The presence of various biotypes of Lactobacillus helveticus seems to be related to the specificity of the ecosystem, which is one of the main determinants of the unique features of cheese produced in particular regions. So far, it has been proved that even biotypes isolated from the same niche differ significantly from each other and many features exhibited by these bacteria are strain-dependent. Therefore, the new strains of L. helveticus T104 and T105 (isolated from traditionally fermented Polish dairy product) were applied in the production of ripened cheeses due to their potential health-promoting properties. We determined changes in fat, fatty acids, protein, amino acids, and some physicochemical characteristics, e.g. antioxidant properties of the produced cheeses (in three stages of ripening). Tricine-SDS-PAGE and MALDI-TOF MS analysis showed some differences in protein and peptide profiles. Final products obtained using L. helveticus T105 exhibited the greatest amount of free amino acids, which are important precursor of cheese aroma and flavor. The research indicated that the tested strains could be applied in the manufacture of cheeses. Moreover, the cheese produced using the said strain exhibited the highest free radical scavenging capacity (88.89% after pre-ripening and 92.74% in the final products) even in comparison to the control cheese variant produced using the industrial L. helveticus strain. Obtained findings indicate that the tested strains exhibit technological and functional potential that provide a reference for further study and might contribute to the development of functional food products with novel, valuable characteristics.