HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHODS FOR THE QUANTIFICATION OF TETRAHYDROBIOPTERIN BIOSYNTHETIC-ENZYMES

Tetrahydrobiopterin is a cofactor in hydroxylation reactions, includin g phenylalanine 4-monooxygenase, tyrosine 3-monooxygenase, tryptophan 5-monooxygenase, alkyl glycol ether monooxygenase and nitric oxide syn thase, Determination of its biosynthesis is carried out to diagnose in herited diseases leading to partial defects in tetrahydrobiopterin syn thesis. In addition, tetrahydrobiopterin synthesis is induced by proin flammatory cytokines, and intracellular levels of tetrahydrobiopterin in many cases limit the activity of tetrahydrobiopterin-dependent reac tions, such as nitric oxide synthase in intact cells. Biosynthesis of tetrahydrobiopterin from guanosine 5'-triphosphate (GTP) requires the action of three enzymes, GTP-cyclohydrolase I(E.C. 3.5.4.16), 6-pyruvo yl tetrahydropterin synthase (E.C. 4.6.1.10) and sepiapterin reductase (E.C. 1.1.1.153). Methods for quantification of biopterin and related pteridines in biological matrices by HPLC and application of these fo r determining the activity of the three tetrahydrobiopterin biosynthet ic enzymes are reviewed in this article.