Glutamate is linked to the glycolytic process, particularly when co-ingested with carbohydrate, but its effects on glucose metabolism are poorly characterised. The present study aimed to (1) specifically examine the effects of carbohydrate administration on circulating glutamate concentrations and (2) investigate the effect of increased glutamate availability, independent of carbohydrate ingestion, on glucose metabolism. A total of nine participants underwent four trials: (1) glutamate supplement+carbohydrate drink (GLU+CHO); (2) glutamate supplement+placebo drink (GLU); (3) placebo supplement+carbohydrate drink (CHO); (4) placebo supplement+placebo drink (CON). Following a fasting blood sample, participants ingested monosodium l-glutamate (MSG; 150 mg/kg body weight) or placebo capsules at each trial followed by a 75 g carbohydrate or a non-energy placebo drink 30 min later. Blood samples were taken at 0, 10, 20, 30, 40, 50, 60, 75, 90, 105 and 120 min. Plasma glutamate concentrations were significantly elevated relative to baseline during the GLU (approximately 10-fold) and GLU+CHO trials (approximately 6-fold). The glucose response to a carbohydrate load was blunted when glutamate was increased in the circulation (peak serum glucose: 5·50 (se 0·54) mmol/l during the GLU+CHO trial v. 7·69 (se 0·53) mmol/l during the CHO trial, P< 0·05). On average, c-peptide results revealed that insulin secretion did not differ between the GLU+CHO and CHO trials; however, four participants demonstrated increased insulin secretion during the GLU+CHO trial and five participants demonstrated decreased insulin secretion under the same conditions. In conclusion, when administration is staggered, MSG and carbohydrate supplementation can be used to manipulate plasma glutamate; however, future studies should control for this dichotomous insulin response.