Phenylketonuria Testing

PHENYLKETONURIA TESTING

Phenylketonuria (PKU) is an inborn error of metabolism in which the body is unable to convert surplus phenylalanine (PA) to tyrosine for use in the biosynthesis of, for example, thyroxine, adrenaline and noradrenaline. This results from a deficiency in the liver enzyme phenylalanine 4-mono-oxygenase (phenylalanine hydroxylase). A secondary metabolic pathway comes into play in which there is a transamination reaction between PA and αketoglutaric acid to produce phenylpyruvic acid (PPVA), a ketone and glutamic acid. Overall, PKU may be defined as a genetic defect in PA metabolism such that there are elevated levels of both PA and PPVA in blood and excessive excretion of PPVA (Figure 26.7).

Control of PKU can be achieved simply by resorting to a low PA-containing diet. However, failure to diagnose PKU will result in mental deficiency, and thus early diagnosis is essential. In 1968, the UK Medical Research Council Working Party on PKU recommended the adoption of the Guthrie test as a convenient method for screening newborn infants. This assay employs Bacillus subtilis as the test organism. In minimal culture medium, growth of this bacterium is inhibited by β-2-thienylalanine (Figure 26.8a) and is competitively reversed in the presence of PA (Figure 26.8b) or PPVA. The use of filter-paper discs impregnated with blood or urine permits the detection of elevated levels of PA and PPVA. The test can be quantified by the measurement of the diameter of the growth zone around the filter-paper disc and comparing it with a calibration curve constructed from known concentrations of PA or PPVA (Figure 26.8c). If positive, the Guthrie test provides presumptive evidence for the presence of PKU. This is then confirmed by other chemical means. The test is still in widespread use but is gradually being phased out in favour of modern genetic testing techniques.