In the course of a screening program directed toward the isolation and evaluation of inhibitors of carboxyl (acid) proteinases, we have found 0.39% of the investigated strains of microorganisms as producers of pepsin inhibitors. In more than 1543 strains which were isolated from various geographical areas and deposited with the IMET Culture Collection, Jena-GDR, and which belong to different genera of actinomycetes such as Streptomyces, Actinomadura, Actinoplanes, Micromonospora and non-identified strains, we have found 6 Streptomyces strains only. Surprisingly, the screening efficiency of the used casein agar spot test and the fibrin-blue-technique was very low. As compared with the inhibitor activity of pepstatin the yield of the fermentation broth of 3 wild type strains ranged from 6 to 27 micrograms/ml. A significant influence of the composition of culture fluids on the inhibitor yield was observed. Supplements by crude glutamine or mixtures of amino acids in culture fluids increased the yield from 3 to 15 times as compared with that of fermentation broth without supplements. After NMG-mutagenic treatment and use of familiar selection procedures 3.1% inhibitor-blocked mutants were detected among 480 strains selected.