The EZ: Faast family of amino acid analysis kits: Application of the GC-FID kit for rapid determination of plasma tryptophan and other amino acids

Abstract

Plasma tryptophan (Trp) and other amino acids (AA) can be determined rapidly by gas (GC) or liquid (LC) chromatography using the Phenomenex EZ:Faast ™ family of kits. Three kits are available: (1) GC-FID or -NPD, (2) GC-MS, (3) LC-MS. The two GC kits can determine 32 AA, whereas the LC-MS can determine five additional AA. All three kits, however, share the same experimental procedure up to and including the preparation of derivatised AA. The method is based on solid-phase extraction (SPE), thus saving time on prior removal of plasma or other proteins and interfering substances, and can be applied to other body fluids and experimental media and to supernatants of extracts of solid material. Briefly, SPE is performed using a proprietary cation-exchange mechanism. The acid solution of the internal standard ensures that the free amino acids are in an anionic form suitable for cationic binding. The alkaline nature of the elution medium ensures that the AA cations are released prior to derivatisation. The latter involves production of chloroformate derivatives of both the amino and carboxylic acid groups. With experience, six plasma samples can be so processed within 12 min. The shortest analytical run is <7 min per sample using the GC-FID/NPD kit. Despite its many steps, the procedure becomes second nature and an enjoyable task. I have now used the GC-FID kit with manual injection to process >1,600 plasma and other samples. Limit of detection of AA is 1 μM or less. The procedure has been validated and optimised for Trp and its main five brain uptake competitors.