Four steroidal alkaloids extracted from the roots and rhizomes of Veratrum californicum were separated by high performance liquid chromatography (HPLC) and identified using high resolution electrospray ionization time of flight tandem mass spectrometry (ESI-TOF-MS/MS) as veratrosine, cycloposine, veratramine, and cyclopamine. This paper compares ethanol and benzene as extraction solvents, HPLC solvent conditions leading to good separation of steroidal alkaloids, and MS/MS fragmentation patterns for the four steroidal alkaloids which have been released to the public database MassBank.jp. The reported Soxhlet extraction method nearly triples the recovery of steroidal alkaloids from V. californicum.