The influence of alanine on plasma amino acid concentrations and fuel substrates as well as cycling performance was examined. Four solutions [6% alanine (ALA); 6% sucrose (CHO); 6% alanine and 6% sucrose (ALA-CHO); an artificially sweetened placebo (PLC)] were tested using a double-blind, randomised, cross-over design. During each trial, ten cyclists ingested 500 mL of test solution 30 min before exercise and 250 mL after 15, 30, and 45 min of exercise. Participants cycled for 45 min at 75% VO(2)max followed by a 15-min performance trial. Blood was collected before beverage consumption and prior to the performance trial. Alanine concentration was increased (p < 0.05) by approximately tenfold for ALA and ALA-CHO and less than twofold for CHO and PLC. Alanine ingestion increased concentrations of most gluconeogenic amino acids. Overall, alanine supplementation tended to produce favourable metabolic effects, but did not influence performance.