Acute caffeine ingestion increases serum NEFA and plasma adrenaline and decreases insulin sensitivity. Although frequently suggested, it is not known if a tolerance to these alterations in glucose homeostasis is developed in habitual caffeine consumers. Our objective was to determine whether acute caffeine ingestion continued to alter insulin, glucose, NEFA and adrenaline during an oral glucose tolerance test (OGTT) following 14 d of caffeine consumption. Twelve caffeine-naive young males underwent four OGTTs over a 4-week period. Subjects ingested a gelatin-filled placebo (PLA) capsule on the first trial day and 5 mg caffeine/kg body weight on the remaining three trial days (day 0, day 7, day 14) before a 2 h OGTT. Following day 0 and day 7, subjects were given six dosages of 5 mg caffeine/kg to consume per d between trials. Serum insulin and blood glucose area under the curve (AUC) were significantly elevated (P < 0.05) v. PLA on day 0 (36 and 103 %, respectively) and were not different from PLA on day 7. On day 14, insulin AUC was 29 % greater than PLA (P < 0.05), and glucose was greater (P < 0.05) during the first hour, although the 50 % elevation in glucose AUC was not different from PLA. Before the OGTT, caffeine resulted in greater (P < 0.05) serum NEFA and plasma adrenaline concentrations in all three caffeine trials, but both NEFA and adrenaline concentrations were decreased (P < 0.05) on day 14 v. day 0. Although 14 d of caffeine consumption by previously caffeine-naive subjects reduced its impact on glucose homeostasis, carbohydrate metabolism remained disrupted.