Despite the vast availability of antibiotics, bacterial infections remain a leading cause of death worldwide. In an effort to enhance the armamentarium against resistant bacterial strains, 1,2,3-triazole (5a–x) and sulfonate (7a–j) analogues of natural bioactive precursors were designed and synthesized. Preliminary screening against two Gram-positive (Streptococcus pneumoniae and Enterococcus faecalis) and four Gram-negative bacterial strains (Pseudomonas aeruginosa, Salmonella enterica, Klebsiella pneumoniae, and Escherichia coli) was performed to assess the potency of these analogues as antibacterial agents. Among all triazole analogues, 5e (derived from carvacrol) and 5u (derived from 2-hydroxy 1,4-naphthoquinone) bearing carboxylic acid functionality emerged as potent antibacterial agents against S. pneumoniae (IC50: 62.53 and 39.33 μg/mL), E. faecalis (IC50: 36.66 and 61.09 μg/mL), and E. coli (IC50: 15.28 and 22.57 μg/mL). Furthermore, 5e and 5u also demonstrated moderate efficacy against multidrug-resistant E. coli strains and were therefore selected for further biological studies. Compound 5e in combination with ciprofloxacin displayed a synergistic effect on multidrug-resistant E. coli MRA11 and MRC17 strains, whereas compound 5u was selective against E. coli MRA11 strain. Growth kinetic studies on S. pneumoniae and E. coli treated with 5e and 5u showed an extended lag phase. 5e and 5u did not show significant cytotoxicity up to 100 μg/mL concentration on human embryonic kidney (HEK293) cells. Transmission electron microscopic (TEM) analysis of bacterial cells (S. pneumoniae and E. coli) exposed to 5e and 5u clearly showed morphological changes and damaged cell walls. Moreover, these compounds also significantly inhibited biofilm formation in S. pneumoniae and E. coli strains, which was visualized by scanning electron microscopic (SEM) analysis. Treatment of larvae of Galleria mellonella (an in vivo model for antimicrobial studies) with 5e and 5u did not cause an alteration in the hemocyte density, thereby indicating lack of an immune response, and were nontoxic up to a concentration of 2.5 mg/mL.