DEFEKTE TETRAHIDROBIOPTERIENHOMEOSTASE IN 'N SWART FAMILIE MET HIPERFENIELALANIENEMIE

Abstract

In 1984 the Department of Biochemistry of the Potchefstroom University for Christian Higher Education introduced a metabolic screening programme. Results from a two year research project on defective amino acid metabolism of 1443 clinically selected patients are presented. Utilization of a dual screening approach consisting of one-dimensional thin layer chromatography and automatic amino acid analysis led to the identification of various abnormalities associated with known metabolic defects. Seven different types of amino acidurias were diagnosed in a total of 15 positively tested patients. Hyperphenylalaninemia accounted for 5 of the amino acidurias of which one was a very rare case of malfunctional phenylalanine metabolism. Amongst the total screened group, atypical ninhydrin positive components were found in approximately 13 % of the analyzed urine samples. It was not possible to link the presence of these components to any known defect of the amino acid metabolism. Given that the presence of the atypical ninhydrin positive components complicates the interpretation of laboratory results, it was necessary to investigate their nature and origin. Different separation techniques were applied in an attempt to isolate the metabolites responsible for the atypical ninhydrin colour complexes. However, it was not possible to isolate these metabolites using the normal protocol for isolation of amino acids. This led to the conclusion that their physico-chemical properties did not correspond with that of known amino acids. GC-MS analysis indicated structural similarities between these atypical compound(s) and amino acid conjugates associated with some organic acidurias. Analysis of a consignment of samples obtained from newborns on antibiotic treatment focused the attention on medication as possible cause of the atypical ninhydrin positive components. GC-MS analysis indicated the presence of a metabolite of the penicillin molecule in the urine of patients receiving antibiotics. These results were confirmed by a penicillin loading test on a healthy individual. Hyperphenylalaninemia (HPA) is not prevalent in Negroids. Therefore, the case of a black baby girl with HP A discovered during this study was exceptional. Using a dual screening procedure, elevated levels of phenylalanine were detected in different plasma specimens of the patient. The extent of the elevation was such that it could not be associated with phenylketonuria (PKU), but rather a variant of the disease. An approach investigating the disease on metabolite as well as enzyme level was formulated. A tentative diagnosis was made on the overall status of metabolites in the phenylalanine hydroxylase system, after which the diagnosis was confirmed by enzymatic analysis. Metabolite analyses revealed the presence of phenolic compounds in the patient's urine. A phenylalanine loading test indicated that the clearance rate of the excess phenylalanine from the patient's bloodstream was impaired. However, her phenylalanine levels did not rise as much as commonly expected in the case of PKU. Although this result was indicative of a tetrahydrobiopterin-defect (BH4), the patient did not react positively to the administration of 20 mg BH4/kg mass. The patient's excretion pattern of urinary pterines, as well as her low levels of active BH4 were consistent with a DHPR-defect. The reduced levels of neurotransmitter metabolites in cerebrospinal fluid of the patient also indicated a DHPR-defect. Information obtained from the metabolite analyses indicated DHPR-defect as the possible cause of hyperphenylalaninemia in the patient. This diagnosis was confirmed when no DHPR-activity could be demonstrated in dried blood spots from the patient. Given that the patient did not react to a high dose of 20 mg BH4 per kilogram of weight, the disorder is referred to as non-responsive dihydropteridine reductase deficiency. The patient described here is not only the second case of non- responsive DHPR-defect in the world, but also the first black patient diagnosed with this disease.