We have reported the synthesis of the new thiobarbiturate derivative ligand (1) and its three Zn(II) complexes (2–4). The ligand and complexes were structurally characterized by various techniques such as ATR-FTIR, 1H NMR, 13C NMR, and elemental analysis. The molecular structure of 3 was determined by the single-crystal X-ray diffraction analysis study. The study revealed that it consists of a zig-zag one-dimensional polymeric chain, in which the Zn(II) ion is five coordinated by three oxygen atoms from the ligand, one nitrogen atom from the ligand, and one nitrogen atom from the ancillary ligand (1-methyimidazole). Also, Addison's five-coordinated index of 3 (τ5 = 0.45) showed that Zn(II) ion has a distorted square pyramidal geometry. Ligand (1) and Zn(II) complexes (2–4) were tested using the MTT cell viability assay on two types of cancer cell lines from human prostate cancer (PC-3), colon cancer (HT-29) and a normal cell line of the bronchial epithelium (BEAS-2B). 1 acted as a potent growth inhibitor on HT-29 cells, while 4 showed cytotoxic effects against PC-3 cells. Their antibacterial activity was determined by the agar well diffusion method. Depending on the 2 and 4 doses, it showed activity against both gram (+) and gram (−) bacteria. In terms of anti and anticancer effects, 4 seems to be a biologically highly active complex and promising biomedically.