Synthesis, Crystal Structure, DFT, MEP, FMOs, Hirshfeld Analysis and Antitumor Activity Assay of 4-[(4-Fluorophenyl)amino]-6-pentylpyrido[4,3-d]pyrimidin-5(6H)-one

Abstract Pyrido[4,3-d]pyrimidine derivatives represent an important class of heterocyclic compounds that play an indispensable role in organic synthetic chemistry and drug discovery. As homologues of pyrimidine antagonists, the pyrido[4,3-d]pyrimidine scaffold holds significant theoretical value and application potential in biochemical research and clinical settings such as tumor chemotherapy. In the present study, the target compound 4-[(4-fluorophenyl)amino]-6-pentylpyrido[4,3-d]pyrimidin-5(6H)-one was synthesized via multi-step organic reactions using malononitrile as the starting material, with its synthetic route encompassing key steps including addition, cyclization, substitution, and Suzuki coupling. Four spectroscopic characterization techniques were utilized to systematically confirm the chemical structure of the target compound. Additionally, its crystal structure was unambiguously determined by X-ray single-crystal diffraction analysis, while the optimized molecular structure was obtained through density functional theory (DFT) calculations. The results demonstrated that the DFT-optimized molecular structure is in good agreement with the crystal structure measured by X-ray single-crystal diffraction, thereby validating the reliability of the theoretical calculations. Furthermore, systematic property investigations were conducted on the target compound, including infrared characteristic vibration analysis, molecular electrostatic potential (MEP) distribution, frontier molecular orbitals (FMOs) analysis, Hirshfeld surface analysis, and in vitro antitumor activity evaluation. These findings provide a robust experimental and theoretical foundation for the further structural modification and pharmacological research of this class of pyrido[4,3-d]pyrimidine derivatives.