STUDY OF THE ANTIOXIDANT ACTIVITY OF SHEEP WHEY PROTEIN IN VITRO AND IN C2C12 CELL LINE Efthalia kerasioti, Alexandros Priftis, Stefanos Aivazidis, Dimitrios Stagos, Demetrios Kouretas Department of Biochemistry & Biotechnology, University of Thessaly, Larisa, 41221 Introduction. Whey is considered as by-product of cheese manufacturing, but it is also described as functional food (1, 2). Whey is the liquid that remains in an aqueous environment after removal of casein. We examined 1) in vitro the scavenging activity of sheep whey protein and compared with that one of beef protein, soy protein and cow whey protein and 2) its antioxidant properties in C2C12 muscle cell line. Methodology. It was determined in vitro the ability of proteins to neutralize DPPH•, ABTS•+, OH•, O2·- radicals as well as their reducing power. Also, we determined the levels of glutathione (GSH) and free radicals (ROS) by flow cytometry, as well as the levels of thiobarbituric reactive substances (TBARS) (marker of lipid peroxidation) spectrophotometrically in muscle cell line C2C12 under the influence of the oxidative agent t-BOOH. Results. All proteins showed a dose-dependent free radical scavenging activity. The IC50 value was determined, showing the concentration that has the ability to scavenge radical by 50% while for reducing power assay, it was determined the RP0.5AU, the concentration that produces an absorbance of 0.5 at 700 nm (Table 1). Table1. IC50 (mg/ml) and RP0.5AU (mg/ml) for sheep whey protein, beef protein, soy protein and cow whey protein. Values are presented as mean±sem. DPPH• ABTS•+ OH• O2·- ΑΔ 3,1±0,06 4,1±0,15 1,8±0,20 - 1,3±0,05 Beef protein 1,3±0,12 0,6±0,02 0,85±0,05 2,5±0,2 0,9±0,03 Soy protein 2,2±0,31 3,1±1,00 1,1±0,34 - 3,1±1,20 Cow 8,2±0,55 3,9±0,09 1,7±0,10 - 4,8±0,02 Proteins Sheep whey protein whey protein As regards the muscle cell line C2C12, sheep whey protein protected from the oxidative agent t-BOOH. Specifically, sheep whey protein at concentrations 0.78, 1.56, 3.12 and 6.24 mg/ml, increased GSH levels by 25%, 112%, 118% και 138% respectively, lowered TBARS levels by 21%, 15%, 25% και 24% respectively and decreased ROS levels by 2.8%, 12.8%, 16.4% και 41.4% respectively, compared to t-BOOH treatment alone. References 1)Kerasioti E, Kiskini A, Veskoukis A, Jamurtas A, Tsitsimpikou C, Tsatsakis AM, Koutedakis Y, Stagos D, Kouretas D, Vaios K. Effect of a special carbohydrateprotein cake on oxidative stress markers after exhaustive cycling in humans. Food Chem Toxicol. 2012 50(8):2805-2810. 2) Kerasioti E, Stagos D, Jamurtas A, Kiskini A, Koutedakis Y, Goutzourelas N, Pournaras S, Tsatsakis AM, Kouretas D. Anti-inflammatory effects of a special carbohydrate-whey protein cake after exhaustive cycling in humans. Food Chem Toxicol. 2013 13:61-66. This work was co-financed by the European Union (European Social Fund – ESF) and Greek national funds through the Operational Program ‘‘Education and Lifelong Learning’’ of the National Strategic Reference Framework (NSRF) – Research Funding Program: Heracleitus II. Investing in knowledge society through the European Social Fund.