Blood sample collection and analysis
Tecan, muscle damage
BLOOD, OXIDATIVE STRESS, INFLAMMATION
All venous blood collections were performed by a qualified medical technologist during the blood collection procedures. The samples were centrifuged at 3000 rpm (1006.2 g) for 10 minutes under 4°C. Plasma was collected in the supernatant and frozen at −80°C for later analysis.Blood samples were collected at the 10, 20, 30, and 40 km marks during the exercise challenge. Blood glucose was measured using an automated glucose analyzer (YSI Life Sciences, Yellow Springs, OH, USA). Using an automatic photometric analyzer (Hitachi 7020, Japan), commercially available kits were applied to determine the plasma concentrations of NEFA (WAKO, Germany), CK (Cayman Chemical Company, ANN Arbor, MI, USA), LDH, and UA (Kanto Chemical, Kanagawa, Japan). For TAC and MDA, absorbance was measured at wavelengths of 734 and 534 nm, respectively, using the UV-visible spectrophotometer (Tecan GENios, A-5082, Austria). Similarly, the absorbance of cytokine IL-6 and serum TNF-α was read at 450 and 570 nm wavelengths to calculate concentrations in the serum samples. BioLegend reagents (Bio-Legend Inc, San Diego, CA) were used for the spectrophotometer measurements. A more detailed account of the evaluation for energy metabolism, oxidative stress, inflammation, and muscle damage can be found in our previously published studies [