Thermomyces lanuginosus strains RT9 and MH4 were studied to find favourable cultivation conditions and to compare their abilities to produce xylanolytic enzymes in three media on different substrates at 50 degrees C or 55 degrees C under shake-culture conditions. Both organisms produced xylanases free of cellulase at widely different levels in all cultivation conditions employed. Wheat bran, corn cobs and xylan induced xylanases in increasing order of producing with both cultures. T. lanuginosus RT9 demonstrated the highest xylanase production in all cultivation conditions but with lower soluble protein, reducing sugar, beta-xylosidase and debranching enzymes levels (arabinosidase, acetylxylanesterase, mannanase) when compared to T. lanuginosus MH4. The study reveals that xylanase production was highly influenced by nitrogen sources and their concentrations and by the initial pH in the cultures, The two strains may therefore be unique, when technical application is considered in terms of the quantity and quality of the xylanolytic enzymes produced.