In this study we investigated in rats whether hydroxycitric acid reduces the postprandial glucose response by affecting gastric emptying or intestinal glucose absorption. We compared the effect of Regulator HCA (310 mg/kg) (HCA) and vehicle (control) on the glucose response after an intragastric (IG) or intraduodenal (ID) glucose load to investigate the role of altered gastric emptying. Steele's one compartment model was used to investigate the effect of HCA on systemic glucose appearance after an ID glucose load, using [U-¹³C]glucose and D-[6,6-²H₂]glucose. Since an effect on post-absorptive glucose clearance could not be excluded, the effect of HCA on the appearance of enterally administered glucose in small intestinal tissue, liver, portal and systemic circulation was determined by [U-¹⁴C]glucose infusion. The data show that HCA treatment delays the intestinal absorption of enterally administered glucose at the level of the small intestinal mucosa in rats. HCA strongly attenuated postprandial blood glucose levels after both IG (p<0.01) and ID (p<0.001) glucose administration, excluding a major effect of HCA on gastric emptying. HCA delayed the systemic appearance of exogenous glucose but did not affect the total fraction of glucose absorbed over the study period of 150 min. HCA treatment decreased concentrations of [U-¹⁴C]glucose in small intestinal tissue at 15 min after [U-¹⁴C]glucose administration (p<0.05), in accordance with the concept that HCA delays the enteral absorption of glucose. These data support a possible role for HCA as food supplement in lowering postprandial glucose profiles.