The present study reports the new pyridine-containing analogs bearing thiazolidinone moieties (1–15), which were synthesized and their biological activity was assessed against thymidine phosphorylase and α-glucosidase enzymes. Some of the analogues with the same basic structure but different substituents were discovered to have strong potential against the targeted enzymes, while other analogues were found to have medium to moderate activity in the tested series. These analogues showed a varied range of inhibition in the case of thymidine phosphorylase, 1 (IC50 = 5.70 ± 0.20 µM), 5 (IC50 = 4.50 ± 0.50 µM), 6 (IC50 = 4.60 ± 0.20 µM) and 12 (IC50 = 6.30 ± 0.20 µM), compared with standard 7-Deazaxanthine, 7DX (IC50 = 32.68 ± 1.20 µM). Similarly, excellent inhibitory profiles were also shown by these analogues in the case of alpha-glucosidase inhibition activity, 1 (IC50 =1.80 ± 0.20 µM), 5 (IC50 =1.40 ± 0.10 µM) 6 (IC50 =2.50 ± 0.60 µM) and 12 (IC50 =3.10 ± 0.30 µM) when compared with standard drug acarbose (IC50 = 3.30 ± 1.30 µM). These analogues were also subjected to molecular docking studies against using the PDB id of 1UOU for thymidine phosphorylase (PDB Id: 1UOU) and alpha-glucosidase ((PDB Id: 3w37), for exploration of binding interactions of molecules with active site of the enzymes. ADME studies also predicted the drug-like properties of these analogues.