The goal of this project is to investigate whether cellobiose, a type of sugar, has a beneficial effect on gut health when consumed by humans. Cellobiose will be obtained from spent coffee grounds through enzymatic modification, repurposing waste biomass for a potentially valuable purpose. The study will only focus on conducting in vivo human trials using a commercial form of cellobiose from Savanna Ingredients GmbH, which has recently been approved as a novel food by the EU (https://eur-lex.europa.eu/eli/reg\_impl/2023/943/oj)

We will also be testing two other commercial products: Orafti® P95, an oligofructose-based product from Beneo GmbH, and maltodextrin from Special Ingredients Limited. These products will be compared to evaluate their potential in promoting gut health by nourishing beneficial bacteria, The study will involve feeding these products to volunteers and observing their impact on gut health.

The key questions this project aims to address include:

• Does cellobiose, exhibit any gut health promoting effects in humans?
• How does cellobiose compare to other commercial prebiotic products, such as Orafti® P95, in terms of its effects on gut health?

To conduct the study, volunteers will be provided with the three different products to consume over a specified period. Their faecal samples will be collected at various points to assess changes in gut microbiota composition and other indicators of gut health. This will involve close monitoring of participants' health and adherence to ethical guidelines to ensure their safety and well-being throughout the study.

Overall, this project seeks to expand our understanding of the potential health benefits of cellobiose and other prebiotic carbohydrates. It aims to provide valuable insights into the role of these substances in promoting gut health.

In this randomised, double-blinded, parallel-group trial, the investigators will enroll 36 healthy adult volunteers and randomise them 1:1:1 to one of three dietary-supplement arms. Participants will consume their assigned product once daily-5 g during a one-week adaptation phase followed by 10 g/day for a three-week intervention phase. Faecal samples will be collected at baseline (Visit 1) and at study completion (Visit 5). Total bacterial counts will be determined by fluorescence in situ hybridisation coupled with flow cytometry (FISH-FLOW), and short-chain fatty acid concentrations quantified by gas chromatography-flame ionisation detection (GC-FID).

The primary outcomes are the change from baseline to Visit 5 in (1) absolute abundance of Bifidobacterium spp. and Lactobacillus spp., and (2) total faecal short-chain fatty acid concentration. Secondary outcomes include gut microbiota diversity metrics (α- and β-diversity) and daily gastrointestinal symptom incidence. Exploratory outcomes will assess correlations between shifts in specific bacterial taxa and metabolite profiles, as well as changes in the abundance of other key genera.

Inclusion Criteria:

Inclusion criteria:

1. Participants aged 18 years old and above
2. Participants are willing to comply with the study instructions
3. Participants have a stool frequency of at least 3 bowel movements per week
4. Participants are healthy at the time of signing the consent form

Exclusion criteria:

1. Not pregnant or planning to be pregnant
2. Previously or currently diagnosed gastrointestinal diseases and disorders that might affect the gut environment.
3. Food allergies or intolerances
4. Previously or currently diagnosed with cancer or taking a long-term medication.
5. Use of medication (e.g, antibiotics, aspirin, proton pump inhibitors) influencing gastrointestinal function (8 weeks before intervention)
6. Participants are currently involved or will be involved in another clinical or food study

De-identified individual participant data underlying the primary and secondary outcomes (microbiome absolute counts, SCFA concentrations, GI symptom diaries), along with the study protocol and statistical analysis plan, will be made available to qualified researchers beginning six months after publication. Requests can be submitted to the corresponding author (r.a.rastall@reading.ac.uk) and will require a signed data-use agreement. Data will be shared in CSV format.

Six months after publication, with data available for up to five years thereafter.

The data will be available to qualified researchers who submit a reasonable research proposal and execute a data-use agreement.