Chemical, crystal structures, properties, thermal behaviour, kinetic parameters, and antitumor activity of synthesized 3-((2,5-Difluorophenyl)diazenyl)-6-ethyl-4-hydrox-2H-pyrano[3,2-c]quinoline-2,5(6H)-dione (DFPDAEHPQD) were investigated and characterized. Different techniques are used to identify the structure of DFPDAEHPQD, such as 1H nuclear magnetic resonance (NMR) 13C NMR, X-ray diffraction (XRD) analysis, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and thermogravimetric analysis (TGA) measurements. XRD analysis showed that DFPDAEHPQD had a polycrystalline nature, and TGA results illustrated that the DFPDAEHPQD was thermally stable up to about 300 oC. Optical parameters and absorption coefficient of DFPDAEHPQD were calculated using diffused reflectance (DR) measurements in a wide range of wavelengths (200 to 1600 nm). Two optical bandgap values (2.35 and 2.25 eV) were obtained in the applied range of photon energy using Kubelka–Munk theory (KMT). The first value (2.35 eV) is responsible for the direct transition, while the other value (2.25 eV) is responsible for the indirect transition. The AC conductivity obeyed the universal power-law σAC(ω) = Aωs. Moreover, DFPDAEHPQD showed highly significant antitumor activity and cytotoxicity against two human cancer cell lines: HepG-2 and HCT-116, with the corresponding IC50 values of 3.07 and 7.15 μg/ml, respectively. Finally, we hope that DFPDAEHPQD turns out to be environmentally friendly and more energy efficient a novel semiconductor for safe use in various electronic and biomedical applications.