The V8 endoprotease from Staphylococcus aureus hydrolyzes the cyclic lipopeptide, surfactin, at 100 mu M. The fragmentation lies between the two residues L-Glu1 and L-Leu2 leading to an open-chain lipopeptide. The structure of the product arising from the enzymic hydrolysis was established by chemical methods and mass spectrometry. The enzyme action has been suggested to be likely limited by the aggregation state of the lipopeptide in solution.