To develop pharmacologically active molecules, a series of analogues of 3-(1,2-dihydro-6-substituted phenyl-2-thioxopyrimidin-4-yl)-1-methylquinolin-2(1H)-one (compounds 4a–j) were synthesized through condensation and ring-closure reactions, yielding novel pyrimidine derivatives. These newly synthesized 2-quinolone-linked pyrimidin-2-thiol analogues were structurally characterized using various spectroscopic techniques. The compounds were evaluated for their antioxidant activity using the DPPH (2,2-diphenyl-1-picrylhydrazyl) assay, and for anti-Alzheimer potential through acetylcholinesterase (AChE) inhibition, assessed via Ellman’s method. Among the series, compounds 4c and 4e, containing electron-donating groups on the phenyl ring, showed excellent radical scavenging activity. In AChE inhibition studies, compound 4c [3-(1,2-dihydro-2-thioxo-6-p-tolylpyrimidin-4-yl)-1-methylquinolin-2(1H)-one] exhibited notable inhibitory activity, likely due to the influence of the p-tolyl moiety.Compound 4g [3-(1,2-dihydro-6-(3-nitrophenyl)-2-thioxopyrimidin-4-yl)-1-methylquinolin-2(1H)-one] also demonstrated considerable AChE inhibition.Kinetic studies provided further insights, compound 4e showed a competitive inhibition pattern at both low and high inhibitor concentrations. Compound 4g displayed mixed-type inhibition, suggesting its ability to interact with both the catalytic anionic site (CAS) and the peripheral anionic site (PAS) of AChE. Molecular docking of compounds 4a–j supported the experimental findings, with docking scores ranging from moderate to good compared to the reference drug. Notably, compound 4g had the best docking score of −10.134, forming a key hydrogen bond with PHE295, correlating well with its strong AChE inhibition. Further molecular dynamics simulations of compound 4g within the AChE binding site confirmed its stable interactions and potential binding mode. Based on these findings, compound 4g emerges as a promising lead molecule for the development of novel anti-Alzheimer agents. Finally, drug-likeness properties of all synthesized compounds were evaluated, supporting their potential for further development.