Effect of Daily Consumption of a Novel Biscuit Enriched With Edible Mushrooms, on Intestinal Health-related Parameters

Study Overview
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Brief Summary
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The purpose of the study is to assess the effect of dietary intervention that includes the daily consumption of a novel biscuit (enriched with the selected mushrooms rich in β-glucans) on gut health related parameters of healthy subjects over 60 years old.

Detailed Description
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After having been informed about the study and the potential risks, all subjects giving written informed consent will undergo screening to determine eligibility for study entry (inclusion criteria). At baseline, the participants will provide biological samples and they will be randomly assigned in a double-blind manner (investigator and participant) to one of the intervention groups, i.e. consumption of the novel biscuit (biscuit enriched with mushroom powder containing 3g of β-glucans) or consumption of the placebo biscuit for 3 months,. After a washout period of 2 months, the subjects will consume the novel biscuit or the placebo biscuit in a cross-over design for 3 more months. Throughout the interventional period subjects' dietary intake, anthropometrical data and gastrointestinal symptoms will be monitored. Meanwhile at baseline and at the end of each trimester physical activity, mental health and sleep habits will be evaluated with the use of questionnaires and biological samples will be provided for further analysis.

Official Title
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Development of a Novel Biscuit Enriched With β-glucans Isolated From Edible Mushrooms of Greek Habitats and Implementation of a Dietary Intervention That Includes Daily Consumption of This Biscuit

Conditions
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Healthy

Intervention / Treatment
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* Other: Novel biscuit enriched with mushroom powder containing 3g of β-glucans
* Other: Placebo biscuit


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: healthy individuals aged 60-80 years old, of both sexes/genders Exclusion Criteria: no recent weight loss and extreme dietary behaviors; no history of gastrointestinal disease, chronic constipation, chronic/acute diarrhea, autoimmune disease, coronary disease, liver and/or kidney malfunction; no consumption of antibiotics two months prior to the initiation of the intervention; no consumption of probiotics and/or prebiotics and/or dietary fiber supplements two weeks prior to the initiation of the intervention.

Ages Eligible for Study
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Minimum Age: 60 Years
Maximum Age: 80 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Prevention
Allocation: Randomized
Intervention Model: Crossover Assignment
Masking: Triple


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: Biscuit enriched with mushroom powder<br>Daily consumption of a novel biscuit enriched with mushroom powder containing 3g of β-glucans | Other: Novel biscuit enriched with mushroom powder containing 3g of β-glucans<br>* Daily consumption of a novel biscuit enriched with mushroom powder containing 3g of β-glucans<br>|
| Placebo Comparator: Placebo biscuit<br>Daily consumption of a placebo biscuit | Other: Placebo biscuit<br>* Daily consumption of placebo biscuit<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change from baseline in Gastrointestinal tolerance of novel biscuit during the interventional period and at 3 months. | Gastrointestinal (GI) symptomatology will be recorded through a 7d-questionnaire in certain time periods.The intensity of each GI symptom (abdominal pain, distension, ﬂatulence and borborygmi) will be measured daily on a scale of 0-4, where '0' represents absence of symptoms and '4' severe symptoms. The possible range for each weekly symptom score is 0-28, and for the total symptom score 0-112. Frequency and consistency of evacuations will be also noted, and presence of diarrhoea will be defined. | Gastrointestinal symptoms will be evaluated at baseline, during and at the end of each intervention. (3 months period) | 
| Change from baseline to the effect on the intestinal microbial populations and their metabolic products [i.e. Short-Chain Fatty Acids (SCFA)] (prebiotic effect) at 3 months. | Microbial populations will be quantified with 16SRNA sequencing and quantitative Polymerase Chain Reaction (PCR). SCFAs will be quantified with Gas chromatography. | The levels of microbial populations and their metabolic products e.g. SCFA will be measured at baseline and at the end of each interventional period.(3 months period) | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change from baseline in Metabolic health at 3 months. | Blood samples will be obtained after an overnight, 12-h fasting. Blood serum analysis will include assessment of lipidemic profile [i.e. Total Cholesterol (TC), Low-Density Lipoprotein Cholesterol (LDL-C), High-Density Lipoprotein Cholesterol (HDL-C), Triglycerides (TG)], glucose metabolism [i.e. Fasting Blood Glucose (FBG)] and uric acid. (units of measure: mg/dL) | Metabolic health will be measured at baseline and at the end of each interventional period.(3 months period) | 
| Change from baseline in blood serum Insulin at 3 months. | Blood samples will be obtained after an overnight, 12-h fasting. Blood serum analysis will include assessment of glucose metabolism [i.e. Insulin (INS)]. (units of measure: μIU/mL) | Blood serum Insulin will be measured at baseline and at the end of each interventional period.(3 months period) | 
| Change from baseline in blood serum 25-hydroxy vitamin D at 3 months. | Blood samples will be obtained after an overnight, 12-h fasting. Blood serum analysis will include assessment of bone biomarkers (i.e. 25-hydroxy vitamin D).(units of measure: ng/mL) | Blood serum 25-hydroxy vitamin D will be measured at baseline and at the end of each interventional period.(3 months period) | 
| Change from baseline in blood serum Parathyroid Hormone at 3 months. | Blood samples will be obtained after an overnight, 12-h fasting. Blood serum analysis will include assessment of bone biomarker [i.e. Parathyroid Hormone (PTH)].(units of measure: pg/mL) | Blood serum Parathyroid Hormone will be measured at baseline and at the end of each interventional period.(3 months period) | 
| Change from baseline in Immune system reinforcement at 3 months. | The production of Interleukins (IL) (i.e. IL-1β, IL-6, IL-10) and Tumor-necrosis factor alpha (TNF-a) in vitro by peripheral blood mononuclear cells (PBMCs) cultured and isolated from healthy immunocompetent subjects, after mitogen stimulation, will be determined in cell culture supernatants by ELISA techniques with commercially available kits. The cytokine expression levels will be determined in cell pellets by Quantitative Real Time Polymerase Chain Reaction (qRT-PCR). Concomitantly, the levels of the same cytokines will be assessed by ELISA in the serum collected from the same subjects. | Immune system reinforcement will be measured at baseline and at the end of each interventional period.(3 months period) | 
| Change from baseline in Dietary intake at 3 months. | Dietary intake will be evaluated with the use of 3-days (2-weekdays and 1 weekend day) diet intake records. | Dietary intake will be measured at baseline and at the end of each interventional period.(3 months period) | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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β-glucans, edible mushrooms, functional food, prebiotic activity, gut microbiota, older adults