The effect of non-caloric sweeteners compared with sucrose on the gut microbiome, cardiovascular risk factors, and sex hormones in women with obesity

This study aimed to investigate the effect of non-caloric sweeteners compared with sucrose on the gut microbiome, cardiovascular risk factors, and sex hormones in women with obesity.

A 4-arm triple-blind, randomized parallel controlled clinical trial. A total of 160 eligible participants will be assigned to four intervention groups (n=40) using block randomization.

This study will be conducted in the Nutrition and Food Security research center, Shahid Sadoughi University of Medical Sciences, Yazd, Iran. The intervention period will last 3 months. The four tested beverages will be provided in the same sweetness, shape, color, and appearance and will be packed in the same bottles. All bottles will be labeled with four codes (A, B, C, and D) by a person who is not aware of the study objectives. Therefore, participants and administrators will be blinded to the content of the bottles until data analyses. Physical activity assessment, three-day weighted food record, anthropometric and blood pressure measurements will be conducted at the baseline, mid-intervention, and the end of the intervention period, and blood samples, 24- hour urine samples, and stool samples will also be collected from participants at the start and the end of each intervention period.

Women with obesity who are experiencing menopausal transition (aged 42-52 years) without any other chronic diseases who do not smoke or drink alcohol, experience infection or antibiotic use, and give consent to enter the study

After a 4-week run-in period, 160 eligible women will be randomly assigned to four groups (n=40) in which they will receive daily a 500mL bottle containing sucrose (50 g), or aspartame (268 mg), or saccharin (125 mg), or steviol glucosides (200 mg).

Gut microbiome, Cardiometabolic markers, Sex hormones

An independent statistician will use a statistical software [statistical package for social sciences (SPSS)] which generates random numbers to block-randomize participants into one of the four trial groups (labeled as A, B, C, and D), ensuring that all the subjects will receive treatments in a balanced and random order. Each label will be assigned to one of the four sweeteners used as treatment (aspartame, saccharine, steviol glycosides, and sucrose).

The concealment will be done by putting the randomly assigned group for each participant in sealed envelopes. The allocation concealment will be conducted by an independent researcher. The four tested beverages will be provided in the same sweetness, shape, color, and appearance and will be packed in the same bottles. In addition, natural flavorings with similar color, appearance, taste, and texture will be added to all of the four treatment groups by an independent researcher from the Department of Pharmaceutics, School of Pharmacy, Shahid Sadoughi University of Medical Sciences. Participants and administrators will be unaware of the content of the bottles until data analyses. All bottles will be labeled with four codes (A, B, C, and D) by a responsible person who is not aware of the study objectives. The codes will not be released until after the statistical analyses; therefore, neither the study participants nor the personnel and the data analysts will be aware of the intervention bottles.

We plan to register and deposit data that relates to individual primary publications and not the whole study database. This enables other researchers to replicate published analyses and results as well as to re-use the data for additional analyses such as meta-analysis. General characteristics of participants (without the identity of the participants) as well as baseline and after-treatment data values of primary and secondary outcome variables, will become available for researchers after a request from interested investigators.

Data will be available after the individual, related papers become published.

Data will become available for all investigators interested in the subject.

All efforts will be made to protect the privacy and to de-identify the data. Therefore, data will be shared on request under the following conditions: - A meaningful study question by the requester - Outline of the planned analyses - Valid methodology - Signed data sharing agreement that contains a confidentiality agreement and approval of the study team members.

The data can be obtained from principal investigators: - Principal investigator from Iran: Dr. Amin Salehi-Abargouei, MSc, Ph.D. in Nutrition. Department of Nutrition Faculty of Public Health Sahid Sadoughi University of Medical Sciences Yazd, PO Code 8915173160 Iran, Tel:+98-35-31492229, Fax:+98-35-38209119, Email: abargouei@ssu.ac.ir Alternate email: abargouei@gmail.com - Principal investigator from Switzerland: Prof. Oscar Franco, MD, Ph.D., FESC, FFPH. Professor of Epidemiology & Public Health. University of Bern, Mittelstrasse 43, 3012 Bern, Switzerland, Tel +41316313350 Email: oscar.franco@ispm.unibe.ch

The interested investigators should provide a brief description of the data needed and their reason for asking for the data by contacting principal investigators. They will assess the possibility of sharing the data and if applicable will ask for a signed data sharing agreement that contains a confidentiality agreement and approval of the study team members.