Aspergillus flavus reduces cottonseed value by producing aflatoxins during seed infection. Xyloglucans are plant-derived polysaccharides critical to the structural integrity of cell walls of plants. Cottonseed hulls contain relatively high xylan concentrations. A. flavus AF36 secreted a xylanase activity when grown on a medium containing xylan as a sole carbon source. Enzyme activity was assayed using xylan conjugated to Remazol Brilliant Blue R as substrate. A survey of several closely related fungi revealed secretion of a similar xylanase activity from A. oryzae, A. parasiticus, A. nomius, and A. tamarii, but not from A. sojae. This xylanase was thermostabile up to 60 C and was tolerant of a wide pH range (5.0-8.0) with no optimum. A concentrated sample of the AF36 xylanase activity was subjected to gel filtration chromatography on a P-60 column. A small protein peak that coincided with a peak of xylanase activity eluted very close to the included volume of the column. Data suggest this hydrolytic activity is associated with a low molecular weight (<20kD) thermostabile protein, and is consistent with a highly mobile (diffusible), environmentally stable plant cell wall degrading factor. The identified xylanase activity may play a role in infection and contamination of cottonseed by A. flavus.