A fast high-performance liquid chromatographic method for the measurement of plasma concentration and specific...

A fast high-performance liquid chromatographic (HPLC) method for the measurement in plasma of phenylalanine concentration and specific activity is reported. One-to-two mL of acidified plasma are applied to an ion-exchange resin. The eluted amino acids are enzymatically converted into the corresponding alpha-ketoacids, i.e. phenylalanine is converted into phenylpyruvic acid. After a two-step extraction, phenylpyruvic acid is separated by reverse phase chromatography within 8-10 min. The use of an internal standard allows precise quantitation of plasma concentrations. The radioactivity eluted from the HPLC is divided by the amount injected to yield the specific activity. Concentration and rate of appearance of phenylalanine in man, calculated with the L-[2,6-3H]-phenylalanine tracer, are in the range of published data.