Technical Abstract: A new GC-MS method was developed for the analysis of ascaulitoxin, its aglycone, and 4-amino-D-proline, which are phytotoxins with potential herbicidal activity produced by Ascochyta caulina. The method involved directly treating the lyophilized culture filtrate with a derivatizing reagent, converting the mixture of toxins in the filtrate to their corresponding trimethylsilyl derivatives, and consequent analysis by EI-MS. The method is rapid, sensitive and highly specific for the identification and analysis of the toxins in a complex sample matrix. Analysis of culture filtrates using this method suggested that phytotoxicity correlates with the level of ascaulitoxin in the culture filtrate. A new method for the purification of 2,4,7-triamino-5-hydroxyoctandioic acid, the aglycone of ascaulitoxin, is also described.