Identification of Phenylthiohydantoin Derivatives of Amino Acids
The breakthrough which has led to this advancement is the development of an identification and detection system for phenythiohydantoin (PTH) derivatives of amino acids by reverse-phase high-pressure liquid chromatography (RP-HPLC). This chapter describes in detail those methods which employ methanol and acetonitrile as an organic modifier. To minimize the High Performance Liquid Chromatography (HPLC) and column problems, it is strongly recommended that the samples for PTH amino acid identification be analyzed on one type of HPLC if possible. The separation of PTH amino acids by RP-HPLC has been found to be dependent largely on the buffer system, flow rate, and type of the reverse-phase column, and to a lesser extent on the instrumentation. Commercial unavailability of standard dimethylaminobenzene thiohydantoin (DABTH) derivatives of amino acids necessitates that these derivatives by synthesized and purified in the laboratory. DABTH derivatives of arginine and histidine are repeatedly dissolved in water and lyophilized.