A novel series of substituted 1,4-naphthoquinone derivatives were synthesized and evaluated for their antibacterial and antifungal activity. The structures of the novel products were characterized by spectroscopic methods. Among the tested compounds, 2,2,3,3-alkoxy substituted naphthoquinones, S,O-substituted naphthoquinone, and N,S-substituted naphthoquinone derivatives are the most potent antifungals against C. tenuis. 2,3-Thio-2,3-alkoxy substituted naphthoquinones are the most effective antifungal compounds against A. niger. Supplemental materials are available for this article. Go to the publisher's online edition ofPhosphorus, Sulfur, and Silicon and the Related Elementsto view the free supplemental file.