Analysis of plasma free amino acid levels is important for diagnosis of inborn errors of metabolism. Traditionally, this is performed using commercially available dedicated amino acid analyzers, but few such instruments are available in Thailand, and many are not used in routine operations. Here, the authors describe the analysis of plasma free amino acid levels in 57 normal children by reverse-phase HPLC and pre-column derivatization with phenylisothiocyanate. Plasma free amino levels are reported as mean +/- SD and 95 per cent confidence interval of mean for each of 5 age groups: 0-6 months; 6-12 months; 1-3 years; 3-6 years; 6-12 years. Mean amino acid levels were generally similar in all age groups (p > or = 0.01), except that hydroxyproline tended to be higher in the 0-6 months age group compared to other age groups (p<0.01). Comparisons were made between the present data with the normal free plasma amino acid levels in children of similar age groups reported both in Thailand and overseas in terms of both mean +/- SD and maximum and minimum values. Overall, our methodology involving HPLC can identify 35 amino acid derivatives, including all the major amino acids except for cysteine, which is substantially more than the number reported in earlier work on plasma free amino acid levels in normal Thai children. Moreover, the present methodology gives mean +/- SD values similar to an overseas report. For these reasons, HPLC should be considered as an alternative approach in laboratories, where demand does not justify the need for dedicated amino acid analyzers. However, there can be substantial variations between the results from different laboratories, and each laboratory should establish its own normal values.