The Effects of Vitamin D Supplementation on Aerobic Fitness in Athletes

A medical screening questionnaire will be completed prior to the test. Athletes resting blood pressure will be measured twice to rule out resting hypertension. After a standardised warm-up, starting speed will be 8/km/hr at 1% incline, until 17/km/hr, after which incline will increase by 1% every minute until VO2max is met. A metabolic cart (Metalyzer 3B) will measure the athlete's ventilation (oxygen uptake and carbon dioxide output) throughout the test.

The test will be terminated when two of the following criterion are met*:

Respiratory exchange ratio reaches/exceeds 1.15 AND VO2 plateau observed OR Heart rate within 10bpm of age-predicted maximum

• Except when volitional exhaustion occurs Athletes will be asked to refrain from strenuous activity for 24 hours prior to VO2max testing in order to control for last bout effects.

Post-exercise lactate concentration (age and gender adjusted) will further-verify that VO2max was attained using a Lactate Pro device.

Hand grip dynamometry will be used to assess athletes grip strength. This will be measured on both dominant and non-dominant hands three times. The athlete will be asked to hold the device and grip as tightly as possible whilst holding the device alongside their body. There will be a 10 second rest period between measurements.

Athletes will be asked to reach upwards as far as possible, from a standing position and mark this point on a wall. After this they will be asked to perform a counter-movement jump a total of 3 times with a 10 second rest-period between jumps, reaching as high as possible and making a mark each time. Vertical jump height is calculated from the difference between standing reach height and the best height reached when performing a counter-movement jump.

Forced expiratory volume at 1 second (FEV1) and forced vital capacity (FVC) will be measured. A calibrated Carefusion® MicroLab portable spirometer will be used to measure these variables.The athletes will need to take a maximal inhalation and then perform a full exhalation as fast as possible into a disposable mouthpiece.

This procedure will be repeated three times and an average taken at each time-point. The timing of repeats will be at the athletes discretion.

Dietary vitamin D and calcium intake will be estimated using a validated food frequency questionnaire at one time point (baseline only). This is due to studies highlighting equivocal variation in dietary vitamin D intake throughout the year.

24hr recalls will be completed at baseline and at 12-weeks in order to derive mean nutrient intakes for the vitamin D and placebo treatment groups respectively.

An online questionnaire will allow athletes to record if they have any symptoms that are associated with upper respiratory infections. The athlete will be asked to mark a series of boxes to indicate how many symptom(s) if any that they have had each week.

The 'Modified Recent Physical Activity Questionnaire' produced by the Medical Research Council (a validated physical activity questionnaire) will be completed on two occasions to estimate physical activity levels.

Body composition will be measured using the BodPod, employing air plethysmography (displacement of air) to accurately measure percentage fat mass and percentage fat-free mass.

25-hydroxyvitamin D (25(OH)D) will be quantified, in serum, using liquid chromatography-tandem mass spectrometry (LCMS/MS). Other biomarkers of vitamin D metabolism (adjusted calcium and parathyroid hormone) will be quantified using a clinical chemistry analyser and commercially available enzyme-linked immunosorbent assay.

A range of immune markers such as interleukin-6, cathelicidin (LL-37) and C-reactive protein will be measured using a commercially available multiplex immunoassay.