State-of-the-art of high-performance liquid chromatographic analysis of amino acids in physiological samples

Abstract

Nowadays, the fast and sensitive high-performance liquid chromatographic (HPLC) analysis of derivatized amino acids is a good alternative to the ion-exchange chromatography (IEC) method for plasma amino acids. However, several precautions have to be taken in order to obtain reliable data on the concentration of amino acids in physiological fluids. These include the collection, centrifugation, storage conditions and the method of deproteinization. Futhermore, the method of pre-column derivatization in connection with the protein precipitant used the choice of the chromatographic system which determines the overall resolution are important factors. HPLC methods using pre-column derivatization with o-phthaldialdehyde were suitable for the accurate determination of many primary amino acids in plasma because of their high sensitivity, simplicity, speed and reliability. When the determination of secondary amino acids and cystine was also necessary, the phenylisothiocyanate method was the preferred technique. The intra-laboratory variability of the HPLC method was satisfactory while the inter-laboratory variation of this method was found to be similar to that of IEC. HPLC methods capable of separating over forty physiological amino acids seem promising for the analysis of urine samples.