Thiamine requirements for humans are generally expressed as absolute values per day (mg/d) or in relation to total caloric intake. Limited data are available on the relation between thiamine requirements and the intake of carbohydrates. This study was performed to investigate the influence of stepwise increases of carbohydrate intake on the status of thiamine in healthy volunteers under isocaloric conditions. During an adaptation phase of four days, the carbohydrate intake of twelve healthy volunteers (6 male, 6 female) was 55% of total energy intake. During the subsequent intervention periods, carbohydrate intake was increased to 65% of total energy for four days and to 75% for another four days. Thiamine intake, total energy intake, and physical activity were kept constant throughout the study. HPLC analysis was used to measure thiamine in plasma, urine and feces. Erythrocyte transketolase activity (ETK) was determined enzymatically. During the intervention periods thiamine decreased significantly (p < 0.05) in plasma (from 19.3 +/- 3.3 to 16.4 +/- 4.0 nmol/l) as well as in urine (from 72 +/- 56 to 58 +/- 21 mumol/mol creatinine). ETK and feces content of thiamine remained unchanged. An increase of dietary carbohydrate intake from 55% to 65% and 75%, respectively, of total caloric intake for four days per period at isocaloric conditions causes a decrease of plasma and urine levels of thiamine without affecting enzyme activities.