Dutch elm disease (DED) is a vascular wilt disease which has decimated populations of American elm (Ulmus americana L.) throughout North America. Mansonones are a family of phytoalexins found within the genus Ulmus and are thought to be involved in the resistance mechanism to DED. There have been no previous studies which have investigated the variation in production of mansonones within a large group of related elm. The purpose of this study was to investigate the production of mansonones within progeny from several parent trees within 6 populations of American elm located in Manitoba, Canada. In this study a method to measure mansonones by high performance liquid chromatography was developed with a shorter run time and a simple solvent system procedure than previously published by other researchers. Mansonone production varied significantly between genotypes. Production of mansonones within progeny from one parent within a population were significantly different than other parents within the same population. No significant differences in mansonone production were found between populations. Throughout the study when seedlings were measured for mansonone production through in vitro callus cultures it was found that mansonones were also produced in control treatments. Further experiments performed to address this problem revealed that mansonone production was initiated approximately at the same time callus tissue was formed. No previous studies have reported this incidence. Vascular staining, a symptom of DED, within progeny varied significantly between parents and populations. When mansonone production and vascular staining were compared among progeny from the same parent trees and populations, no correlation was found (r2 = 0.01). At the juvenile stage, it appears that neither mansonone production nor vascular staining is an adequate indicator of resistance. This is the first study to show large differences in the production of mansonones over such a large study group. Future research on mansonone production and DED resistance must concentrate on more mature material which can show visual symptoms of DED more accurately.