An LC-ESI-MS (negative ions) method was developed for the determination of cyclopiazonic acid (CPA), a mycotoxin produced by many Aspergillus and Penicillium genuses, in milk samples. First, the acquisition parameters of the ESI mass spectrometer were optimised for deprotonated CPA and MS-MS measurements were performed, giving a fragmentation pattern. After this stage, LC separation was applied to milk extracts (with or without CPA spikes) by means of an aminic column, using the selective reaction monitoring (SRM) acquisition mode for MS detection. The CPA response was linear over three decades of concentration and an LOD of 5 ng mL(-1) was estimated; moreover, the extraction procedure produced almost quantitative recoveries of CPA from milk. Twenty different milk samples were analysed and three of them were found to be contaminated with CPA to various extents.