An improved method for the measurement of tissue metabolites associated with cellular energetic state by capillary electrophoresis is described. This method allows 17 compounds present in a mixture of standards to be determined simultaneously within 43 min with good reproducibility. ATP, ADP, AMP, UTP, IMP, inosine, hypoxanthine, creatine, phosphocreatine, UDP-galactose, NAD and NADH were detected in samples of either rat heart tissue or rat neonatal cardiomyocytes. This method can detect compounds at concentrations of 5 microm in samples. Recoveries for ATP and phosphocreatine added to cardiomyocyte samples were 99.4 +/- 2.1% and 103.1 +/- 3.3%, respectively (mean +/- SEM, n = 3). Our method has been comprehensively validated and is capable of measuring a wider range of tissue metabolites important in assessing cellular energy status than existing methods.