Beet Supplementation, Exercise, Inflammation
Condition: Inflammatory Response; Metabolic Disturbance; Immune Suppression
MeSH terms: Inflammation
Study type: Interventional

Summary: 
      This study will determine if ingesting a beet-based supplement with nitrates for 2 weeks
      moderates exercise-induced inflammation.
    

Detailed description: 
      RESEARCH DESIGN: Subjects will come to the lab for orientation/baseline testing, pre-and
      post-supplementation blood sample collections (2 weeks supplementation with POSSIBLE Beet
      Pre-Workout supplement (Standard Process, Kannapolis, NC) or placebo), two 2.25 h cycling
      sessions, and additional lab visits to provide 1-day 7:00 am recovery blood samples) (thus 7
      total lab visits). The total amount of time subjects will be asked to volunteer for this
      study is about 20 hours at the Human Performance Laboratory (over a 6-week period).

      A. Orientation/Baseline Testing (Visit #1) After voluntarily signed IRB-approved consent
      forms, study participants will be tested for maximal aerobic capacity (VO2max) during a
      graded, cycling test with continuous metabolic monitoring with the Cosmed CPET metabolic
      device (Cosmed, Rome, Italy). Body composition will be measured with the seca BIA and Bod Pod
      body composition analyzer (Life Measurement, Concord, CA). Demographic and training histories
      will be acquired with questionnaires.

      B. Pre-Supplementation Lab Visit #2 and 2-Week Supplementation Protocol: Two weeks prior to
      the first cycling session, subjects will report to the Human Performance Laboratory at
      approximately 7:00-8:00 am. A blood sample will be collected in an overnight fasted state to
      coincide with the same time of the day for the post-supplementation blood draw. A 2-week
      supply of POSSIBLE Beet Pre-Workout supplement or placebo supplement will be given to the
      participants. Subjects will mix the supplement powder in one cup of cold water and consume
      one dose in the early morning and a second dose mid-day.

      POSSIBLE Beet Pre-Workout supplement (subjects will consume 2 scoops per day, 28 grams total,
      with one dose in the early morning, and the second dose mid-day). Note: the placebo will
      include excipients only, with none of the ingredients listed below.

      Ingredients (per 14 g scoop dose) Beets: Beets in combination with fermented beets to provide
      nitrates (106 mg of nitrates per serving).

      Caffeine (100 mg) from Green Tea Extract. Camu Camu to provide vitamin C (22 mg or 25% of the
      Daily Value). Camu camu is a sour berry native to the Amazon rainforest that has a high
      vitamin C content.

      Quinoa Sprouts: Source of B-vitamins and will provide 20% of the Daily Value for thiamin,
      riboflavin, niacin, and vitamin B6. Quinoa is a plant with edible seeds that are rich in B
      vitamins.

      Mushroom Blend of three common mushrooms (1.5 g evenly divided from Hericium erinaceus or
      lion's mane mushroom, Cordyceps sinensis or caterpillar fungus, and Inonotus obliquus or
      chaga). These three mushrooms may support exercise performance, but more scientific evidence
      is needed.

      C. 2.25 h Cycling Session (Lab Visit #3): During the 3-day period prior to the 2.25-h cycling
      session, subjects will taper exercise training and ingest a moderate-carbohydrate diet using
      a food list restricting high fat foods, visible fats, caffeine, and polyphenols (1,2). Study
      participants will report to the Human Performance Lab in an overnight fasted state and
      provide a blood sample, ingest the supplement (POSSIBLE Beet Pre-Workout supplement or
      placebo), and then cycle 2.25 h at high intensity (70% VO2max) while ingesting water alone (3
      ml/kg every 15 minutes). Blood samples will be collected at 0 h, 1.5 h, 3.0 h, and 24 h
      post-exercise.

      Testing protocol during the lab sessions with the 2.25-h cycling session:

        1. 7:00 am: Provide blood sample, delayed onset of muscle soreness (DOMS) rating (1-10
           scale), and profile of mood states ratings (POMS).

        2. 7:10 am: Ingest supplement with 1 cup water.

        3. 7:30 am: Start the 2.25 h cycling session. Subjects will cycle on trainers at 70% VO2max
           (~race pace) for 2.25 hours. Oxygen consumption, carbon dioxide production, respiratory
           exchange ratio, and ventilation will be measured using the Cosmed Quark CPET metabolic
           cart every 30 minutes. Subjects will consume 3 ml/kg water every 15 min. No other
           beverage or food containing energy or nutrients will be allowed during the 2.25-h
           exercise sessions.

        4. ~10:00 am to 1:00 pm: Blood samples will be taken via venipuncture immediately after
           completing the cycling session, and then 1.5-h and 3.0-h post-exercise. Subjects will be
           allowed to shower and change clothes. The DOMS and POMS questionnaires will be
           administered each time blood samples are collected. Subjects will ingest no food or
           beverage other than water (7 ml/kg during the 1.5 h post-exercise period). After the 1.5
           h post-exercise blood draw, subjects will ingest a Boost beverage (8 kcal/kg body
           weight). Another blood sample will be collected 3-h post-exercise. Afterwards, subjects
           will be allowed to leave the lab with instructions to adhere to the food list
           requirements, and to keep exercise training moderate.

        5. 7:00 am, next morning: 24-h blood sample and DOMS and POMS ratings (Lab visit #4).

      D. Washout/Crossover/Repeat Subjects will engage in a 2-week washout period without the
      supplements, crossover to the opposite treatment arm, and then repeat all procedures (Lab
      visits #5,6,7).

      STATISTICAL ANALYSIS AND POWER CALCULATION The study participant number (N = 20) will provide
      84% power to detect a difference with an effect size 0.7 at alpha 0.05 using 2-sided paired
      t-tests (5). Data will be checked for normality of the residuals using Q-Q plots. All data
      will be analyzed using the generalized linear model (GLM), and a 2 (condition) x 6 (time)
      repeated-measures ANOVA, within-participants design (IBM SPSS Statistics for Windows, Version
      24.0, IBM Corp, Armonk, NY, USA). Changes over time within conditions will be contrasted
      between trials using paired t-tests, with the alpha level set at p,0.01. Principle component
      analysis (PCA) and heatmap analysis of the oxylipin data will be performed with the R
      programming language (https://www.r-project.org/, version 3.3.1).
    

Intervention name(s): Beet; Placebo
Intervention type(s): Dietary Supplement; Dietary Supplement
Intervention description(s): Beet-based powder with other ingredients; Placebo powder.
Allocation: Randomized
Intervention model: Crossover Assignment
Intervention model description: Randomized treatment, double blinded procedures, crossover
Masking: Double (Participant, Investigator)
Primary purpose: Basic Science
Keywords: Beets; exercise; inflammation; immune function; oxylipins; proteomics