Methods were developed for bioproduction and extraction of rubratoxin B from liquid cultures of Penicillium rubrum P-13 (NRRL A-11785). A maximum of 874.7 mg of toxin per liter of medium was attained in 21 days using stationary cultures of Mosseray9s simplified Raulin solution enriched with 2.5% malt extract. Malt extract was required for rubratoxin production. Rubratoxin was not produced in either shake flasks or in fermentors with restricted aeration. Crystalline toxin was obtained by liquid-liquid extraction of concentrated culture medium with ethyl ether. Adhering colored impurities were removed by column chromatography and by recrystallization from acetone. Images