New Schiff bases bearing 5-nitroisatin-thiocarbohydrazone ( 1-9 ) have been prepared. The chemical structures of these compounds have been clarified by spectroscopic methods (IR, 1 H NMR, and 13 C NMR), and elemental analysis. Besides, they have been tested for antimicrobial activity. The antimicrobial activities of the novel compounds have been evaluated using a twofold serial dilution method for Minimum Inhibitory Concentration (MIC) against three gram-positive bacteria, three gram-negative bacteria, and two fungal pathogens. The obtained data showed that compound 7 has excellent activity against gram-strain positive and gram-strain-negative bacteria. Compound 7 showed better activity than amoxicillin, which is one of the standard drugs used against the E. faecalis pathogen, but showed the same activity as tetracycline. Synthesized compounds could be formed the basis for the development of better antibacterial agents. The total antioxidant activity of compounds 3 and 5 showed better antioxidant activity than standard BHT, while compounds 4, 8, 7, 2 , and 6 showed better antioxidant activity than standard Trolox. The urease inhibitory effect of compound 7 was better than thiourea. Also, the structure-activity relationship study showed that the scores of molecular models revealed that the compounds with the highest number of binding with urease enzyme were compounds 3 and 7 with total bond numbers of 19 and 18, respectively.(c) 2022 Elsevier B.V. All rights reserved.