HPLC measurement of phenylalanine by direct injection of plasma onto an internal-surface reversed-phase silica support.

Direct-injection analysis for phenylalanine in plasma by isocratic "high-performance" liquid chromatography (HPLC) is described. For chromatographic separation I use an internal-surface reversed-phase silica support. The method involves ultraviolet detection, requires as little as 10 microL of plasma (routinely 20 microL), and supplies results in less than 7.5 min from injection. The intra-batch coefficient of variation for phenylalanine in human plasma samples (197-1104 mumol/L) ranged between 3.0% and 6.0% and the range of linearity was 10-1250 mumol/L for a 20-microL sample injection loop. The upper limit can readily be extended by using a smaller volume loop. Phenylalanine results from direct plasma injection (y) correlated well with those of a protein precipitation/HPLC technique (x): y = 0.931x + 46.4 (r = 0.951, n = 41, range 78-1344 mumol/L).