Natural Modulators of Immune Function and Mood
Condition: Stress
Study type: Interventional

Summary: 
      The objective of this proposal is to recruit 20 healthy subjects per ingredient arm (5 arms x
      20 subjects; N= 120 total) to participate in a research study investigating the effects of
      dietary supplementation for one-month with different dietary supplement formulations (versus
      placebo; N=20) that may have benefits for supporting respiratory health, psychological
      stress, and quality of life.

      Supplements to be studied (all are commercially available):

        1. Chaga (mushroom extract)

        2. Arava (algae concentrate)

        3. Thymoquin (black cumin seed oil)

        4. Thymoquin + Astaxanthin (antioxidant carotenoid)

        5. Thymoquin + Omega 3 (fish oil)

        6. Placebo (corn starch)

      In order to "stress" volunteers, participants will train for and complete a strenuous
      off-road trail marathon to induce both physical and mental stress.
    

Detailed description: 
      The main hypothesis is that the group(s) supplementing with natural stress modulators will
      demonstrate superior subjective assessments of both physical health (e.g. respiratory
      function, mobility, flexibility) and mental stress (e.g. mood, energy, focus).

      Stress-related factors following intense endurance activity can lead to a range of adverse
      physical manifestations including muscle soreness, physical fatigue, and general malaise.
      Furthermore, stressful events frequently lead to negative emotions including temporary
      reductions in perceived mood and energy with heightened irritability and tension.

      Each of the ingredients under consideration for this study, Chaga, Algae extract, Black Cumin
      Seed Oil, Astaxanthin, and Omega-3s, have a number of laboratory and animal studies
      demonstrating anti-stress benefits including improvements in physical mobility (e.g.
      flexibility), mental energy, and overall well-being.

      Each of these ingredients may be considered as natural "stress-modulators" that can help to
      balance overall well-being and quality of life. Such natural substances represent an emerging
      approach to buffer the detreimnetal effects of physical and mental stressors.

      Mushrooms, particularly Chaga, have a long history of use in traditional medicine for
      supporting general well-being. An abundance of in-vitro evidence suggests that a wide range
      of mushroom bioactives may modify stress responses, with promising preliminary research in
      humans.

      Algae concentrate and in particular Astaxanthin, the natural red carotenoid pigment found in
      microalgae, has been shown to be a broad-spectrum stress modulator, likely due to its actions
      in reducing oxidative and inflammatory stress and thus protecting mitochondria from damage
      and controlling cellular senescence.

      Black cumin seed oil and its primary bioactive compound, thymoquinone, have been used in
      traditional Middle Eastern Medicine for thousands of years for its broad-spectrum benefits
      related to longevity and overall well-being.

      Omega-3 fatty acids are well-described in the scientific literature for their
      anti-inflammatory effects and thus benefits in a wide range of health parameters. Because of
      the close relationship between well-being and inflammatory balance, omega-3 fatty acids may
      help to support quality of life parameters, particularly after stress, which is known to
      disrupt both inflammatory signaling and psychological mood state.

      Key Outcome Measures

        -  Mental and Physical Stress (Vivosmart)

        -  Mood, Energy, Focus, etc = Profile of Mood States (POMS)

        -  Microbiome Assessment (BiomeTracker)

        -  Salivary Cortisol (morning waking sample)

        -  Well-Being Symptoms (Health Log)

        -  Blood Pressure (in 4 arms containing Black Cumin Seed OIl and Placebo)

        -  Blood Omega 3 (in 2 arms containing Omega3 and Placebo)

      Volunteers will perform all study measurements in the privacy of their own home.

      Mental & Physical Stress

        -  Stress (mental and physical) is assessed by heart rate variability (the time interval
           between heart beats)

        -  "Readiness" is assessed as "Body Battery Energy" - a combination assessment of stress,
           heart rate variability, sleep quality, and physical activity.

        -  Sleep Quality is assessed by tracking total, light, deep, and REM stages of sleep.

        -  Each of these parameters is automatically uploaded by the subject to a secure
           password-protected online database (Garmin Connect) where the investigators can access
           to collect and collate data.

      Health Log The daily health log contains questions related to overall health status and
      general well-being (e.g. stress, energy, flexibility/mobility, etc).

      Mood Assessment (POMS; Profile of Mood States) Longer term changes in mood/focus/energy will
      be assessed using the research-validated Profile of Mood States (POMS) questionnaire to
      measure 6 primary psychological factors (tension, depression, anger, fatigue, vigor, or
      confusion) plus the combined global mood state as an indication of subjective well-being. The
      POMS methodology has been used in ~3,000 studies, and its validity is well established. The
      POMS profile uses 65 adjective-based intensity scales scored on a 0-4 hedonic scale (e.g.
      "not at all" to "extremely"). The 65 adjective responses are categorized into the 6 mood
      factors (tension, depression, anger, fatigue, vigor, or confusion), tabulated, scored, and
      analyzed. The output of the POMS questionnaire is an assessment of the positive and negative
      moods of each subject at baseline and post-supplementation.

      Microbiome Assessment (BiomeTracker) Microbiome analysis of fecal samples will be carried out
      using the complete BiomeTracker system (Wasatch Scientific, Murray, UT). Briefly, fecal
      samples will obtained by nylon swab and placed into preservative binding buffer to lock the
      composition of bacteria in place. DNA is then purified using DNA columns and ~20ng of DNA
      from each sample is added to the reaction mixtures. Samples are processed on an ABI 7500 Fast
      (Applied Biosystems) instrument in duplicate. A "microbiome composite score" is generated as
      an overall average of many different aspects of microbiome balance, including
      Bifidobacterium, Lactobacillus, Akkermansia, Firmicutes/Bacteroidetes (F/B) ratio, and
      others.

      Volunteers will be provided with take-home kit to obtain fecal samples in the privacy of
      their home for analysis. The kit will include detailed instructions and postage paid
      packaging to be returned directly to Wasatch Scientific Laboratories. Each kit will be
      numerically coded so that samples are blinded to the lab.

      Salivary Cortisol Cortisol is the major glucocorticosteroid hormone produced in the adrenal
      cortex. Cortisol is actively involved in the regulation of calcium absorption, blood pressure
      maintenance, anti-inflammatory function, gluconeogenesis, gastric acid, pepsin secretion, and
      immune function. Cortisol production has a circadian rhythm. Levels peak in the early morning
      and drop to the lowest concentration at night. Levels rise independently of circadian rhythm
      in response to stress. In the blood only 1 to 15% of cortisol is in its unbound or
      biologically active form. The remaining cortisol is bound to serum proteins. Unbound serum
      cortisol enters the saliva via intracellular mechanisms, and in saliva the majority of
      cortisol remains unbound to protein. Salivary cortisol levels are unaffected by salivary flow
      rate or salivary enzymes. Studies consistently report high correlations between serum and
      saliva cortisol, indicating that salivary cortisol levels reliably estimate serum cortisol
      levels.

      Volunteers will be provided with take-home kit to obtain saliva samples in the privacy of
      their home for cortisol analysis. The kit will include detailed instructions and will be
      returned to CRC for shipment to Wasatch Scientific Laboratories. Each kit will be numerically
      coded so that samples are blinded to the lab.

      The CRC will confirm with volunteers and Wasatch Scientific that these samples have been
      collected and received by the lab for analysis.

      Blood Pressure Blood pressure will be measured with Omron Series 3 Upper Arm Blood Pressure
      Monitor, which will be provided to 10 randomly selected participants in each of the three
      study arms containing Thymoquin as well as 10 randomly selected participants in the placebo
      group.

      Blood Omega-3 Index Blood levels of omega-3 fatty acids will be assessed in the group taking
      Omega-3 fish oil supplements, using a self-collected at-home fingerstick blood sample
      (OmegaQuant). Subjects will receive all supplies and instructions (see Instruction card
      below) to apply a drop of blood to the collection card and mail sample to the laboratory in a
      pre-paid envelope.
    

Intervention name(s): Chaga Mushroom; Algae Extract; Black Cumin Seed Oil; Black Cumin Seed Oil + Astaxanthin; Black Cumin Seed Oil + Omega 3; Placebo
Intervention type(s): Dietary Supplement; Dietary Supplement; Dietary Supplement; Dietary Supplement; Dietary Supplement; Dietary Supplement
Intervention description(s): Effect of chaga on immune/mood support; Effect of algae extract on immune/mood support; Effect of black cumin seed oil on immune/mood support; Effect of Black Cumin Seed Oil + Astaxanthin blend on immune/mood support; Effect of Black Cumin Seed Oil + Omega3 blend on immune/mood support; Placebo control
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: treatment (5 dietary supplement blends) versus placebo (control)
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Prevention