Short-Chain Fatty Acids in Lypopolysaccharide-Induced Inflammation and Executive Function

Investigating the Role of Short-Chain Fatty Acids in Endotoxemia-Induced Inflammation and Core Executive Functioning: A Randomized, Triple-Blind, Placebo-Controlled Trial

Short-chain fatty acids (SCFAs) are substances produced by gut bacteria when they ferment dietary fiber. They can act as signals between the gut and the brain and may help regulate the body's immune and stress responses. Earlier work has shown that delivering SCFAs to the colon can lower the stress hormone response to a mental stress task in healthy people, and laboratory studies suggest SCFAs can reduce inflammation. However, it is not yet known whether SCFAs can reduce inflammation in humans, or whether doing so protects mental performance.

This study uses a controlled, acute, and short-lasting challenge. Healthy adults receive a single, low dose of a bacterial substance called lipopolysaccharide (LPS) through a vein. LPS briefly activates the immune system and causes mild, flu-like symptoms (such as fatigue, headache, and feeling unwell) that pass on their own within a few hours. It does not cause a real infection.

Participants are randomly assigned to take either SCFA capsules or inactive placebo capsules on the morning of the test day, before the LPS challenge. Neither the participants, the researchers running the visit, nor the researchers analyzing the results know who received which capsules until the study is complete (triple-blind).

The main goal of the study is to test whether SCFAs, compared with placebo, reduce the inflammatory response to LPS, measured by markers of inflammation in the blood. The study also examines whether SCFAs reduce the associated sickness symptoms and whether they protect performance on tasks that measure core executive functions: the ability to keep relevant information in mind (working memory), the ability to hold back automatic responses or thoughts (inhibition), and the ability to switch flexibly between tasks or rules (cognitive flexibility). Blood, saliva, and stool samples are collected to measure inflammation markers, SCFA levels, stress hormones, and gut bacteria.

Participants attend a screening visit, a test day with monitoring for several hours, and a short follow-up visit the next day.

Study Overview

• Status: Recruiting
• Conditions: Inflammation; Endotoxemia; Executive Function (Cognition)
• Intervention / Treatment: Dietary supplement: Short-Chain Fatty Acid Colon-Delivery Capsules; Biological: Lipopolysaccharide (LPS); Dietary supplement: Placebo capsules

Detailed Description

INFLEX is a single-center, randomized, triple-blind, placebo-controlled, parallel-group interventional trial conducted at UZ Leuven / KU Leuven. Its primary aim is to test whether a single acute dose of colon-delivered SCFAs, relative to placebo, attenuates the systemic inflammatory response induced by experimental endotoxemia in healthy adults. Secondary aims are to test whether SCFAs attenuate LPS-induced sickness behaviour and mitigate endotoxemia-induced changes in core executive functioning, and to characterize associations between systemic SCFA uptake, the inflammatory response, and these outcomes.

Eligible participants are randomized to receive SCFA colon-delivery capsule or placebo capsules, taken on the morning of the test day with a standardized low-fiber breakfast. One hour after capsule intake, participants receive a single intravenous bolus of purified Escherichia coli lipopolysaccharide (0.4 ng/kg body weight), which serves as a fixed inflammatory challenge applied to all participants. The timing is designed so that peak circulating SCFA concentrations (approximately 3-4 hours after colonic delivery) coincide with the peak of LPS-induced cytokines and sickness behaviour (approximately 2-3 hours post-injection).

Participants are monitored for a minimum of 6 hours after the LPS challenge. Vital signs, sickness symptoms, and blood and saliva samples are collected hourly to quantify inflammatory markers (including IL-6 and hs-CRP), serum SCFA, cortisol, and ACTH. Core executive-function tasks are administered before and after the LPS challenge and at a 24-hour follow-up visit. A fecal sample collected at home prior to the test day is used for gut microbiota profiling. To standardize endogenous SCFA production, participants follow a low-fiber diet for three days before the test day and avoid caffeine, alcohol, and strenuous exercise beforehand. The night prior to the test day, participants are instructed to consume a standard dinner that is provided to them.

The study comprises three visits: a screening visit (~1 hour), the laboratory test day (~8 hours), and a follow-up visit (~30 minutes) 24 hours after the LPS challenge.

• Study Type: Interventional
• Enrollment (Estimated): 56
• Phase: Not Applicable

Contacts and Locations

• Study Contact: Name: Franco Ruiz, M.D.; Phone Number: +32456716114; Email: franco.ruiz@kuleuven.be
• Study Contact Backup: Name: Dina Satriawan, M.D., Mnsci(Adv); Phone Number: +32 16 71 03 66; Email: dina.satriawan@kuleuven.be

Study Locations

• Belgium
  • Leuven, Belgium, 3000
    • Recruiting
    • University Hospital Leuven (UZ Leuven)
    • Contact: Franco Ruiz, M.D.; Phone Number: +3256716114; Email: franco.ruiz@kuleuven.be
    • Contact: Dina Satriawan, M.D.; Phone Number: +32 16 71 03 66; Email: dina.satriawan@kuleuven.be

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult
• Accepts Healthy Volunteers: Yes

Description

Inclusion Criteria:

• Voluntary written informed consent obtained prior to any study procedure
• Healthy, with no gastrointestinal or psychological complaints
• Age 18-45 years
• BMI 18.5-27 kg/m2
• Proficiency in English and/or Dutch

Exclusion Criteria:

• History of previous or current neurological disorder (e.g., epilepsy, multiple sclerosis, migraine with aura)
• History of previous or current psychiatric disorder (e.g., depression, anxiety disorders, bipolar disorder, schizophrenia)
• History of previous or current gastrointestinal disorder (e.g., Crohn's disease, ulcerative colitis, irritable bowel syndrome)
• History of previous or current endocrine disorder (e.g., diabetes, thyroid disorders, polycystic ovary syndrome)
• History of immune system disorder, including inflammatory, autoimmune, or severe allergic conditions (e.g., rheumatoid arthritis, lupus, celiac disease, or severe allergies such as anaphylaxis)
• History of neuropsychiatric disorder (e.g., ADHD, autism spectrum disorder, learning disorder, Tourette's syndrome)
• History of major head trauma (e.g., concussion with loss of consciousness or long-term symptoms)
• History of severe infection (e.g., sepsis, pneumonia requiring hospitalization, meningitis, endocarditis, or other systemic infection requiring hospitalization, intravenous antibiotics, or intensive care)
• One or more diagnoses based on the Mini International Neuropsychiatric Interview (MINI)
• One or more diagnoses based on ROME-IV criteria for gastrointestinal disorders
• Any disorder that, in the investigator's opinion, might jeopardise the participant's safety or compliance with the protocol
• Any prior or concomitant treatment that might jeopardise the participant's safety or compromise the integrity of the study
• Participation in an interventional trial with an investigational medicinal product (IMP) or device
• Current or recent (past month) prescribed medication use, with particular attention to cardiovascular drugs, steroids, NSAIDs, and centrally-acting drugs (excluding isolated over-the-counter use such as ibuprofen or paracetamol, and hormonal contraceptives in women)
• Use of psychotropic medication within the past year (e.g., antidepressants, anxiolytics, antipsychotics)
• Use of antibiotics within three months preceding the study
• Use of pre- or probiotics within one month preceding the study
• Current or recent (past month) infection (e.g., common cold, influenza, COVID-19)
• Recent (past month) vaccination
• Pregnant or lactating women
• Previous or current substance or alcohol dependence or abuse (>2 units/day or >14 units/week)
• Smoking
• Night-shift work
• Adherence to special diets (e.g., vegan, vegetarian, weight-loss, lactose-free, gluten-free)
• Previous experience with or knowledge of any of the cognitive tasks used in the study (questionnaires excluded)
• C-reactive protein higher than 0.5mg/dL on the day of the injection.
• Colour vision deficiency or colour-blindness

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Basic Science
• Allocation: Randomized
• Interventional Model: Parallel Assignment
• Masking: Quadruple

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: SCFA Colon-Delivery Capsules; A single acute oral dose of colon-delivery capsules delivering a short-chain fatty acid mixture (approximately 186 mmol total SCFA: 111 mmol acetate, 43 mmol propionate, 32 mmol butyrate; molar ratio approximately 60:23:17), taken on the morning of the test day with a standardized low-fiber breakfast.	Dietary supplement: Short-Chain Fatty Acid Colon-Delivery Capsules; Single acute oral dose of pH-dependent colon-delivery capsules delivering a short-chain fatty acid mixture to the colon. The mixture comprises sodium acetate,sodium butyrate, and sodium propionate (54.4%, 21.2%, and 24.4% by weight), providing approximately 186 mmol total SCFA per dose (111 mmol acetate, 32 mmol butyrate, 43 mmol propionate). Capsules are sub-coated with hydroxypropylcellulose and coated with a pH-dependent layer (Eudragit FS 30 D with PlasACRYL T20). Taken on the morning of the test day with a standardized low-fiber breakfast.; Biological: Lipopolysaccharide (LPS); Single intravenous bolus of purified Escherichia coli-derived lipopolysaccharide (0.4 ng/kg body weight), administered one hour after capsule intake to all participants as a standardized experimental inflammatory challenge common to both study arms.; Other Names: LPS; Endotoxin
Placebo Comparator: Placebo; Single acute oral dose of placebo capsules containing microcrystalline cellulose and matching excipients, matched in appearance and number to the SCFA capsules.	Biological: Lipopolysaccharide (LPS); Single intravenous bolus of purified Escherichia coli-derived lipopolysaccharide (0.4 ng/kg body weight), administered one hour after capsule intake to all participants as a standardized experimental inflammatory challenge common to both study arms.; Other Names: LPS; Endotoxin; Dietary supplement: Placebo capsules; Single acute oral dose of placebo capsules containing microcrystalline cellulose and matching excipients, an inert non-fermentable substrate. Matched in appearance and number to the SCFA capsules and taken on the morning of the test day with a standardized low-fiber breakfast.; Other Names: Placebo; microcrystalline cellulose

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Serum interleukin-6 (IL-6) response to the LPS challenge	Effect of SCFA versus placebo on the systemic inflammatory response to experimental endotoxemia, indexed by serum IL-6 concentrations measured at serial timepoints across the test day following the intravenous LPS bolus. The primary analysis evaluates the treatment-by-timepoint interaction on the IL-6 time course using a linear mixed model, with treatment (SCFA vs. placebo) as a between-subject factor and timepoint as a within-subject factor.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Serum pro- and anti-inflammatory cytokine response to the LPS challenge	Effect of SCFA versus placebo on serum concentrations of TNF-alpha, IL-1-beta, IFN-gamma, IL-8, IL-4, and IL-10. Analyzed as treatment-by-timepoint interactions using linear mixed models.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection
High-sensitivity C-reactive protein (hs-CRP) response to the LPS challenge	Effect of SCFA versus placebo on serum high-sensitivity C-reactive protein (hs-CRP) concentrations, measured at selected timepoints on the test day and at the 24-hour follow-up visit.	2 hours before LPS injection, and at 2, 6, and 24 hours after LPS injection
LPS-induced sickness behaviour (SicknessQ)	Effect of SCFA versus placebo on subjective sickness behaviour, assessed with the Sickness Questionnaire (SicknessQ), analyzed across timepoints on the test day.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection
Body temperature response to the LPS challenge	Effect of SCFA versus placebo on body temperature (degrees Celsius).	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection
Heart rate response to LPS challenge	Effect of SCFA versus placebo on heart rate (beats per minute) across the monitoring period.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection
Blood pressure response to the LPS challenge	Effect of SCFA versus placebo on systolic and diastolic blood pressure (mmHg).	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection
Salivary cortisol response to the LPS challenge	Effect of SCFA versus placebo on salivary cortisol concentrations (ng/ml).	At 0, 1, 2, 3, 4, and 6 hours after LPS injection
Plasma ACTH	Effect of SCFA versus placebo on plasma ACTH concentrations (ng/l)	At 0, 1, 2, 3, 4, and 6 hours after LPS injection
Working memory performance (3-back task)	Effect of SCFA versus placebo on working memory, assessed with the 3-back task (primary index: d-prime). Assessed at baseline (pre-LPS), post-LPS on the test day, and at the 24-hour follow-up.	2 hours before LPS injection, 2.25 hours after LPS injection, and 24 hours after LPS injection
Response inhibition (Stop Signal Task)	Effect of SCFA versus placebo on response inhibition, assessed with the Stop Signal Task (outcome: stop-signal reaction time, SSRT, in milliseconds).	2 hours before LPS injection, 2.25 hours after LPS injection, and 24 hours after LPS injection
Cognitive flexibility - task switching (Number-Letter task)	Effect of SCFA versus placebo on cognitive flexibility, assessed with the Number-Letter task (outcome: switch cost, in milliseconds).	2 hours before LPS injection, 2.25 hours after LPS injection, and 24 hours after LPS injection
Cognitive inhibition (Stroop task)	Effect of SCFA versus placebo on cognitive inhibition, assessed with the Stroop task (outcome: congruency effect, in milliseconds).	2 hours before LPS injection, 2.25 hours after LPS injection, and 24 hours after LPS injection
Cognitive flexibility - set shifting (Wisconsin Card Sorting Task)	Effect of SCFA versus placebo on cognitive flexibility, assessed with the Wisconsin Card Sorting Task (outcome: number of perseverative errors).	2 hours before LPS injection, 2.25 hours after LPS injection, and 24 hours after LPS injection
Systemic SCFA uptake (serum SCFA concentrations)	Serum concentrations of acetate, propionate, and butyrate, assessed as a measure of systemic SCFA uptake to confirm target engagement of the colon-delivery intervention, and used as a mediator in mechanistic analyses of treatment effects on the inflammatory response.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, and 6 hours after LPS injection

Other Outcome Measures

Outcome Measure	Measure Description	Time Frame
Baseline gut microbiota composition	Baseline fecal gut microbiota composition, characterized by quantitative microbiome profiling from a single fecal sample collected before the test day. Examined in exploratory analyses as a baseline predictor of inter-individual variability in systemic SCFA uptake and treatment response.	Single fecal sample collected within the 3 days prior to the test day
Mood - valence (MDMQ)	Effect of SCFA versus placebo on self-reported mood valence (good-bad dimension), assessed with the Multidimensional Mood Questionnaire (MDMQ/MDBF). Score range 1-40; higher scores indicate a more positive mood.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
Mood - alertness (MDMQ)	Effect of SCFA versus placebo on self-reported alertness (awake-tired dimension), assessed with the Multidimensional Mood Questionnaire (MDMQ/MDBF). Score range 1-40; higher scores indicate greater alertness.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
Mood - calmness (MDMQ)	Effect of SCFA versus placebo on self-reported calmness (calm-restless dimension), assessed with the Multidimensional Mood Questionnaire (MDMQ/MDBF). Score range 1-40; higher scores indicate greater calmness.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
Negative affect (PANAS)	Effect of SCFA versus placebo on self-reported negative affect, assessed with the negative affect subscale of the Positive and Negative Affect Schedule (10 items, 5-point Likert; score range 10-50). Higher scores indicate greater negative affect.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
Positive affect (PANAS)	Effect of SCFA versus placebo on self-reported positive affect, assessed with the positive affect subscale of the Positive and Negative Affect Schedule (10 items, 5-point Likert; score range 10-50). Higher scores indicate greater positive affect.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
State anxiety (STADI)	Effect of SCFA versus placebo on self-reported state anxiety, assessed with the anxiety subscale of the State-Trait Anxiety and Depression Inventory (STADI), state form (4-point Likert). Higher scores indicate greater state anxiety.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
State depression (STADI)	Effect of SCFA versus placebo on self-reported state depression, assessed with the depression subscale of the State-Trait Anxiety and Depression Inventory (STADI), state form (4-point Likert). Higher scores indicate greater state depressive symptoms.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
Sleepiness (SSS)	Effect of SCFA versus placebo on self-reported sleepiness, assessed with the Stanford Sleepiness Scale (single item, range 1-7). Higher scores indicate greater sleepiness.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection
State anxiety (STAI-S)	Effect of SCFA versus placebo on self-reported state anxiety, assessed with the state subscale of the State-Trait Anxiety Inventory (20 items, 4-point Likert; score range 20-80). Higher scores indicate greater state anxiety.	2 hours before LPS injection, and at 0, 1, 2, 3, 4, 5, 6 and 24 hours after LPS injection

Collaborators and Investigators

• Sponsor: Universitaire Ziekenhuizen KU Leuven
• Collaborators: University Hospital, Essen; Flemish institute of biotechnology (VIB)
• Investigators: Principal Investigator: Lukas Van Oudenhove, MD, PhD, Laboratory for Brain-Gut Axis Studies, Translational Research Center for Gastrointestinal Disorders, Department of Chronic Diseases and Metabolism, KU Leuven, 3000 Leuven, Belgium; Principal Investigator: Kristin Verbeke, PhD, Translational Research Center for Gastrointestinal Disorders, Department of Chronic Diseases and Metabolism, KU Leuven, 3000 Leuven, Belgium; Principal Investigator: Boushra Dalile, PhD, Laboratory for Brain-Gut Axis Studies, Translational Research Center for Gastrointestinal Disorders, Department of Chronic Diseases and Metabolism, KU Leuven, 3000 Leuven, Belgium

Study record dates

Study Major Dates

• Study Start (Actual): October 24, 2025
• Primary Completion (Estimated): March 30, 2027
• Study Completion (Estimated): March 30, 2027

Study Registration Dates

• First Submitted: June 30, 2026
• First Submitted That Met QC Criteria: July 14, 2026
• First Posted (Actual): July 20, 2026

Study Record Updates

• Last Update Posted (Actual): July 20, 2026
• Last Update Submitted That Met QC Criteria: July 14, 2026
• Last Verified: June 1, 2026

More Information

Terms related to this study

• Keywords: inflammation; healthy volunteers; executive function; microbiota; gut-brain axis; working memory; systemic inflammation; short-chain fatty acids; endotoxemia; cognitive inhibition; inhibition; butyrate; response inhibition; microbiota-gut-brain axis; acetate; cognitive flexibility; experimental; lps; task switching; scfa; proprionate; lypopolysaccharide; lps challenge; experimental inflammation; experimental endotoxemia; sickness behaviour
• Additional Relevant MeSH Terms: Pathologic Processes; Infections; Sepsis; Systemic Inflammatory Response Syndrome; Bacteremia; Toxemia; Pathological Conditions, Signs and Symptoms; Behavior; Inflammation; Endotoxemia; Inhibition, Psychological; Van der Woude syndrome; Lipids; Biological Factors; Carbohydrates; Polysaccharides; Glycoconjugates; Bacterial Toxins; Toxins, Biological; Antigens; Antigens, Bacterial; Polysaccharides, Bacterial; microcrystalline cellulose; Endotoxins; Lipopolysaccharides
• Other Study ID Numbers: S69844

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: UNDECIDED

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No