Description

Abbreviations

• GSRS-IBS: Gastrointestinal Symptom Rating Scale for Irritable Bowel Syndrome
• GIQLI: Gastrointestinal Quality of Life Index
• HADS: Hospital Anxiety and Depression Scale
• FSS: Fatigue Severity Scale
• LBP: Lipopolysaccharide-Binding Protein

Inclusion Visit (visit 0 \[V0\])

At the inclusion visit (V0) at Avicenne Hospital (Bobigny), participants will receive study instructions and materials. They will have up to 15 days to:

• Complete questionnaires (modified GSRS-IBS, Bristol Stool Scale, GIQLI, HADS, FSS)
• Collect a baseline stool sample, stored at -18°C at home until their first hospital visit (V1).

Wash-out phase (Day-15 \[D-15\] to Day 0 \[D0\]):

Participants will follow a gluten-free and low-fructan diet for 15 days. During this period:

• Daily food intake will be recorded in a diary, with meal photos sent to the research team via a dedicated email
• On D-8, participants will complete the modified GSRS-IBS and Bristol Stool Scale.
• On D-1, participants will perform a 24-hour urine collection after ingestion of lactulose (10g) and mannitol (10g) in 200 mL water, divided into three containers:
• First morning urine
• 0-5 h collection after sugar ingestion (fasting except water)
• 5-24 h collection Stool will also be collected on D-1 and stored at 4°C until V1.

First investigation day (Day 0 \[D0\] - V1):

At 9 a.m., participants arrive the Clinical Research Center (at Avicenne Hospital) with their stool and urine samples.

After completing the questionnaires (modified GSRS-IBS, GIQLI, HADS, FSS, and Bristol Stool Scale), a venous catheter will be inserted for blood sampling.

A 10 mL blood sample will be collected before the test meal. Participants will then consume the test meal (105 g of "artisanal" bread with water).

Postprandial blood samples (2 mL each) will be collected every 30 minutes for 3 hours (total blood volume: 22 mL) The modified GSRS-IBS questionnaire will be completed every hour during the postprandial period If a bowel movement occurs during the visit, the stool sample collected at that time may be used to replace the one collected the previous day.

Intervention phase (D1 to D14):

Participants will continue the same gluten-free, low-fructan diet, with the addition of the "artisanal" bread, which will serve as the sole source of gluten source and the main source of fructans.

• D1: 75 g of bread
• D2: 100 g of bread
• D3-14: 125 g of bread daily (≈ 8.5 g gluten, \<1.2 g fructans) Bread consumption may be distributed throughout the day. On D8, participants will again complete the modified GSRS-IBS and Bristol Stool Scale.

Second investigation day (D15 - V2):

On D14, participants will repeat the 24-hour urine and stool collection as previously described. On D15, they will return to the research center and undergo the same procedures as V1.

Measurements

• Questionnaires: modified GSRS-IBS (gastrointestinal symptoms), GIQLI, HADS, FSS.
• Blood: LBP (marker of inflammation), zonulin (marker of intestinal permeability), glucose, insulin.
• Urine: lactulose and mannitol, with calculation of the lactulose/mannitol ratio as a marker of intestinal permeability.
• Stool: microbial community analysis (microbiota) and fecal calprotectin (marker of inflammation). Baseline stool samples (before wash-out phase) will also be used to inoculate an artificial colon model for later investigation of the effects of bread on the microbiota.