Characterizing germplasm collections of autochthonous cultivars for fruit quality traits could be a successful approach for selection, improvement of organoleptic quality and levels of antioxidants of crop produce, and development of new market opportunities and coherent strategies for conservation and valorization. The aim of the study was the evaluation of fruit physicochemical traits as well as the content of bioactive compounds and the antioxidant capacity in 25 sweet cherry autochthonous cultivars. Cultivars were a source of statistically significant variation for all evaluated traits. Notably, average fruit ascorbate levels ranged from 34.45 to 244.68 µg g-1 fresh weight (FW) , total flavonoids from 1396.40 to 4694.75 µg quercetin equiv. g-1 FW, monomeric anthocyanins from 4.80 to 360.90 µg g-1 FW, and total antioxidant capacity from 1.53 to 2.58 nmol Trolox equiv. mg-1 FW. Fruit profiling of eight cultivars by high-resolution mass spectrometry identified a total of eight different anthocyanins and twenty-five non-anthocyanin polyphenolic compounds - mostly coumaroylquinic acid and neochlorogenic acid. Among the better-performing cultivars for fruit quality traits, Mulegnana Nera and Pagliarella shared high fruit levels of phenolics, flavonoids and antioxidant capacity. This is a forerunner work on the characterization of genetic resources, which is critical to researchers and breeders for exploitation of the genetic potential of cultivars and for their conservation. © 2016 Society of Chemical Industry.