In the current era, an ever-emerging threat of multidrug-resistant (MDR) pathogens pose serious health challenges to mankind. Researchers are uninterruptedly putting their efforts to design and develop alternative, innovative strategies to tackle the antibiotic resistance displayed by varied pathogens. Among several naturally derived and chemically synthesized compounds, quinones have achieved a distinct position to defeat microbial pathogens. This review unleashes the structural diversity and promising biological activities of naphthoquinones (NQs) and their derivatives documented in the past two decades. Further, realizing their functional potentialities, researchers were encouraged to approach NQs as lead molecules. We have retrieved information that is dedicated on biological applications (antibacterial, antifungal, antiparasitic) of NQs. The multiple roles of NQs offer them a promising armory to combat microbial pathogens including MDR and the ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) group. In bacteria, NQs may exhibit their function in the following ways (1) plasmid curing, (2) inhibiting efflux pumps (EPs), (3) generating reactive oxygen species (ROS), (4) the inhibition of topoisomerase activity. Sparse but meticulous literature suggests the mechanistic roles of NQs. We have highlighted the possible mechanisms of NQs and how the targeted drug synthesis can be achieved via molecular docking analysis. This bioinformatics-oriented approach will explicitly lead to the development of effective and most potent drugs against targeted pathogens. The mechanistic approaches of emerging molecules like NQs might prove a milestone to defeat the battle against microbial pathogens.