Lawsone (2-hydroxy-1, 4-napthoquinone),also known as hennotannic acid, is an orange red dye used as a popular skin and hair colorant. The dye is produced in the leaves of Lawsonia inermis L, often referred to as the ‘‘henna’’ tree. In this study, we report the production of lawsone by an endophytic fungus,Gibberella moniliformis isolated from the leaf tissues of Lawsonia inermis. The fungus produced the orange-red dye in potato dextrose agar and broth, independent of the host tissue. Presence of lawsone was confirmed spectrometrically using HPLC and ESI–MS/MS analysis. The fragmentation pattern of lawsone was identical to both standard lawsone and that extracted from plant tissue. This is a first report of lawsone being produced by an endophytic fungus, independent of the host tissue. The study opens up interesting questions on the possible biosynthetic pathway through which lawsone is produced by the fungus.