The responses of plants to abiotic stress involve the up-regulation of numerous metabolic pathways, including several major routes that engage thiamine diphosphate (TDP)-dependent enzymes. This suggests that the metabolism of thiamine (vitamin B1) and its phosphate esters in plants may be modulated under various stress conditions. In the present study, Zea mays seedlings were used as a model system to analyse for any relation between the plant response to abiotic stress and the properties of thiamine biosynthesis and activation. Conditions of drought, high salt, and oxidative stress were induced by polyethylene glycol, sodium chloride, and hydrogen peroxide, respectively. The expected increases in the abscisic acid levels and in the activities of antioxidant enzymes including catalase, ascorbate peroxidase, and glutathione reductase were found under each stress condition. The total thiamine compound content in the maize seedling leaves increased under each stress condition applied, with the strongest effects on these levels observed under the oxidative stress treatment. This increase was also found to be associated with changes in the relative distribution of free thiamine, thiamine monophosphate (TMP), and TDP. Surprisingly, the activity of the thiamine synthesizing enzyme, TMP synthase, responded poorly to abiotic stress, in contrast to the significant enhancement found for the activities of the TDP synthesizing enzyme, thiamine pyrophosphokinase, and a number of the TDP/TMP phosphatases. Finally, a moderate increase in the activity of transketolase, one of the major TDP-dependent enzymes, was detectable under conditions of salt and oxidative stress. These findings suggest a role of thiamine metabolism in the plant response to environmental stress.