Twelve Lactobacillus isolates from Brazilian starter-free ovine cheeses were evaluated for their probiotic potential. The strains were identified by 16S rDNA sequencing as Lactobacillus plantarum (7), Lb. brevis (2), Lb. casei (2) and Lb. parabuchneri (1). All strains showed variable resistance to gastric juices and relative tolerance to pancreatin and bile salts. Only five strains of Lb. plantarum could not deconjugate the sodium salt of taurodeoxycholic acid. Autoaggregation ability after 24 h was above 50% and hydrophobicity was higher than 60% for most strains. All lactobacilli could inhibit linolenic acid oxidation, except Lb. parabuchneri strain, whereas none of them could scavenge DPPH radical. β-Galactosidase activity ranged from 47·7 to 2503 Miller units. Inhibition of food pathogens Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, Escherichia coli and Salmonella typhimurium was demonstrated and the production of organic acids could be associated with this effect. The Lactobacillus strains from Brazilian regional ovine cheese showed interesting functional characteristics, mainly the strains Lb. brevis SM-B and Lb. plantarum SM-I. Both presented high acid tolerance. In addition, Lb. brevis SM-B also displayed remarkable antioxidant activity and Lb. plantarum SM-I was the highest β-galactosidase producer, exhibited high autoaggregation and hydrophobicity properties.