A new nitro-nitrosyl complex [RuNO(Phen)(NO2)2OH] (1) was synthesized and characterized by X-ray diffraction, where Phen = 1,10-phenanthroline. The complex was crystallized in two different modifications without (1) and with solvent molecule of DMF (1a). The photolysis process together with the determination of the quantum yield of NO release was investigated in acetonitrile solution using a special flow-through system for simultaneous registration of infrared (IR) and optical absorption (UV-vis) spectra under irradiation with 450 nm. The quantum yield of photoinduced NO release was 4.0±0.2%. DFT calculations have shown that the main contribution to the absorption band at 450 nm is made by the HOMO/HOMO-1→LUMO transitions, which are represented by the transfer of electron density from the -OH and -NO2 ligands to the orbitals located on the Ru-NO bond. The dark and photoinduced cytotoxicity of the complex was studied against the human breast adenocarcinoma (MCF-7), lung carcinoma (A549) cell lines and human non-tumor lung fibroblasts (MRC5). Complex shows a low cytotoxicity on MCF-7 cells (〖IC〗_50^dark = 90.6±6.2 μM and 〖IC〗_50^(irr.) = 95.3±11.4 μM) and a moderate dark cytotoxicity on A549 and MRC5 cells (〖IC〗_50^dark = 33.4±2.6 μM and 〖IC〗_50^dark = 62.6±3.1 μM, respectively), that slightly increase after irradiation (〖IC〗_50^(irr.) = 21.2±3.3 μM and 〖IC〗_50^(irr.) = 47.2±2.3 μM, respectively).