Ashbya gossypii is a filamentous fungus which overproduces riboflavin as a pseudo-secondary metabolite. Vitamin E supplemented at 1, 2.5 and 5 mu M levels in the growth medium of A. gossypii increased the extracellular secretion of riboflavin and at 50, 100 and 240 mu M levels reduced the biomass and riboflavin yield. With 2.5 mu M vitamin E total riboflavin production and extracellular riboflavin secretion on day 2 was higher than non-supplemented control. By day 3 the production in supplemented was nearly the same as in non-supplemented, but the intracellular riboflavin levels were lower and extracellular levels higher. Supplemented cells showed increased levels of catalase, glutathione peroxidase, lipid peroxides and membrane lipid peroxides, and decreased glutathione indicating that vitamin E, a well-known antioxidant, had acted as a pro-oxidant at low levels of 2.5 mu M and had increased the oxidative stress. Menadione, a well known oxidant also increased riboflavin production and secretion at 1.0, 2.5 and 5.0 mu M level. This is the first report were vitamin E and menadione effects support the concept that overproduction of riboflavin is a stress induced phenomenon. These findings are not only of scientific interest but also useful for improving the industrial production of riboflavin. (C) 2009 Elsevier Ltd. All rights reserved.