Establishment of reference range of plasma amino acids for younger Chinese children by reverse phase HPLC

Due to lack of country-specific norms in China, we established the reference range of plasma amino acids for younger Chinese children by reverse phase high-performance liquid chromatography (HPLC).

Plasma proteins were precipitated with ethanol. L-Norvaline served as an internal standard. This HPLC method was based on automated precolumn derivatization using o-phthalaldehyde 3-mercaptopropionic acid for primary amino acids and 9-fluorenylmethyl chloroformate for secondary amino acids. Twenty-three amino acid derivatives were separated by a Zorbax Eclipse AAA column and detected fluorometrically. Plasma amino acids were measured in 108 healthy Chinese children (ages 0-5 years, 59 boys and 49 girls).

The assay was linear from 7.2 to 925.0 micromol/L for all amino acids. Recovery of amino acids added to plasma samples was 93%-107%. Within- and between-run reproducibility was 0.18%-6.27% and 2.94%-16.15%, respectively. Sex- and age-specific plasma amino acid reference range for younger Chinese children was established. In our study, the boys had significantly higher levels of glutamine, citrulline, and tryptophan than girls (p < 0.05), and the girls had a significantly higher level of alanine than boys (p < 0.05). Compared with the 0- to 1-year group, the 1- to 5-year group had significantly higher levels of citrulline, valine, phenylalanine, isoleucine, and sarcosine and lower levels of aspartate, glutamate, serine, threonine, alanine, methionine, and tryptophan (p < 0.05).

This study validates the HPLC method described here as a simple, rapid, and reliable assay. The reference range of plasma amino acids for younger Chinese children is different from that for Caucasian children and will facilitate our clinical diagnosis in the future.