As pyrazoles are familiar for showing various biological properties, a series of novel isocoumarin tethered carbothioamide linked pyrazole derivatives were prepared successfully in the presence of environmental favor conditions and evaluated for their pathogenic resistance against four bacteria and two fungi, showing significant activity against different microbial except compound 5d. Among the active compounds, 5f displayed highest antimicrobial property. In fact, 5b and 5l motifs showing good inhibitory activity. Further, a detailed structure activity relationship also discussed, which revealed that the presence of electron withdrawing -NO2 group in 5f compound may delivered highest biological property, while the amino group attached in compound 5d delivered inactive antibacterial property. Moreover, computer aided studies of biologically effective compounds to determine the interactions and docking score with the crystal structure of Escherichia coli 24 kDa DNA gyrase subunit B in complex with clorobiocin. It was observed that the amino acid residues present in the active binding site of the protein1KZN can frequently interact with targets.