Structure of sporostatin (M5032), an inhibitor of cyclic adenosine 3',5'-monophosphate phosphodiesterase.

EnzymeAssay Bovine heart CAMP-PDEactivity was measured as follows: The reaction mixture contained in a final volume of 1.0 ml, Tris-HCl buffer (pH7.0) 40him, MgSO4 2mM, CAMP0.6mMas a substrate and the enzyme 7.5mU/ml. The reaction was started by adding the substrate. Incubation was carried out 30°C for 30 minutes, and the reaction was terminated by adding 0.1ml of 55% perchloric acid, and the mixture was centrifuged at 3000 rpm for 10 minutes. The reaction product, 5'-AMP in the supernatant, was analyzed with HPLC. The HPLC was performed on a reversed phase silica gel column (Hitachi, #3056, 4mm i.d.x150mm) with lOmM