AbstractRecent advances in analytical chemistry have provided an array of techniques to analyze cyanotoxins in a variety of samples. Mass spectral analysis by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) has proven to be a reliable and rapid tool to detect and identify microcystin variants in very small samples such as single Microcystis colonies. Mass signals presumptive of microcystin variants can be characterized further and identified by postsource‐decay fragmentation and comparison of observed fragment spectra with theoretical ones or with those of pure reference compounds. By analyzing Microcystis colonies from water blooms, the high metabolic diversity among individual colonies in the communities could be established. For monitoring microcystins in environmental samples, MALDI‐TOF MS can provide considerable support to HPLC by identifying microcystin variants not available as purified standards, especially in Planktothrix‐dominated blooms in which demethylated variants are predominant. A short description of the principles of MALDI‐TOF MS is provided, followed by examples of its application and a discussion of its potential use in cyanotoxin research. © 2002 Wiley Periodicals, Inc. Environ Toxicol 17: 367–374, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/tox.10073