The effects of prolonged feed deprivation were assessed on blood plasma metabolites and free amino acid levels in Solea senegalensis. Juvenile specimens were maintained at two experimental conditions (24 h fasted and 21 days feed-deprived). In feed-deprived fish, relative growth rate and hepatosomatic index as well as plasma triglycerides and proteins levels were significantly lower. However, plasma cortisol levels were higher in feed-deprived fish, while plasma glucose and lactate values were not significantly different between treatments. Furthermore, feed-deprived fish showed higher levels of total plasma free amino acids than 24 h fasted fish. In 21 days feed-deprived sole, an increase in plasma cortisol levels may suggest a functional role in mobilizing energy due to the catabolic action of this hormone in teleosts. Higher levels of glutamine, arginine and ornithine in 21 days feed-deprived fish may be indicative of a dual role for these amino acids: ammonia detoxification and carbon source for gluconeogenesis. The increased plasma glucogenic and branched-chain amino acid levels, together with the maintenance of plasma glucose in 21 days feed-deprived sole, suggest active liver gluconeogenic processes supported by tissue proteolysis.