Evaluation of Molecular Hydrogen Supplementation to Enhance the Efficacy of Weight Loss Retreat in Overweight and Obese Adolescents

Body mass index is calculated as the ratio of body mass (kg) to the square of body height (m2). Body mass is measured using a digital scale to the nearest 0.1 kg and body height is measured using a stadiometer to the nearest 1 cm.

Six-minute walk test is measured by recording the total distance the participant is able to walk on a flat surface within six minutes. The participant is instructed to walk at his/her own pace in order to walk as far as possible in the given time.

Body fat is measured using bioelectrical impedance analysis (InBody 270, InBody, Seoul, South Korea).

30-second sit-to-stand test is measured by counting the number of repetitions in which the participant rises from sitting in a chair and sits back down within 30 seconds.

Maximal isometric grip force is measured using a hand dynamometer. The test is performed seated, with the elbow flexed at 90 degrees, and wrist in a neutral position.

Maximal wall sit is measured by recording the maximum duration for which the participant can maintain a static isometric squat position against a wall.

The level of resting blood lactate is measured using the Lactate Scout+ (EKF Diagnostics, Cardiff, United Kingdom). Capillary blood is collected in a fasting state from the fingertip using a sterile disposable lancet. Immediately after collection, the blood sample is applied to the test strip of the lactate meter and analyzed in real time.

Homeostatic model assessment of insulin resistance is calculated based on fasting glucose and fasting insulin levels measured from blood samples obtained via venipuncture by a healthcare specialist. Plasma glucose is separated by centrifugation and analyzed using an enzymatic colorimetric method. Serum insulin is separated by centrifugation and analyzed using a chemiluminescent immunoassay.

The level of total cholesterol is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation and analyzed using an enzymatic colorimetric method.

The level of low-density lipoprotein is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation and analyzed using an enzymatic colorimetric method.

The level of high-density lipoprotein is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation and analyzed using an enzymatic colorimetric method.

The level of triglycerides is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation and analyzed using an enzymatic colorimetric method.

The level of cortisol is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The serum is separated by centrifugation and analyzed using a chemiluminescent immunoassay.

The level of fibroblast growth factor 21 is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until ELISA analysis.

The level of total antioxidant capacity is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until analysis using the TROLOX equivalent antioxidant capacity assay.

The level of malondialdehydes is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until ELISA analysis.

The level of protein carbonyls is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until ELISA analysis.

The level of interleukin-6 is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until ELISA analysis.

The level of tumor necrosis factor alpha is measured from a fasting blood sample obtained via venipuncture by a healthcare specialist. The plasma is separated by centrifugation, divided into cryotubes, and stored at -80°C until ELISA analysis.

Waist circumference is be measured at the narrowest part of the torso, above the iliac crest and below the lowest rib, using a non-elastic tape.

Blood pressure is measured using a mercury sphygmomanometer.

Hydration status is assessed by measuring the urine specific gravity of morning urine sample using a refractometer (SUR-NE, ATAGO, Tokyo, Japan).

Subjective evaluation of lower limb soreness is measured using a Visual Analog Scale before and after a wall-sit exercise, with the duration individually adjusted to 65% of each participant's maximum hold time. The Visual Analog Scale consists of a 100 mm line ranging from 0 mm (no soreness) to 100 mm (unbearable soreness).

Heart rate variability is measured using an ECG device (DiANS PF8, DIMEA Group, Olomouc, Czech Republic). ECG recording is performed during the orthoclinostatic maneuver in a calm room without acoustic and visual disturbances.

The level of physical activity is assessed using a wrist-worn tri-axial accelerometer (AX3, Axivity, Newcastle, United Kingdom). This accelerometer provides detailed data on the intensity, frequency, and duration of physical activity.

Sedentary behavior is assessed using a wrist-worn tri-axial accelerometer (AX3, Axivity, Newcastle, United Kingdom). This accelerometer identifies frequency and duration of sedentary bouts.

Sleep duration and quality is assessed using a wrist-worn tri-axial accelerometer (AX3, Axivity, Newcastle, United Kingdom). This accelerometer detects sleep patterns and provides total sleep time, sleep efficiency, and the timing of sleep periods.