Tryptanthrin is a valuable tetracyclic alkaloid, which displays a wide variety of biological activities. The application of this type of scaffold as a starting material for the discovery of new drug candidates is of major importance in medicinal chemistry. In this work, we report one of the few examples of tryptanthrin-based multicomponent reaction approaches for drug discovery, and the first using the Petasis reaction. The optimized BINOL-catalyzed reaction conditions allowed the synthesis of a library of new tryptanthrin derivatives bearing considerable structural diversity. An asymmetric version was also established, achieving the desired enantiomerically pure derivative with 99% ee and 71% yield. The resulting library was screened against one Gram-positive and one Gram-negative bacteria, two yeasts, and three filamentous and four dermatophyte fungal strains with clinical relevance, with compound 5bea displaying moderate fungicidal activity. © The Royal Society of Chemistry and the Centre National de la Recherche Scientifique.