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Aflatoxins can be produced on a synthetic medium in submerged culture. Glucose, sucrose, or fructose are the preferred carbon sources, and Casamino Acids are the preferred nitrogen source. Ammonia is almost as good a nitrogen source. Zinc is required at levels of at least 0.4 mg per liter. Concentrations of aflatoxin of 60 to 80 mg per liter (as determined by optical-density measurements of a chloroform extract of the unfiltered broth) can readily be obtained in indented shake flasks; somewhat lower yields were obtained in 5-liter fermentors.

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We have found whole human platelets, granulocytes, and mononuclear leukocytes to possess high affinity for the toxic lipopolysaccharide from all gramnegative bacteria tested. We have extracted these cells and platelets with nbutanol-water; all endotoxin-binding activity resided in the organic phase. These endotoxin-binding extracts did not block serologically active groupings on endotoxins or receptors on the erythrocytes. The specificity of these still crude materials was less than that of the highly purified erythrocyte lipopolysaccharide receptor previously described by us, since they bound some bacterial antigens not related to endotoxins. Depending on source, the n-butanol extracts contained 40 to 52% glycerophosphatides (most active), 15 to 22% sphingomyelin, 17% cholesterol, <2 to 5% triglycerides, and 7 to 13% inactive peptide. The most active substances in the n-butanol extract were soluble in petroleum ether, whereas the peptide and sphingomyelin were not. Thus, no constituent protein, carbohydrate, or nucleic acid was present in the most highly active material. Polyacrylamide gel electrophoresis of the petroleum ether-soluble material showed for each extract one lipid band only, which was well defined and migrated similarly to phosphatidyllipids. Because of the lipidic nature of the inhibitory substances from leukocytes and platelets we also tested the lipid A component of bacterial endotoxins and some of its derivatives. Lipid A inhibited endotoxin coating of erythrocytes. De-O-acylation of lipid A left amide-linked 3-D-hydroxymyristic acid intact and increased the inhibitory activity of lipid A 20fold. Complete de-Oand de-N-acylation destroyed its inhibitory effect. MATERIALS AND METHODSHuman blood was collected into citrate-phosphate-dextrose anticoagulant solution USP (63 ml/pint; ca. 134 ml/liter), and single units were used 978 on July 16, 2020 by guest http://iai.asm.org/ Downloaded from Bacteriol. 95:1572-1579.