Cultural and Nutritional Factors that Control Rubratoxin Formation

Penicillium rubrum P3290 was grown in a glucose-salts broth with various supplements and cultures were incubated under different conditions. Maximum mold growth and toxin production was favored by 20% glucose. The highest yield of rubratoxin A appeared in 6 days and of rubratoxin B in 12 days. Rubratoxin A disappeared from cultures after 22 days of incubation; rubratoxin B was gone after 26 days. All amino acids tested supported synthesis of rubratoxin B; highest yields were obtained when the medium contained asparagine or glutamic acid. Glycine, methionine, and serine supported good yields of rubratoxins A and B (3.3 to 6.8 mg of rubratoxin A/100 ml; 36.0 to 46.2 mg of rubratoxin B/100 ml), whereas aspartic acid, lysine, tyrosine, and tryptophan allowed production of only rubratoxin B. Yields of toxin were enhanced by 0.2–0.3% ammonium sulfate and mold growth was maximal when 5% was in the medium. Inorganic phosphate generally enhanced toxin production; it was optimal at 0.6% and inhibitory at 9.6%. Zinc and iron were required for toxin production but manganese and magnesium were not. A temperature of 25–28 C was optimal; reducing it to 18 C caused a 65% decline in toxin production. When the temperature was increased from 28 to 37 or 45 C, a 60–85% decline in, or complete inhibition of, toxin synthesis occurred because the fungus grew poorly at the higher temperatures. An initial pH value of 4–5 enhanced toxin synthesis, although limited toxin production occurred at pH 10 but not at pH 1. A relative humidity of 67% allowed rubratoxin formation in 7-day old cultures, of 77% supported highest yields of toxin, and 85% allowed accumulation of moderate amounts of rubratoxin.