Eight normal controls and nine non-insulin-dependent diabetes mellitus diabetics were, after an overnight fast, infused for 3 h with [6-3H]- and with [U-13C]glucose with six 13C carbons at rates from 0.03 to 0.15 mg.kg-1.min-1. Plasma glucose and lactate were assayed by gas chromatography-mass spectroscopy. Several parameters of glucose metabolism were calculated from the mass isotopomer distribution. Glucose production (GP) determined with [6-3H]- and [U-13C]glucose agreed closely. GP was 1.9 +/- 0.16 (range 1.3-2.5) mg.kg-1.min-1 in controls and 2.8 +/- 0.29 (1.7-4.5) mg.kg-1.min-1 in diabetics (P < 0.05). The correlation in diabetes between plasma glucose and GP (r = 0.911, P < 0.01) was close. Recycling of carbon (8 vs 7%) dilution by unlabeled carbon (2- vs 2.3-fold), and dilution via the tricarboxylic acid cycle (1.5-fold) were similar in controls and diabetics. Gluconeogenesis was 0.90 +/- 0.08 (0.5-1.3) mg.kg-1.min-1 in controls and 1.30 +/- 0.13 (0.8-1.9) mg.kg-1.min-1 in diabetics (P < 0.05). Gluconeogenesis contributions to GP were 46.6 +/- 4.0% (26-61%) in the controls and 48.8 +/- 5.7% (32-83%) in diabetics. We show that, using [U-13C]glucose infusion of 2-5% of glucose turnover (0.03-0.10 mg.kg-1.min-1), a large number of parameters of glucose metabolism may be determined in humans.