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Since 1990, 20 diagnostically confirmed phenylketonuria (PKU) patients have been screened with a tetrahydrobiopterin (BH4) loading test, in which plasma phenylalanine and urinary pterin metabolites were investigated, ind activity of dihydropteridine reductase (DHPR) was determined as well. The results showed that there was no statistical difference between the concentrations of plasma phenylalanine before and after BH4 (20mg/kg) administration in all patients, and values of urinary neopterin and biopterin were within the range of classic PKU. All patients but one had normal activity of DHPR in red cells. This suggests that incidence of BH4 deficiency in PKU patients amounts to five percent (1/20) which is almost the same as reported abroad.

We describe the clinical, neurologic, and biochemical findings in 10 patients with 6-pyruvoyl tetrahydropterin synthase (6-PTS) deficiency from seven families, all of whom originate from one large tribe in Saudi Arabia. This deficiency presents with severe, early onset of failure to thrive, neurologic deterioration, and morbidity and mortality secondary to repeated episodes of bronchopneumonia or cardiorespiratory abnormalities. The urinary pterin excretion pattern indicates deficient activity of 6-PTS, which has been confirmed by direct enzyme assay in red blood cells of three patients. We treated our patients with combined use of tetrahydrobiopterin 20 mg/kg/d, L-dihydroxyphenylalanine 15 mg/kg/d, carbidopa 3.75 mg/kg/d, and L-5-hydroxytryptophan 5 mg/kg/d. Neurologic findings improved significantly in all after 5 to 24 months. Although head circumference and weight returned to the lower limit of normal in four, height normalized only in one of seven patients. Despite an unrestricted diet during combined therapy, blood phenylalanine and urinary excretion of neopterin and biopterin returned to normal.

New catalytic functions of human carbonyl- and aldose reductase in tetrahydrobiopterin biosynthesis are proposed. 6-Pyruvoyl tetrahydropterin, an intermediate in the biosynthesis of tetrahydrobiopterin, was converted to 6-lactoyl tetrahydropterin and 1'-hydroxy-2'-oxopropyl tetrahydropterin by carbonyl reductase under anaerobic condition. 1'-Hydroxy-2'-oxopropyl tetrahydropterin was subsequently metabolized to tetrahydrobiopterin by aldose reductase. Based on these results alternative pathways for the synthesis of tetrahydrobiopterin in patients with genetic defects of sepiapterin reductase are suggested.

Refsum's disease (Heredopathia atactica polyneuritiformis) is caused by accumulation of phytanic acid in all body tissues due to an inherited failure of alpha-oxidation of branched chain fatty acids. Plasmapheresis has been reported to be beneficial by removal of phytanic acid from the blood. We describe a patient with Refsum's disease in whom long-term plasmapheresis did not improve clinical, biochemical or electrophysiological parameters.

Three novel pteridines have been isolated from the urine of patients with a new variant of 6-(L-erythro-1',2'-dihydroxypropyl)-5,6,7,8-tetrahydropterin (tetrahydrobiopterin) deficiency, showing hyperphenylalaninemia. From the results of high performance liquid chromatography, oxidative degradation, and gas chromatography-electron impact mass spectrometry, their structures were identified as 7-(D-erythro-1',2',3'-trihydroxypropyl)-pterin (7-neopterin), 7-(L-erythro-1',2'-dihydroxypropyl)-pterin (7-biopterin), and 6-oxo-7-(L-erythro-1',2'-dihydroxypropyl)-pterin (6-oxo-7-biopterin). The ratio of biopterin to 7-biopterin in the patients' urines was 1:1, and after oral loading with tetrahydrobiopterin, 7-biopterin excretion rose parallel to biopterin. This finding suggests that 7-substituted pterins may be formed endogenously by a yet unknown isomerization reaction. The cause of hyperphenylalaninemia is still unclear. The activities of the enzymes involved in tetrahydrobiopterin biosynthesis and regeneration were found to be normal in the patients, and no effect of 7-biopterin on these enzymes was observed in vitro. However, compared with the normal cofactor, tetrahydrobiopterin, the Km values of tetrahydro-7-biopterin for phenylalanine hydroxylase and dihydropteridine reductase are 20 and 5 times higher, respectively.

An assay is described for GTP cyclohydrolase I activity in human mononuclear cells isolated from 20 ml of heparinized blood. The activity of this enzyme was low in unstimulated cells and increased 5-10 times after stimulation by phytohemagglutinin (formation of 0.8-1.3 pmol dihydroneopterin triphosphate/min per mg protein at 37 degrees C, n = 15) or mixed lymphocyte culture. No activity was detected in phytohemagglutinin-stimulated mononuclear cells of a patient with proven GTP cyclohydrolase I deficiency in liver; the samples from the father and mother of the patient showed 30 and 46%, respectively, of the mean of 15 healthy controls. In unstimulated cells, neopterin was the main component of the total intracellular pterins (after oxidation). After stimulation, dihydroneopterin triphosphate, measured as neopterin triphosphate by high performance liquid chromatography, was increased 10-30 times; neopterin and pterin were increased only 2- to 6-fold. Since the immunoreactive cells from this patient were unable to produce pterins and all immunological tests on the patient were normal, it is concluded that neither dihydroneopterin triphosphate, nor one of its metabolites are of primary importance for an immune reaction. The assay described can be used for the detection of heterozygotes of GTP cyclohydrolase I deficiency.

A 44-year-old man sustained a transmural inferolateral myocardial infarction and began to show signs of postmyocardial infarction syndrome (Dressler's syndrome) one week after infarction. Anticoagulant therapy had been initiated for suspected pulmonary thromboembolism. Administration of steroids did not improve the patient's clinical condition or the results of laboratory investigations. A massive pericardial effusion was diagnosed clinically, and this diagnosis was confirmed by a pericardial scan using 99m technetium. Subsequently, 1,800 ml of bloody fluid was removed from the pericardial cavity, and following the pericardiocentesis, the patient became asymptomtic. This case reemphasizes the hazards of anticoagulant therapy in patients with the postmyocardial infarction syndrome.

A 23-yr-old male student presented with clinical and biochemical evidence of Cushing's syndrome. One month later, his elevated plasma and urinary adrenal steroids had returned to normal. At surgery, an adrenal adenoma was removed from his right side. We postulate that he either underwent a temporary spontaneous remission of his disease without treatment, prior to surgery, or that his adenoma secreted glucocorticoids in a cyclical fashion.

GTP cyclohydrolase I from human liver and Escherichia coli is competitively inhibited by 8-aminoguanosine triphosphate with a dissociation constant (Ki) of 0.25 mumol/l. 8-Aminoguanosine triphosphate, prepared from GTP and hydroxylamine-O-sulfonic acid, was coupled to Sepharose 4B and used as an affinity adsorbent for a 309-fold purification of GTP cyclohydrolase I from human liver. GTP cyclohydrolase I from human liver is a relatively heat-stable enzyme with a half-life of 2 min at 80 degrees C, an isoelectric point (pI) of about 5.6, and a Km for GTP of 31 mumol/l. Addition of KCl (0.3 mol/l) increased the Km to 153 mumol/l. No cofactors were required for activity. L-erythro-5,6,7,8-Tetrahydrobiopterin, L-erythro-7,8-dihydrobiopterin, L-sepiapterin and 8-aminoguanosine triphosphate were strong inhibitors.

An increase in total urinary neopterin was observed in 12 of 13 patients with acquired immunodeficiency syndrome (AIDS), seven of 13 patients with lymphadenopathy, one of six healthy homosexual males, seven of ten adult patients with staphylococcal pneumonia, 11 of 12 children with viral infections, four of seven children with bacterial infections, and 12 of 13 children with various immune defects. Extremely high values of total urinary neopterin and monapterin were observed in severely ill patients with AIDS and those with familial hemophagocytic lymphohistiocytosis. Neopterin excretion was normal in two AIDS patients with Kaposi's sarcoma, but without opportunistic infections at that time. On reexamination of one of these patients later on, elevated neopterin values were noted. Parallel increases in neopterin and monapterin were found, whereas biopterin was usually normal. The increase in total neopterin was mainly due to 7,8-dihydroneopterin and was accompanied by an increase in 3'-hydroxysepiapterin. Increased neopterin in urine is assumed to reflect the increase in GTP pool and GTP cyclohydrolase I activity as observed in stimulated monocytes. Thus, neopterin, as a measure of the activation of the nonspecific cellular immune system, may be used diagnostically to detect allograft rejection after transplantations and to follow-up HTLV-III positive patients.

Three unknown compounds present in the urine of a patient with mild hyperphenylalaninemia were identified to be L-erythro-7-iso-biopterin, D-erythro-7-iso-neopterin, and L-erythro-6-oxo-7-iso-biopterin. The newly identified pterins were named primapterin, anapterin, and 6-oxo-primapterin, respectively. Primapterin and anapterin are present in very low concentrations in every human urine, as well as in the liver of man and mouse, whereas 6-oxo-primapterin was detected in the patient's urine only. Substantial amounts of primapterin were excreted in the patient described. The metabolic origin of primapterin and anapterin is still obscure.

In our laboratory study 19 students effected 4 repetitive tasks of 70 min each. All four conditions had a motor task at the base and in addition two had to deal with discrimination. In the 2 tasks with discrimination, the electromyography of the neck muscles was distinctly higher than in the 2 other conditions. The tasks with discrimination were accompanied by a higher subjective tension, a lower heart rate variability and a slightly increased urinary excretion. The main cause for an augmentation in the muscle tonus might be the immobilisation of the neck.

A 4-year-old patient is described with hyperphenylalaninemia, severe retardation in development, severe muscular hypotonia of the trunk and hypertonia of the extremities, convulsions, and frequent episodes of hyperthermia without infections. Urinary excretion of neopterin, biopterin, pterin, isoxanthopterin, dopamine, and serotonin was very low, although the relative proportions of pterins were normal. In lumbar cerebrospinal fluid, homovanillic acid, 5-hydroxyindoleacetic acid, neopterin and biopterin were low. Oral administration of L-erythro tetrahydrobiopterin normalized the elevated serum phenylalanine within 4 h, serum tyrosine was increased briefly and serum alanine and glutamic acid for a longer time. Urinary dopamine and serotonin excretion were also increased. Administration of an equivalent dose of D-erythro tetrahydroneopterin was ineffective and demonstrated that this compound is not a cofactor in vivo and cannot be transformed into an active cofactor. GTP cyclohydrolase I activity was not detectable in liver biopsies from the patient. The presence of an endogenous inhibitor in the patient's liver was excluded. This is the first case of a new variant of hyperphenylalaninemia in which the formation of dihydroneopterin triphosphate and its pterin metabolites in liver is markedly diminished. Normal activities of xanthine oxidase and sulfite oxidase were apparent since uric acid levels were normal and no increase in hypoxanthine, xanthine, and S-sulfocysteine concentrations could be observed in urine. It is concluded that the molybdenum cofactor of these enzymes may not be derived from dihydroneopterin triphosphate in man. Also, since no gross abnormalities in the patient's immune system could be found, it seems unlikely that dihydroneopterin triphosphate metabolites, such as neopterin, participate actively in immunological processes, as postulated by others. See Note added in proof.

The occurrence, properties and functions of GTP-cyclohydrolases in mammalian and non-mammalian systems is reviewed. GTP-cyclohydrolases catalyse the removal of C-8 atom from GTP as formic acid. GTP-cyclohydrolase I (EC 3.5.4.16) converts GTP into D-erythro-7, 8-dihydroneopterin triphosphate, whereas GTP-cyclohydrolase II forms 2,5-diamino-4-oxo-6-(5'-phosphoribosyl) -amino pyrimidine, a possible intermediate in the biosynthesis of riboflavin. GTP-cyclohydrolase I is the first enzyme in the biosynthesis of tetrahydrobiopterin, a cofactor of the monooxygenases of the aromatic amino acids. It is the rate limiting enzyme in many mammals, but not in man. Recently, patients with GTP -cyclohydrolase I deficiency were described, a variant form of tetrahydrobiopterin-deficient hyperphenylalaninaemia.

Urines of two children with 17 alpha-hydroxylase deficiency contained a number of 5-pregnane- and pregnenediols, -triols and -tetrols with a hydroxy or oxo group in position 11 of the steroid ring. They are formed mainly from progesterone via 11-hydroxyprogesterone, pregnanolone and corticosterone, respectively, or from pregnenolone. Three metabolites not previously described, 16-hydroxypregnenolone, 6,21-dihydroxypregnanediol and 6-hydroxytetrahydrocorticosterone, were identified.

D-erythro-7,8-Dihydroneopterin triphosphate (NH2TP) formed from guanosine triphosphate (GTP) by GTP cyclohydrolase I (EC 3.5.4.16) in the presence of EDTA was oxidized to neopterin triphosphate (NTP) by iodine, separated from substrate and other compounds by ion-paired reverse-phase HPLC, and quantitated by fluorometric detection at 365/446 nm. Excess GTP at the end of reaction was controlled by simultaneous detection of guanine nucleotides at 254 nm. The method required only 15 mg of liver tissue for the measurement of GTP cyclohydrolase I and is suitable for activity measurement in liver biopsies. The detection limit was 4 pmol of NTP at a signal to noise ratio of 10:1. The activity of GTP-cyclohydrolase I in homogenates of human liver (n = 10) was 45 pmol NH2TP (range 32-60) formed per milligram protein per hour at 37 degrees C. Liver homogenates from Wistar rats (n = 8) formed 47 pmol NH2TP (range 35-61) per milligram protein per hour.

A 60-year-old man presented with complaints on angina pectoris and was found to have a coronary artery fistula between his left main truck and main pulmonary artery. Particles of 99m Tc-Albumin were injected in the ostium of the left coronary artery, and differential radioactive counts were injected in the ostium of the left coronary artery, and differential radioactive counts were obtained over both lung fields and myocardium. The degree of left to right shunt was calculated at 56% of total left coronary artery flow. The patient underwent ligation of the fistula without any complications. This case report represents a new application of myocardial radioisotopic scanning.

A semiautomated fluorometric method for the quantitative determination of urinary dopa, 3-(3,4-dihydroxyphenyl)-L-alanine, is described. It provides a simple, sensitive and reproducible analytical technique for routine use. Dopa is isolated from interfering substances, especially catecholamines, by adsorption onto aluminium oxide, elution with 0.1 M HCl, then passed through a cation-exchange column. By mild oxidation with potassium ferricyanide, dopa is cyclized to dopachrome. This is isomerised with strong alkali to 5,6-substituted indole, which is highly fluorescent but very unstable in the presence of oxygen. The addition of ascorbic acid and 3-mercaptopropionic acid to the alkaline solution stabilizes the fluorescence. Concentrations of chemicals, pH and reaction times are made optimal for maximal sensitivity. Recovery of dopa added to the urine samples averaged 79% (range, 75 to 83). Reproducibility of results from the same urine specimen gave a coefficient of variation of 2.4%. In healthy adults, we found daily excretion ranges from 17.3 to 54.9 (mean: 36.1) microng/24 h or from 0.015 to 0.048 (mean: 0.021) microng/mg cretinine.