In the present work, carbonyl pyridoquinolones 8–33 were synthesized and their ability to inhibit gram-positive stains Bacillus subtilis and Staphylococcus aureus, gram-negative stains Escherichia coli and Pseudomonas aeruginosa, and fungi Candida albicans was studied. Carbonyl piperazines attached at C-3 position of pyridoquinolone exhibited good antibacterial activity when compared to the standard drugs ciprofloxacin and gatifloxacin. Indeed carbonyl piperazine derivatives 24–32 when attached at the C-3 position of pyridoquinolone increased the activity of their parent compound 6 than that of carbonyl ureas 8–23 exhibited lower antibacterial activity compared to carbonyl piperazine derivatives 24–32.