Use of a Selected Mixture of Probiotic Strains for Degrading Gluten During Digestion

NCT05798689 | Completed

• 1 other identifier: GlutProbiotic
• Study Type: interventional
• Target: 70
• Locations: 1 country
• Sites: 1

Brief Summary

Gluten intake spreads worldwide, being the major food protein consumed in the Western diets (up to 20 g gluten/d). But gluten has unique and unusual features. It resists the complete luminal digestion by gastric, pancreatic and intestinal brush border enzymes, and is susceptible to post-translational modification (deamidation) by mucosal transglutaminases. Apart from partial digestion, gluten per se has a negative impact on a consistent part of the worldwide population, which mainly results in the manifestations of celiac disease (CD) or other gluten-related disorders. This study will enable to test in vivo a novel multi-species probiotic that in vitro has proven to degrade gluten to non-immunotoxic peptides.

Conditions

Probiotics

Keywords

Gluten; Gut microbiome; Celiac disease

Outcome Measures

Primary Outcomes (5)

• Hydrolyzed gluten amount in feces by competitive Elisa R5 antibody (gluten ppm)

  The collection of faecal samples will be at baseline (T0); 10 days of GFD (T1); 4 days of 50 mg/day gluten intake (T2); 4 days of 1 g/day gluten intake (T3); 4 days of 3 g/day gluten intake (T4); 20 days of 10 g/day gluten intake (T5), of which 10 last days were the wash-out (T6).The gluten degrading efficiency of the probiotic will be assessed in all above mentioned time points with competitive Elisa using the R5 antibody for hydrolysed gluten (gliadin epitopes). The concentration of gliadin (ppm) will be then converted to gluten according to the multiplication factor (2) reported by the manufacturer.

  6 months

• Gut microbiome

  Fecal samples DNA from baseline, intervention period and wash out will be sequenced targeting the V4 region of 16s rRNA gene to evaluate the alterations of the gut microbiome and possible modulation in the PR arm due to the probiotic mixture administrated

  7 months

• Volatile compounds determination by GC-MS using SPME extraction

  Fecal samples at the beginning of intervention (T1), end of intervention (T5) and wash-out (T6) will be evaluated for their organic volatile compounds by GC-MS. Volatile compounds and short chain fatty acids composition will be compared between the intervention groups PL vs PR. 4-methyl-2pentanol (final concentration 1 mg L-1) will be used as an internal standard in all analyses, to quantify the identified compounds by interpolation of the relative areas versus internal standard area.

  6 months

• Short chain fatty acids determination by GC-MS using SPME extraction

  Fecal samples at the beginning of intervention (T1), end of intervention (T5) and wash-out (T6) will be evaluated for SCFA by GC-MS. SCFA compounds and short chain fatty acids composition will be compared between the intervention groups PL vs PR. A stock solution containing the mixture of SCFA standards (acetic acid, butyric acid, propionic acid, isobutyric acid and valeric acid) will be dissolved in ultrapure water to obtain a calibration curve ranging from 1 μg mL-1 to 250 μg mL-1.The calibration curve will be constructed by plotting the normalized peak area versus concentration of individual SCFA. The relative peak of SCFA in faecal sample will be integrated and the concentration of SCFA will be calculated by the calibration curve equation

  6 months

• Persistence and colonization ability of probiotic preparation by quantitative PCR (copy numbers)

  The persistence and colonization ability of the probiotic preparation will be evaluated by qPCR on cDNA and DNA at T1, T5 and T6. For each species belonging to the probiotic preparation, species- specific primers will be used. The qPCR results (cycle threshold, CT) will be converted in Copy Number (CN) based on standard curves previously constructed by using serial dilutions of DNA extracted from pure cultures. The CN and CN(Log) will be calculated based on DNA concentration and amplicon length. The standard curves will be obtained by CT and CN(Log) interpolation.

  2 months

Study Arms (2)

PR

ACTIVE COMPARATOR

Fifty healthy volunteers were randomly allocated in the probiotic arm. To eliminate residual traces of gluten and similar proteins from the faecal material, both groups underwent a gluten-free diet (GFD) from day-1 to day-10. After 10 days, gluten administration started. The increasing administration plan was as follows: 50 mg/day for 4 days; 1 g/day for subsequent 4 days; 3 g/day for subsequent 4 days; and 10 g/day (in this case, reintroducing an equivalent amount of wheat-based bread - 4 slices) for subsequent 20 days. At this stage (10 + 4 + 4 + 4 + 10 days = total of 32 days), the administration of the probiotic preparation was interrupted, with a period of 10 days of wash-out.

Combination Product: Probiotic administration and gluten

PL

PLACEBO COMPARATOR

Twenty healthy volunteers were randomly allocated in the placebo.To eliminate residual traces of gluten and similar proteins from the faecal material, both groups underwent a gluten-free diet (GFD) from day-1 to day-10. After 10 days, gluten administration started. The increasing administration plan was as follows: 50 mg/day for 4 days; 1 g/day for subsequent 4 days; 3 g/day for subsequent 4 days; and 10 g/day (in this case, reintroducing an equivalent amount of wheat-based bread - 4 slices) for subsequent 20 days. At this stage (10 + 4 + 4 + 4 + 10 days = total of 32 days), the administration of placebo preparation was interrupted, with a period of 10 days of wash-out.

Combination Product: Placebo administration and gluten

Interventions

Probiotic administration and gluten COMBINATION_PRODUCT

Probiotic preparation including multi-species strains of Lactobacillus and Bacillus. The probiotic was administered at baseline and interrupted after 32 days. At the same time the other arm received placebo. Gluten was provided after 10 days of gluten free diet in increasing amounts

PR

Placebo administration and gluten COMBINATION_PRODUCT

The placebo was administered at baseline and interrupted after 32 days. At the same time the other arm received probiotic. Gluten was provided after 10 days of gluten free diet in increasing amounts

PL

Eligibility Criteria

• Age: 18 Years - 65 Years
• Sex: all
• Healthy Volunteers: Yes
• Age Groups: Adult (18-64), Older Adult (65+)

You may qualify if:

• healthy individuals;
• adherent to Mediterranean diet

You may not qualify if:

• known medical disease;
• known digestive disease symptoms;
• known family history of celiac disease (CD);
• wheat allergy;
• and use of prescription medications (including antibiotics or probiotics in the previous 2 months)

Sponsors & Collaborators

• Free University of Bozen-Bolzano: lead
• Evonik Operations GmbH, Germany: collaborator

Study Sites (1)

Free University of Bolzano-Bozen

Bolzano, 39100, Italy

Location

Related Publications (1)

• De Angelis M, Siragusa S, Vacca M, Di Cagno R, Cristofori F, Schwarm M, Pelzer S, Flugel M, Speckmann B, Francavilla R, Gobbetti M. Selection of Gut-Resistant Bacteria and Construction of Microbial Consortia for Improving Gluten Digestion under Simulated Gastrointestinal Conditions. Nutrients. 2021 Mar 19;13(3):992. doi: 10.3390/nu13030992.

  PMID: 33808622 BACKGROUND

MeSH Terms

Conditions

Celiac Disease

Interventions

Glutens

Condition Hierarchy (Ancestors)

Malabsorption Syndromes; Intestinal Diseases; Gastrointestinal Diseases; Digestive System Diseases; Metabolic Diseases; Nutritional and Metabolic Diseases

Intervention Hierarchy (Ancestors)

Prolamins; Grain Proteins; Plant Proteins; Proteins; Amino Acids, Peptides, and Proteins; Seed Storage Proteins

Study Officials

• Olga Nikoloudaki, Ph.D

  Free University of Bolzano-Bozen

  PRINCIPAL INVESTIGATOR

Study Design

• Study Type: interventional
• Phase: not applicable
• Allocation: RANDOMIZED
• Masking: TRIPLE
• Who Masked: PARTICIPANT, CARE PROVIDER, INVESTIGATOR
• Purpose: SCREENING
• Intervention Model: PARALLEL
• Sponsor Type: OTHER
• Responsible Party: SPONSOR

Study Record Dates

• First Submitted: March 3, 2023
• First Posted: April 4, 2023
• Study Start: October 1, 2020
• Primary Completion: April 30, 2021
• Study Completion: May 31, 2021
• Last Updated: April 4, 2023

Record last verified: 2023-03

Locations

IT (1)