Determination of histidine and 3-methylhistidine in physiological fluids by high-performance liquid chromatography.

High-performance liquid chromatography based on pre-column derivatization of histidine and 3-methylhistidine with o-phthalaldehyde-mercaptoethanol has been used to determine these amino acids in small volumes of plasma and urine. The elution is performed in 45 min on a 5-micron Resolve C18 column by a multi-step gradient. The eluted analytes are measured with a fluorescence detector which provides detection limits of less than 1 pmol per 20-microliter injection. The correlation between concentration and the integrated area of amino acids gives a linear relationship between 10 and 150 pmol per injection. Some preliminary results from patients with chronic renal failure under variation of diet are presented.