impermeable to light.

The trial-group assignments will be concealed from patients, physician, and investigators (enrolling, assessing, and analyzing) until the end of the study and data analysis, with the exception of the study pharmacist. The study pharmacist will provide a randomization list, sequentially numbered drug containers, and opaque and sealed envelopes.

Fifty-two eligible patients will be randomly assigned to either the intervention group (n = 26) or the control group (n = 26). Patients in the intervention group will receive an identical propolis tablet containing 450 mg Iranian green propolis extract (100 mg polyphenol compounds and 67 mg flavonoids) twice a day, one tablet before lunch and one tablet before dinner, for 6 weeks. Propolis sample is obtained from honey bee colonies located in Rasht, the north area of Iran. Patients in the control group will receive an identical placebo tablet twice a day, before lunch and dinner, for 6 weeks. Propolis and placebo tablets will be similar in color, odor, taste, shape, size, and weight and will be produced by the School of Pharmacy, Mashhad University of Medical Sciences, Iran. Patients will be asked to continue their usual diet, physical activity, and medicines until intervention completion. Also, they will be instructed on how to use their supplements. Patients will be followed through weekly phone calls and intermediate visits (t₃) to the clinic for assessing treatment compliance and adverse events. At the end of the trial (t₇), participants will be informed of the usual dietary recommendations of IBS.

The primary outcome of this clinical trial is the change in the severity of IBS from baseline to the sixth week of the intervention. The secondary outcomes of this clinical trial are the change in IBS quality of life, weight, and waist circumference from baseline to the sixth week of the intervention.

Demographic variables, including sex, age, race, education, income, marital status, smoking status, housing status, employment status, medical history, and family history, will be obtained from each participant at the beginning of the study (t₀). Besides, for controlling of confounder factors, changes in dietary intakes, physical activity level, and anxiety will be evaluated in the current trial.

At the beginning of the trial, demographic information, food intakes, physical activity, weight, height, BMI, waist circumference, anxiety, IBS quality of life, and IBS severity will be collected by validated tools.

We used Hern’s single-stage phase II methodology to calculate the sample size [34]. Based on results from other dietary supplements in patients with IBS [35] in which 15% of patients in the placebo group (P₀) and 40% of patients in the intervention group (P₁) had complete remission from symptoms of IBS, we need 22 patients in each trial group with a power of 80%. To compensate for withdrawal during follow-up, we increase the final sample size by 20%, to 26 patients in each group.

In this study, the dose of propolis supplementation was extrapolated from the study conducted by Sabuncuoglu et al. [18] in which the administration of propolis at a dose of 100 mg kg⁻¹ improves the function of the GI tract and gut microbiota in rats. Based on the following formula for the conversion of animal dose to human [36] and reference body weight of 60 kg, the dose of propolis for a human is approximately 900 mg. At this dose of propolis, no adverse events were reported in the previous clinical trials.

Patients will be asked to avoid taking the assigned supplement following the occurrence of adverse events and to immediately call the study physician or other investigators. One of the study researchers will be instructed to call the participants weekly by phone and ask them about experiencing any adverse events. Tabriz University of Medical Sciences is responsible to follow up on any adverse events and reports them to the Medical Ethics Committee of Tabriz University of Medical Sciences for decision making. If the reported adverse events are related to the consumed supplements, the study will be stopped and patients will be referred to a specialist for immediate treatment in which can be treated free of charge. Participants who are unwilling to continue collaborating on the study will be able to discontinue the trial at any time.

One of the researchers will check the coding, security, and storage of data. Also, she/he will check data entry, and data values double times. If any patient reports the occurrence of adverse events, more information is needed for the decision making about excluding the patient from the study.According to the Medical Ethics Committee Criteria, unblinding is permissible in this situation, and we can specify whether the patient has received the propolis or placebo.

All patients will be instructed on how to use their supplements at the baseline, each phone call, and intermediate visits (t₃). All patients will be asked to complete a daily calendar immediately after consumption of their assigned treatments. The adherence to intervention refers to the degree to which the behavior of trial participants corresponds to the intervention assigned to them. Adherence to the assigned treatments will be checked by means of a daily calendar at each phone call and will be rechecked by counting the returned tablets at the end of the trial (t₇). Compliance will be calculated based on the following formula:

The compliance less than 80% will be classified as poor compliance [37].

We will use the following measures to ensure minimal loss to follow-up:

The SPSS software (Version 16, SPSS Inc., and Chicago, IL, USA) will be used for statistical analysis. Data will be reported as the mean and standard deviation for normally distributed variables, the median and interquartile range for non-normally distributed variables, and frequencies for nominal variables. We will use the Kolmogorov-Smirnov test to determine the distribution of quantitative data. Chi-square test and Fisher’s exact test will be used to determine differences in frequencies of nominal variables. We will use the independent sample t test and Wilcoxon rank-sum test for within-group comparisons and paired sample t test and Mann-Whitney U test for between-group comparisons. We will also use the analysis of covariance (ANCOVA) test to adjust the potential confounding factors, such as a change in anxiety levels, physical activity, and intake of energy and nutrients, especially for FODMAPs, during the study period.

The binary logistic regression test will be used to calculate the multivariable-adjusted odds of improvement in the primary outcome with propolis supplementation. According to the patterns of missing data, a suitable multiple imputation approach will be followed for completing missing data. We will use an intention-to-treat analysis as our primary analytic approach. Linear mixed model ANOVAs may be used as a substitute statistical analysis method [38]. Statistical differences will be shown by P values and 95% confidence intervals. A P value < 0.05 will be considered statistically significant.

Not applicable.

Each propolis tablet contains 450 mg Iranian green propolis extract, including 100 mg polyphenol compounds and 67 mg flavonoids. Propolis sample is obtained from honey bee colonies located in Rasht, the north area of Iran. The propolis and placebo will have the same shape, color, odor, taste, size, Lot number, and container and will be produced by the School of Pharmacy, Mashhad University of Medical Sciences, Iran. Also, all containers will be the same in terms of shape, color, odor, size, and weight and will be kept inside consecutively numbered, opaque, sealed envelopes.

Enrollment; protocol version: 2 on 13 February 2020; the recruitment began on 31 July 2019 and will be approximately completed on 18 June 2020.