Inulin and Nitrate's Effects on Vascular Health and Gut Microbiome

Not Applicable

Completed

• Conditions: Gut Microbiota Dysbiosis and Nutrition; High Blood Pressure; Endothelial Dysfunction
• Interventions: Dietary Supplement: Inulin; Dietary Supplement: Potassium nitrate and inulin

• Registration Number: NCT06502535
• Lead Sponsor: University of Exeter

Brief Summary

Previous research has revealed that adults with pre-hypertensive and hypertensive blood pressure levels typically exhibit impaired vasodilation, which is a crucial aspect of blood vessel function, due to decreased nitric oxide production. Flow-mediated dilation (FMD) is a widely used technique for assessing vasodilation, which gauges the capacity of blood vessels to widen in response to increased blood flow. Nitrate supplements have been shown to enhance blood vessel function both in the short and long term by elevating nitric oxide (NO) production. Similarly, acetate, a by-product of fibre digestion, has demonstrated anti-inflammatory effects in the gut and may also increase NO production. However, further investigation is needed to determine whether these two supplements work synergistically to improve FMD responses by augmenting NO production in the human body. This study will involve administering inulin plus nitrate supplements to adults aged 45-74 with untreated high blood pressure (120-139/80-89 mmHg and 140/90 mmHg or higher) for a period of 4 weeks, followed by a subsequent 4-week period where only inulin supplements are consumed. The supplements will be provided in the form of powder, to be mixed with water, and will be given in a cross-over and counterbalanced order. Various measurements, including brachial artery function, faecal samples, blood pressure, plasma nitrate and nitrite levels, plasma short-chain fatty acid levels, red blood cell and whole blood S-Nitrosothiols, and salivary nitrate and nitrite levels, will be taken at the baseline and after each supplementation period. Additionally, acute supplementation measurements will be collected on two separate occasions before the start of the longer-term supplement regimen.

Detailed Description

Endothelial dysfunction contributes to the increased cardiovascular risk associated with high blood pressure levels (e.g., 120-139/80-89 mmHg and 140/90 mmHg or higher), and it is considered as an early event in the development of atherosclerotic vascular disease. Mechanistically, endothelial dysfunction is characterised by a reduced bioavailability of the endothelium-derived molecule nitric oxide (NO), which is reflected by impaired brachial artery flow-mediated dilation (FMD). However, endothelial dysfunction is largely preventable by its early detection, thus approaches that might restore the loss of NO in the body could have obvious therapeutic potential.

Consumption of the prebiotic fibre inulin has been shown to increase the fermentation of bacterial-derived metabolites such as acetate by our gut microbiome. While increased acetate has local benefits in the gut (i.e., anti-inflammatory, lower luminal pH), it also has the potential to improve endothelial health. Research has shown that acetate can increase NO bioavailability via the activation of the NO synthase pathway by increasing phosphorylation of endothelial NO synthase (eNOS), a key enzyme in the maintenance of endothelial homeostasis, following a 15-day supplementation with inulin. Similarly, NO is produced following the consumption of nitrate, with individual responses being dependent on the basal eNOS activity. Therefore, individuals with compromised eNOS activity (e.g., pre- and hypertensive) might show an augmented response to nitrate supplementation. Since both nitrate and inulin supplementations can increase NO production, it is possible that their combined ingestion might enhance endothelial function to a greater extent. When supplemented in isolation, nitrate proved an effective dietary strategy to ameliorate endothelial dysfunction in older and clinical populations. For example, a 4-week chronic dietary nitrate supplementation (400 mg nitrate/day) was associated with an increased peak brachial artery FMD of 1.0% (95% CI, 0.3-1.5; p \< 0.001), which was not evident in the placebo group. Similarly, following a 4-week dietary nitrate supplementation (400 mg nitrate/day), brachial artery FMD (%) values increased significantly in the nitrate group compared to the placebo group. Unlike nitrate, the effects of inulin on brachial artery FMD in humans are yet to be established. Preliminary findings demonstrated an enhancement of endothelial function after inulin supplementation in mice. These findings were further supported by another study which demonstrated a brachial artery FMD increase of \~1% following a 3-month prebiotic supplementation with 12 g fructo-oligosaccharides in clinical patients with less severe endothelial dysfunction.

To date, no studies that the investigators are aware of have explored the role of nitrate and inulin in combination; however, based on the above, these two nutrients have the potential to work synergistically. Therefore, the primary aim of this study is to assess whether adding nitrate to inulin during a 4-week chronic supplementation might lead to an increase in brachial artery FMD in middle-aged and youngest elderly adults (45-74 years old) with elevated blood pressure and compare this response to inulin supplemented in isolation. The secondary aims of this study include brachial artery FMD % changes at 4 h post-consumption of a single dose of 15 g inulin and 1300 mg potassium nitrate combined and 15 g inulin supplemented in isolation. This study will also explore whether changes in brachial artery FMD % following the two supplementations are mediated by potential changes in the gut microbiome composition and diversity that may occur following prebiotics supplementation. Further secondary outcomes of the study will investigate differences in plasma nitrate, nitrite, and short-chain fatty acids (SCFAs) following both types of supplementation. Additionally, the study will examine changes in the levels of S-nitrosothiols (RSNO) in red blood cells and whole blood. The study will also compare changes in resting blood pressure between the combined inulin and nitrate supplementation and the inulin-only supplementation.

Study Timeline

• Study Start: 2023-05-09
• Study First Submitted: 2024-05-14
• Study First Submitted QC: 2024-07-08
• Study First Posted: 2024-07-16
• Primary Completion: 2024-07-31
• Study Completion: 2024-07-31
• Status Verified: 2025-01
• Last Update Submitted: 2025-01-07
• Last Update Posted: 2025-01-09

Recruitment & Eligibility

• Status: COMPLETED
• Sex: All
• Target Recruitment: 22

Inclusion Criteria

• Able to provide informed consent
• Adults aged 45-74
• Adults with untreated elevated blood pressure as SBP 120 to 129 mm Hg and DBP < 80 mm Hg or untreated stage 1 hypertension (i.e., pre-hypertension) as SBP 130 to 139 mm Hg or DBP 80 to 89 mm Hg or untreated stage 2 hypertension as SBP equal to or higher than 140 mm Hg or DBP equal to or higher than 90 mm Hg (ACC/AHA and ESC/ESH guidelines and Blood Pressure UK)
• Weight-stable in the 3 months prior to enrolment (self-report)

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Exclusion Criteria

• Body mass index > 40 kg/m^2 at screening;
• History of symptomatic coronary artery disease, stroke, or other known atherosclerotic disease;
• History of gastrointestinal disease (e.g., irritable bowel syndrome, inflammatory bowel disease, coeliac disease, Chron's disease) or former abdominal surgery, weight loss surgery e.g., gastric by-pass (except for appendectomy);
• History of chronic viral hepatitis (including presence of hepatitis B surface antigen or hepatitis C antibody), or other chronic hepatic disorders;
• History of malignancy within the past 5 years, other than non-melanoma skin cancer;
• History of diabetes and/or any endocrine disorder;
• Current use of nitrate medications, including glyceryl trinitrate (GTN), isosorbide dinitrate and isosorbide mononitrate and blood pressure medications;
• Consumption of a low calorie or other special diet during the last month prior to the study, or if following a slimming or medically prescribed diet, or a vegan or macrobiotic lifestyle;
• Use of antibiotics during the three months prior to screening;
• Consumption of pre- or probiotics during the last month prior to screening and during the study, unless discontinued 3 weeks before the start of the trial. This is referred to the use of supplements only and NOT foods containing probiotics and/or prebiotics foods;
• Consumption of dietary supplements such as vitamins C and E during the last month prior to the screening and during the trial, unless discontinued 3 weeks before the start of the trial;
• Consumption of fish oil 6 weeks prior to the screening and during the trial, unless discontinued 6 weeks before the start of the trial;
• Allergies or intolerance against the substances used in the study and/or food intolerances associated with gastrointestinal upset;
• Any apparent dependence on or abuse of alcohol, tobacco, and cannabis products;
• Pregnancy, breast-feeding, or plans to become pregnant during the duration of the study;
• Current use of nitrate supplements in the form of beetroot juice, beetroot crystals, potassium nitrate and sodium nitrate, including L-arginine or L-citrulline supplements.
• Self-reported use of antimicrobial mouthwash or tongue scrapes, unless discontinued during the trial.

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Study & Design

• Study Type: INTERVENTIONAL
• Study Design: CROSSOVER

Arm && Interventions

Group	Intervention	Description
Inulin (15 g)	Inulin	Participants will receive 15 g inulin in powder form once daily for 4 weeks.
Inulin (15 g) and potassium nitrate (1300 mg)	Potassium nitrate and inulin	Participants will receive 15 g inulin in powder form once daily for 4 weeks and 1300 mg potassium nitrate in powder form once daily for 4 weeks.

Primary Outcome Measures

Name	Time	Method
Plasma nitrite and nitrate levels	Day 0, Month 3	Differences in the concentrations of plasma nitrate and nitrite between the 'inulin plus nitrate' and 'inulin' supplementations as assessed by chemiluminescence, measured from baseline to the 4-week chronic supplementation.
Plasma Short-Chain Fatty Acid (SCFA) levels	Day 0, Month 3	Differences in the concentrations of plasma acetate, butyrate, and propionate between the 'inulin plus nitrate' and 'inulin' supplementations as assessed by liquid chromatography coupled with tandem mass spectrometry, measured from baseline to the 4-week chronic supplementation.
Gut microbial composition using 16S sequencing	Day 0, Month 3	Changes in microbial composition, including the relative abundances of Operational Taxonomic Units (OTUs), will be evaluated using the 16S rRNA V3-V4 region. These changes will be observed from baseline to the end of the 4-week intervention following supplementation with inulin plus nitrate and inulin alone.
Flow-mediated dilation (FMD) with ultrasound to detect a change in brachial artery FMD	Day 0, Month 3	Absolute percentage change in FMD response between the 'inulin plus nitrate' and 'inulin' supplementations, measured from baseline to the 4-week chronic supplementation.

Secondary Outcome Measures

Name	Time	Method
Flow-mediated dilation (FMD) with ultrasound to detect a change in brachial artery FMD	Baseline, 4 hour post supplement consumption	Absolute percentage change in FMD response between the 'inulin plus nitrate' and 'inulin' supplementations, measured from baseline to 4 h post-consumption of the supplements.
Body weight	Day 0, Month 3	Changes in body weight in Kg following supplementation with 'inulin plus nitrate' and 'inulin' will be measured with the use of electronic scales.
Plasma nitrite and nitrate levels	Baseline, 4 hour post supplement consumption	Differences in the concentrations of plasma nitrate and nitrite between the 'inulin plus nitrate' and 'inulin' supplementations as assessed by chemiluminescence, measured from baseline to 4 h post-consumption of the supplements.
Rate of habitual fibre intake before and during the intervention	Pre intervention, at baseline	Participants will be categorised based on their habitual dietary fibre intake to determine if the effects of the supplemented inulin are independent of their overall fibre consumption.; The questionnaire includes 5 questions addressing the intake from the five main food groups contributing to fibre intake: fruit, vegetables, bread and cereals, nuts and seeds, and legumes. The intake of these food groups will be evaluated using the following scale: Never Less than 1 serving per month 1-3 servings per month; 1 serving per week 2-4 servings per week 5-6 servings per week; 1. serving per day; 2. servings per day; 3. servings per day; 4. servings per day; 5. servings per day; 6. or more servings per day
Dietary intake	Day 0, Month 3	4-day recall questionnaire filled out during 4 different occasions to evaluate dietary intake, including macronutrients and caloric intake.
Body fat percentage	Day 0, Month 3	Changes in body fat percentage following supplementation with 'inulin plus nitrate' and 'inulin' will be assessed using bioimpedance analysis.
Waist circumference	Day 0, Month 3	The changes in waist circumference, measured halfway between the lower ribs and the iliac crest, following supplementation with 'inulin plus nitrate' and 'inulin', will be used to calculate the waist-to-hip ratio in conjunction with hip measurement, from baseline to the 4-week intervention.
Plasma Short-Chain Fatty Acid (SCFA) levels	Baseline, 4 hour post supplement consumption	Differences in the concentrations of plasma acetate, butyrate, and propionate between the 'inulin plus nitrate' and 'inulin' supplementations as assessed by liquid chromatography coupled with tandem mass spectrometry, measured from baseline to 4 h post-consumption of the supplements.
S-nitrosothiols (RSNO) in red blood cells and whole blood	Day 0, Month 3	Differences in the concentrations of RSNO in red blood cells and whole blood following the 'inulin plus nitrate' and 'inulin' supplementations as assessed from chemiluminescence, measured from baseline to the 4-week intervention.
Resting blood pressure	Day 0, Month 3	Differences in resting blood pressure following the 'inulin plus nitrate' and the 'inulin' supplementations as assessed by an automatic sphygmomanometer, measured from baseline to the 4-week intervention.
Tolerability of inulin defined by gastrointestinal wellbeing	Day 0, Month 3	Tolerability of a 4-week intervention with inulin with or without nitrate, assessed by completion of gastrointestinal wellbeing questionnaires, also known as the Gastrointestinal Symptom Rating Scale (GSRS). GSRS score will be calculated as per Revicki 1997: Response scale; 1. No discomfort at all; 2. Slight discomfort; 3. Mild discomfort; 4. Moderate discomfort; 5. Moderately severe discomfort; 6. Severe discomfort; 7. Very severe discomfort; The 14 items combine into five symptom clusters: Reflux, Abdominal pain, Indigestion, Diarrhoea and Constipation.
Hip circumference	Day 0, Month 3	The changes in hip circumference, measured at the largest circumference around the buttocks, following supplementation with 'inulin plus nitrate' and 'inulin,' will be used to calculate the waist-to-hip ratio in conjunction with waist measurement, from baseline to the 4-week intervention.

Trial Locations

Locations (1)

University of Exeter; 🇬🇧 Exeter, Devon, United Kingdom