The measurement of N-acetylaspartic acid (NAA), N-acetylglutamic acid (NAG), and N-acetylaspartyl-glutamic acid (NAAG) in the whole brain of 3-mercaptopropionic acid (3-MPA)-treated rats has been developed using Liquid chromatography-mass spectrometry with an atmospheric pressure ionization interface system. The recoveries of these compounds were 90.85 +/- 3.43% for NAA, 91.62 +/- 5.47% for NAG, and 92.29 +/- 4.44% far NAAG. The detection Limits for NAA, NAG, and NAAG were 12, 15, and 20 mu g/ml, respectively. After administration of 3-MPA, the concentrations of NAA NAG, and NAAG in the whole brain over 10 min increased 177.25, 134.23, and 127.70%, respectively. These concentrations then decreased over the next 60 min. The simultaneous determination of NAA, NAG, and NAAG using this method was found to be very useful for studies of metabolism of NAA, NAG, and NAAG in biological samples. (C) 1999 Academic Press.