This research focuses on the design of new compounds and their biological applications to address various medical issues and pandemics caused by diverse microorganisms. In this work, two novel isoxazoline derivatives containing chlorine and methoxy groups at the para position were synthesized, characterized by various known techniques and QSAR properties were calculated using Osiris and docking against the protein targets such as 1HNJ, 1KZN and 1T15. The theoretical outcomes are compared with the experimental antimicrobial investigations on B. subtilis, E. coli, S. aureus, K. pneumoniae, C. albicans and A. niger using agar well-diffusion method. The derived derivatives exhibited good results and showed good coincidence with theoretical results. Out of two target compounds, activating methoxy group substituted isoxazoline compound showed better result against bacterial strains (12-16 mm), whereas deactivating chlorine group substituted compound revealed the good result (12-15 mm) against fungus strains. © 2025 Elsevier B.V., All rights reserved.