Zn (II), Cd (II), Hg (II) and U (VI)O22+ complexes of water‐soluble thiosemicarbazone ligand (NaH3PyTSC) have been prepared and characterized using various techniques. Fourier transform‐infrared (FT‐IR) demonstrated that NaH3PyTSC ligand behaves as a binegative NOS tridentate in [Hg(H2PyTSC)(H2O)]2 and [UO2(H2PyTSC)(H2O)]2 complexes via the deprotonated SH, (C=N)az groups from one molecule and SO3− group from another molecule, while it behaves as a binegative NNSO tetradentate in [Cd(H2PyTSC)(H2O)2]2 complex through the deprotonated SH group, the SO3− group and the nitrogen of both the (C=N)az and (C=N)py. Finally, it behaves as a binegative OO bidentate in [Zn(H2PyTSC)(H2O)2]2·2H2O complex by the deprotonated OH group from one molecule and SO3− group from another ligand molecule. The spectral data suggest a tetrahedral coordination around Hg (II) and Zn (II) ions, and an octahedral coordination around Cd (II) and U (VI)O22+ ions. The NaH3PyTSC ligand exhibited maximum luminescent intensity at 501 nm, while Zn (II), Cd (II) and Hg (II) chelates show emission bands at 459, 458 and 358 nm, respectively. Two comparable methods were used to estimate various thermodynamic parameters. Cyclic voltammetry has been studied for Cd (II) complex in solution. Different biological applications of the isolated complexes have been estimated. It was found that [Cd(H2PyTSC)(H2O)2]2 showed the most effective antioxidant and anticancer activity.