Fractionation of the polyphenols constituting a food grade lingonberry extract (Vaccinium vitis-idaea) highlighted a composition more complex than described until now in the berry. Procyanidins B1, B2, and A2 were identified by UPLC/ESI-MS(2) along with the presence of other flavanol oligomers. Processing induced the release of large amounts of aglycones for ferulic acid, p-coumaric acid, and quercetin. The described anthocyanic composition of lingonberry was completed with hexoside derivatives of peonidin, petunidin, malvidin, and delphinidin. Besides confirmation of in vitro antioxidant activity, in vivo study was performed on rats fed a diet inducing oxidative stress. Supplementation with lingonberry extract significantly decreased the total oxidant status and favorably affected antioxidant defense enzymes in red blood cells and liver. A drop in the serum reduced glutathione level was also prevented, and uric acid was maintained at low level, confirming the antioxidant activity of the extract (5% proanthocyanidins) from a dosage of 23 mg/kg of body weight.