Our search for an inhibitor of thermolysin yielded phosphoramidon (I), a compound previously identified as an Ehrlich reaction-positive metabolite in a culture filtrate. Thermolysin, a metalloendopeptidase from Bacillus thermoproteolyticus, was targeted for inhibition. An inhibitor was isolated from actinomycetes, with strain MD706-Y4 (Streptomyces tanashiensis) selected for detailed production and isolation studies. The inhibitor's concentration was quantified via thermolysin-inhibiting activity. Purification involved activated carbon adsorption, DEAE-Sephadex A-25 chromatography, and Sephadex LH-20 chromatography, yielding a compound confirmed as phosphoramidon by comparison with an authentic sample. Physicochemical characterization revealed: melting point 173~178°C (dec.); specific rotation [α]D²⁰ -33.6° (c=1.0, H₂O); elemental composition consistent with C₂₃H₃₂N₈O₁₀P₁Na₂; solubility in water, methanol, and dimethylsulfoxide; positive Ehrlich, ammonium molybdate-perchloric acid, and Rydon-Smith reactions; negative ninhydrin reaction. Inhibitory activity assays (Table 1) demonstrated phosphoramidon is a specific inhibitor of thermolysin (ID₅₀ 0.4 μg/ml), with no effect on trypsin, papain, α-chymotrypsin, or pepsin (ID₅₀ >250 μg/ml). This specificity suggests interaction between phosphoramidon's phosphoric acid moiety and the Zn²⁺ in thermolysin's active site. Phosphoramidon (100 μg/ml) exhibited no antibacterial or antifungal activity and low toxicity—intravenous injection of 1.0 g/kg to mice caused no deaths.