Hyaluronidase (HAdase), which depolymerizes hyaluronic acid (HA), is distributed in various animal tissues and secretions. Previously known HAdase-producing microorganisms are limited to pathogenic bacteria such as Streptococcus group A, Staphylococcus aureus, and Clostridium welchii, and both testicular and bacterial HAdase become labile during purification. To establish industrial microbial production of HAdase, it is essential to obtain non-pathogenic microbes that secrete stable enzymes. The authors successfully isolated a Streptomyces strain capable of constitutively producing a very stable HAdase. This study reports the discovery of this new HAdase produced by the Streptomyces strain, along with its partial purification and some properties. Strains of Streptomyces and mold were isolated from soils using modified Czapek agar containing 0.6% HA and 40 r/ml Achromycin, then screened for HAdase productivity. Streptomyces strain No. 81-10 was selected, which accumulated constitutive HAdase in the culture fluid when grown in a soluble starch-peptone medium (3% soluble starch, 0.5% peptone, 0.5% meat extract, 0.8% ammonium sulfate, 0.15% yeast extract, 0.1% dipotassium phosphate, pH 7.2) at 30℃ for 72 hours with shaking. The enzyme was partially purified through ammonium sulfate fractionation (40~70% saturation, pH 5), DEAE cellulose column chromatography, CM cellulose column chromatography, and Sephadex G-100 gel filtration. The enzyme showed no activity towards chondroitin sulfate, with an optimal pH of 5.0 and an optimal temperature of 65~70℃. It was highly thermostable, with no appreciable activity loss after incubation at 75~80℃ for 30 minutes. Vacuum-dried enzyme powder retained full activity for over 4 months at room temperature, and the purified enzyme solution remained active for at least one month when stored in the refrigerator. The enzyme was almost completely inhibited by 10⁻² M p-chloromercuribenzoate and 10⁻³ M mercuric chloride, as well as by 10⁻² M monoiodoacetate, 10⁻⁴ M potassium permanganate, 10⁻³ M iodine, and 10⁻² M N-bromosuccinimide. This is the first report of microbial HAdase from non-pathogenic microbes, and the HAdase from Streptomyces strain No. 81-10 is clearly different from those derived from bacteria and animals.