The collagenolytic enzyme complexes secreted by two thermophilic Actinomycetes newly isolated from Bulgarian soils were characterized for their possible application for treating collagen-rich materials. The strains were identified as belonging to genera Microbispora and Thermoactinomyces. The molecular masses of the enzymically active fractions determined by gel chromatography on Sephadex G-100 showed values between 46 and 140 kDa. The enzyme complexes produced by both strains were stable against temperature changes and high ethanol concentrations, which allows effective elimination of different by-products (pigments, nutrient medium components, and metabolites). The comparison with clostridiopeptidase A as well as the inhibitory analysis showed that these enzymes might be considered as "collagenase-like" enzymes. Preliminary experiments showed that a mixture of both enzyme preparations could be applied for pre-treatment of collagen-rich raw materials, e.g., in the pelt industry. (c) 2004 Elsevier Ltd. All rights reserved.