Muscle carnosine is an intracellular buffer. Intake of β-alanine combined with carbohydrate and protein enhanced carnosine loading in human muscle. The aim of the present study was to examine whether muscle carnosine loading was enhanced by β-alanine intake and co-ingestion of glucose in male rats. Thirty six male rats were divided into three groups and supplemented for 4 weeks: β-alanine (βA, 1.8% β-alanine in drinking water), β-alanine and glucose (βAGL, 1.8% β-alanine and 5% glucose in drinking water) and control (C, drinking water). During the supplementation period, rats were exercised (20 m·min-1, 10 min·day-1, 4 days·week-1 for 4 weeks). Muscle carnosine concentration was quantified in soleus (n=12) and rectus femoris (n=6) muscles using HPLC. In soleus muscle, carnosine concentration were 2.24±1.10, 6.12±1.08 and 6.93±2.56 mmol/kg dw for C, βA and βAGL. In rectus femoris, carnosine concentrations were 2.26±1.31, 7.90±1.66 and 8.59±2.33 mmol/kg dw for C, βA and βAGL, respectively. In each muscle, βA and βAGL resulted in similar carnosine increases compared to C. In conclusion, 4 weeks β-alanine intake, either alone or with glucose co-ingestion, equally increased muscle carnosine content. It appears that the potential insulin response to fluid glucose intake does not have an effect on muscle carnosine loading in male rats.