Abstract: Taking the fruiting bodies of Isaria cicadae as materials, ultrasonic-microwave synergistic extraction method was used to extract adenosine in I. cicadae mycelium and the extraction conditions were optimized. According to the single factor and orthogonal experiments, the optimal extraction conditions were 150 W of microwave power, 120 s of ultrasonic microwave synergistic extraction time, 1 ∶ 30 of solid-liquid ratio and 20% of methanol volume fraction. The content of adenosine in mycelium of I.cicadae determined was ( 0.53 ± 0.02) mg/g.The adenosine contents in different areas were determined by high performance liquid chromatography.The results showed that the adenosine contents of I. cicadae provided from four areas were ranged from 0.14 to 0.40 mg/g. The content of mycelium adenosine in the strain JJS0017 isolated from Jinggangshan was highest, which was followed by the strain JJS0007 obtained from Lianhua town, Jiujiang city.The lowest content was 0.14 mg/g, which was from the strain JJS0032 from Rao town, Poyang county, Jiangxi province.This article could provide a theoretical basis for the development and utilization of I.cicadae.