Developing tools to detect when our bodies are more resistant towards protein synthesis is valuable for identification of when someone may be at risk of losing body or muscle mass such as with aging or certain diseases. The current study aims to refine our previous breath test method to be more effective at measuring changes in how the body processes protein in different situations, such as resting, reducing physical activity, and doing resistance exercise. We hypothesize that using a lower amount of dietary amino acids in our breath test will be effective at detecting lower amounts of amino acids used after exercise, and a greater amount with step reduction compared to normal activity levels

Maintaining high-quality and abundant lean body mass (LBM) is crucial for growth, health, and performance across all ages, sexes, and activity levels. Body protein, including skeletal muscle, undergoes constant turnover, breaking down old and damaged proteins and using dietary amino acids (AA) to synthesize new proteins, especially after resistance exercise. Unused AA are oxidized for energy and excreted as carbon dioxide (CO2). Studying the proportion of AA used for protein synthesis versus energy production provides insights into acute growth of LBM after a meal in different physiological states (e.g., at rest or exercise). Stable isotope tracers, commonly administered intravenously, are used in protein metabolism research to examine the effects of nutrition and exercise on protein turnover. However, this method may not be feasible for vulnerable populations.

While exercise has been shown to enhance anabolic sensitivity (i.e., greater utilization of dietary AA for protein synthesis), step-reduction leads to fed-state anabolic resistance (I.e., reduced utilization of dietary AA for protein synthesis). Indeed, reduced habitual activity, whether mild or severe, leads to fed-state anabolic resistance, reducing the muscle protein synthesis (MPS) response to amino acids. For instance, one week of reduced daily steps (~1,192 steps/day) decreased MPS rates by approximately 27% in young males who habitually reach ~10,000 steps/day, i.e., a ~75% reduction from habitual.

Therefore, developing metabolic tools to detect anabolic resistance before muscle mass loss occurs would be valuable for both treatment and prevention of age-related muscle loss. Recently, our laboratory demonstrated the effectiveness of a non-invasive stable isotope "breath test" to detect increased anabolic sensitivity in males after resistance exercise. This study, in addition to ongoing metabolic trials in our lab, utilized a protein dose of 0.25g/kg which has been shown to maximize the rate of myofibrillar protein synthesis and to support whole-body protein synthesis. However, this dose may not adequately distinguish between more subtle changes in anabolic sensitivity. Further, a lower protein dose may reduce the duration for which breath samples would need to be collected, which would minimize participant burden and time commitment going forward.

Therefore, the present project will use previously established 'breath test' methodology but with a lower protein dose to assess the following objectives:

1. To investigate whether the 'optimized' non-invasive breath test can detect changes in anabolic sensitivity (i.e., leucine oxidation) across varying physiological states (I.e., rest, step-reduction, whole-body resistance exercise) after feeding, in young healthy adults.
2. To assess the reproducibility, and day-to-day variability of our breath test during habitual activity in both an at-home and a controlled laboratory setting

Inclusion Criteria:

• Healthy young (age: 18-35 years)
• BMI between normal to overweight (18.5-29.9 kg/m2)
• if oral contraceptive (OC) user, must be on monophasic OCs for at least 3 months prior to study
• if non-OC user, then must have regular menstrual cycles (length: 25-35 days) for at least 3 months prior to study and at least 6 months off of OCs

Exclusion Criteria:

• Chronic disease diagnosis (cardiovascular, thyroid, diabetes)
• Current or recent remission of cancer
• Regular use of NSAID (except low-dose aspirin), anticoagulants
• Use of prescription drugs that would impact muscle protein synthesis (e.g., Statins, Lithium, ADHD medication, etc..)
• Insertion of intrauterine device (IUD) - exception: copper
• Smoking
• Use of illicit drugs (growth hormones, testosterone)

Participants will be randomized to either begin their participation with the "at-home" or "in-person" phase of the study. There is no need to conceal the randomization as this study is not blinded - this does not affect our outcome as we are looking at physiological changes with feeding.