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Background: This trial evaluated the maximum tolerated dose (MTD), safety, pharmacokinetics, and activity of volasertib, a selective Polo-like kinase 1 inhibitor that induces mitotic arrest and apoptosis, combined with cisplatin or carboplatin in patients with advanced/metastatic solid tumors (NCT00969761; 1230.6)., Methods: Sequential patient cohorts (3 + 3 dose-escalation design) received a single infusion of volasertib (100-350 mg) with cisplatin (60-100 mg/m(2)) or carboplatin (area under the concentration versus time curve [AUC]4-AUC6) on day 1 every 3 weeks for up to six cycles. Sixty-one patients received volasertib/cisplatin (n = 30) or volasertib/carboplatin (n = 31) for a median of 3.5 (range, 1-6) and 2.0 (range, 1-6) treatment cycles, respectively., Results: The most common cycle 1 dose-limiting toxicities (DLTs) were thrombocytopenia, neutropenia and fatigue. MTDs (based on cycle 1 DLTs) were determined to be volasertib 300 mg plus cisplatin 100 mg/m(2) and volasertib 300 mg plus carboplatin AUC6. Co-administration did not affect the pharmacokinetics of each drug. Partial responses were observed in two patients in each arm. Stable disease was achieved in 11 and six patients treated with volasertib/cisplatin and volasertib/carboplatin, respectively., Conclusions: Volasertib plus cisplatin or carboplatin at full single-agent doses was generally manageable and demonstrated activity in heavily pretreated patients with advanced solid tumors.

Pteridinic derivatives are important biomolecules considered as biomarkers for several diseases, especially in cancer and infectious pathologies. A new fluorimetric-HPLC method for the analysis of nine pteridines in human serum has been reported. Two analytical columns composed by C18 porous and fused core particles were assayed and the results compared. Fused core particle column allows us adequate separation, in only one run and in 15 min. Acid precipitation step of the proteins and clean-up process with an Isolute ENV+ (hydroxylated polystyrene-divinylbenzene copolymer) cartridge of the serum samples have been optimized. Analytes were determined by fluorimetric detection, exciting at 272 nm and measuring the fluorescence emission at 410 nm for isoxanthopterin, at 465 nm for xanthopterin, and at 445 nm for the analysis of the other pteridines. Detection limits between 0.07 and 0.61 ng mL(-1) were calculated according to Clayton criterium. Intraday precision varied from 1.2 to 5.3 and interday precision between 1.2 and 7.4, both expressed as RSD (%). External standard and standard addition calibrations were compared in the analysis of serum samples. The pteridine amounts in serum (expressed as ng mL(-1) ± confidence interval) were 3.69 ± 1.78; 1.35 ± 0.24; 0.46 ± 0.14; 0.54 ± 0.24; 0.84 ± 0.55; 2.10 ± 0.51 and 0.23 ± 0.11 for XAN, NEO, MON, ISO, BIO and 6HMPT, respectively, using the external standard method. Comparable results were obtained by the standard addition method. It is noticeable that 7BIO was not detected in the healthy serum samples analyzed., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Pteridines are a group of endogenous heterocyclic compounds whose concentrations in biological fluids may be increased in some disorders, such as infections, autoimmune disorders and cancer. In particular, pteridine concentrations in urine may represent promising noninvasive markers. However, their specificity requires further investigation. Pteridines can occur in three oxidation states with different stability. In order to enable the analysis of the unstable di- and tetra-hydroforms either an oxidation (mainly with iodine) or stabilization by reducing agents is applied. Due to the high polarity of pteridines, many analytical procedures employed ion-pair, ion-exchange or hydrophilic interaction liquid chromatography using mostly fluorescence detection. In the last decade, MS was found to be applicable. The objective of this Review is to show possibilities and different approaches in pteridine analysis in biological samples.

Occupationally-exposed lead affects the neuromuscular junction and might cause disturbances in the locomotor activity. This study was undertaken to evaluate pteridine metabolism, in which neurotransmitters are synthesized in battery workers. Urinary neopterin, biopterin and creatinine were measured using high performance liquid chromatography. Serum neopterin concentrations were detected by enzyme-linked immunoassay. Blood dihydropteridine reductase (DHPR) activities and deltaaminolevulinic acid (delta-ALA) were measured spectrophotometrically. Blood and urinary lead were detected by atomic absorption spectroscopy. Significantly increased blood and urinary lead levels, urinary neopterin, biopterin and delta-ALA were found in workers, while DHPR activities were indifferent compared to control group. Urinary creatinine decreased. This is the first study to demonstrate that increased activity of the pteridine pathway results in the accumulation of the neurotransmitters that may be responsible for the neurological disorders.

The serotonergic system is believed to play a key role in the pathophysiology of seasonal affective disorder (SAD). Tetrahydrobiopterin is an essential cofactor in the hydroxylation of tryptophan and, therefore, in the synthesis of serotonin, while neopterin is known as a marker of cell-mediated immune activity. The present study was designed to measure levels of biopterin, neopterin and tryptophan in plasma of 19 depressed patients with a history of SAD, before and after light therapy as well as in a control group. In the group of patients a significantly lower plasma biopterin and tryptophan level and a higher neopterin level was demonstrated. After light therapy, the level of biopterin increased to that of the controls but lowered again in summer. Neopterin concentrations remained on the same level after light therapy, whereas tryptophan levels increased slightly after light therapy and reached normal values in summer. It is concluded that the vulnerability for a depressive episode is enhanced by lowered levels of biopterin that, however, in SAD becomes symptomatically manifest in the presence of increased immune activity at the same time.

Background: Pteridine metabolism is impaired in the uraemic state. This may affect cardiovascular function and contribute to malnutrition. We wished to clarify further the impact of impaired pteridine metabolism., Methods: Using the HPLC method, the plasma concentrations of endogenous pteridines were determined in 64 patients with chronic renal failure (33 on intermittent haemodialysis (HD) treatment vs 31 not yet on renal replacement therapy), and in 18 healthy controls. The patients were classified into three groups on the basis of creatinine clearance (Ccr): group (a), Ccr >60 ml/min; group (b), Ccr=10-60 ml/min; group (c), all patients receiving HD., Results: Total neopterin (NP) and biopterin (BP) levels and the NP/BP ratio (a biomarker for macrophage activity) were significantly higher, whereas tetrahydrobiopterin (BH(4))/dihydrobiopterin (BH(2)) ratio (a biomarker for nitric oxide synthase and phenylalanine hydroxylase activities) was significantly lower in group (c) (118.9+/-11.7 ng/ml, 18.8+/-1.2 ng/ml, 6.79+/-0.53, and 0.26+/-0.06) than in healthy subjects (5.17+/-0.29 ng/ml, 2.83+/-0.19 ng/ml, 1.92+/-0.13, and 1.15+/-0.11; P<0.01). These significant differences were also observed between control and group (b) (12.4+/-2.20 ng/ml, 4.48+/-0.36 ng/ml, 2.81+/-0.48, and 0.74+/-0.08; P<0.01). In groups (a) and (b), significant negative correlations were found between Ccr and the total NP level (r=-0.663, P<0.01), the total BP level (r=-0.492, P<0.01), the BH(2) level (r=-0.677, P<0.01), and the NP/BP ratio (r=-0.493, P<0.01). Conversely, significant positive correlations were found between Ccr and the BH(4)/BH(2) ratio (r=0.602, P<0.01)., Conclusion: The reduction of quinoid-type BH(2) to BH(4) is modified in patients with advanced chronic renal failure, before and after the initiation of regular HD treatment. These metabolic alterations may play a role in the impaired macrophage, endothelial constitutive nitric oxide synthase, or phenylalanine hydroxylase (PH) activities observed in such patients.

In the present study, we investigated age-related changes in pteridine levels and enzymatic activity responsible for tetrahydrobiopterin biosynthesis in mouse tissues. Until about 15 weeks after the birth, the remarkable change of tetrahydrobiopterin (BH4) was observed in all tissues tested. Between 20 and 50 weeks after the birth, pteridines levels were almost constant in all of the tissues. Total biopterin levels were decreased and levels of pterin and neopterin were increased in the period exceeding 50 weeks in all of the tissues. Activities of guanosine triphosphate (GTP) cyclohydrolase I, pyrvoyltetrahydropterin synthase, and the production of BH4 were recognized by specific biochemical assays, and we investigated the age-related changes in mouse tissues. The alteration of these enzymatic activities was indicated to be similar to that described in the change of pteridine levels.

Bacterial lipopolysaccharides (LPS) induce the activity of guanosine triphosphate (GTP)-cyclohydrolase I (GTP-CHI), the first enzyme in the biosynthesis of tetrahydrobiopterin (H4bip) from GTP in endothelial cells and macrophages. In these and other cells, LPS also acts costimulatory with cytokines, i.e., mainly tumor necrosis factor-alpha (TNF-alpha). H4bip is the cofactor for nitric oxide synthase (NOS). We were interested in comparing the pteridine and nitrate levels in two baboon models: a hyperdynamic sepsis model and a hemorrhagic traumatic shock model. Our results show a similar response of pteridines (H4bip, neopterin) and nitrite/nitrate levels to an immune stimulus. LPS, which peaks rapidly, induces a sustained increase in pteridine levels in septic animals. Since hemorrhagic animals show very little response in terms of cytokine production, it was not possible to measure the induction of neopterin and nitrite/nitrate. This information could aid our understanding of the regulatory mechanisms in various forms of experimental shock.

Fasting plasma levels of tryptophan, kynurenine and the pteridines, neopterin and tetrahydrobiopterin were measured in seven patients with Gilles de la Tourette syndrome (GTS) and 10 healthy controls. Plasma kynurenine was significantly elevated in the GTS patients. The lowest patient value was higher than the highest control value. Values for tryptophan, neopterin and tetrahydrobiopterin were similar in TS patients and controls. However, in TS patients only, there was a significant negative correlation between tryptophan and neopterin and a significant positive correlation between kynurenine and neopterin when controlling for tryptophan. This finding indicates that activation of cellular immune processes is a possible explanation for the rise in plasma kynurenine.

We report a 19-year-old man with mild form of phenylketonuria. The diagnosis was first made when he was examined for the tremor at 19 years of age. He had not received the Guthrie's screening test for phenylketonuria in infancy. His development of speech and walking was almost normal. Action and positional tremor developed at two years of age, and psychomotor deterioration at five years. His intelligence was of borderline, and he entered the special class for the mentally retarded at elementary school and junior high school. His skin and iris were less pigmented than those of Japanese young adults, and his hair was rather reddish. He had mild action tremor. He showed mild mental retardation, and the WAIS was 46 in PIQ, 70 in VIQ and 53 in total IQ. T2-weighted MRI of the brain showed high signal of the deep white matter around the posterior conus of the lateral ventricles. EEG showed paroxysmal abnormalities. Serum aminogram disclosed a marked elevation of phenylalanine. Analyses of pteridine in the serum and urine disclosed a low ratio of neopterine/biopterine. An assay of the dehydropteridine reductase in erythrocytes was normal. These laboratory data indicated that his condition was caused by a deficiency of phenylalanine hydroxylase deficiency.

Transthyretin (TTR) in plasma is associated with yellow compounds. Their properties differ, and in the chicken protein a major yellow compound has recently been identified as a carotenoid, lutein, also called xanthophyll. We now show that the major yellow component extracted from human TTR has properties like a pterin derivative, 7,8-dihydropterin-6-carboxyaldehyde (2-amino-4-hydroxy-6-formyl-7,8-dihydropteridine). The human TTR derivative has chromatographic and spectral properties identical to a yellow photochemical degradation product of biopterin and a spectrum like that of the pterin aldehyde.

Concentrations of 2,4-diamino-7-hydroxy-pteridines in plasma and cerebrospinal fluid (CSF) are reported for the 0.5-8 g/m2 dose range of methotrexate in children with acute lymphoblastic leukemia or non-Hodgkin's lymphoma. This experiment has revealed that: (1) there is a wide inter-individual but relatively narrow intra-individual variability of the maximal concentrations of 2,4-diamino-7-hydroxy-pteridines in plasma during consecutive methotrexate cycles; (2) the increase in the level of the metabolites in plasma was related to increments of the methotrexate dose, but not above 5 g/m2: this can be explained by a saturable conversion of methotrexate to 2,4-diamino-7-hydroxy-pteridines; (3) significant correlations were found between simultaneous values of 2,4-diamino-7-hydroxy-pteridines in plasma and CSF; (4) the formation of 2,4-diamino-7-hydroxy-pteridines did not depend on the ages of patients receiving the same dose of methotrexate. The presence of these compounds in plasma and CSF in significant amounts creates the potential for a number of competitive interactions with pteridine-dependent metabolism which may open up new possibilities for understanding the metabolic side-effects of methotrexate therapy.

Concentrations of unconjugated pteridines (neopterin, monapterin, biopterin, pterin) were measured in the cerebrospinal fluid (CSF) of 310 patients, using a high performance liquid chromatography (HPCL) method. Our cohort included 209 controls (C), 15 patients with meningism (M), 22 with viral meningitis (VM), 17 with bacterial meningitis (BM), 9 with herpetic meningoencephalitis (HME), 2 with tuberculous meningoencephalitis (TME) and 36 with peripheral systemic infections (PI). These measurements, expressed as nmol/litre, showed a gradation of neopterin concentrations according to the type of infection: 20.1 + 6.5 in group C; 46.9 +/- 29.9 in group PI; 274.3 +/- 231.7 in group VM; 699.2 +/- 711.2 in group BM, 1,101.9 +/- 1,107.9 in group HME and 1,169 +/- 1,171.9 in group TME. There was no such gradation with biopterin. Comparisons of means showed that total concentrations in the pathology groups were very different from those observed in controls and in the neuromeningeal infections of the PI group. There was no correlation between the number of lymphocytes and the concentrations of neopterin or biopterin in the CSF. It is concluded that the concentration of neopterin in the CSF is a sensitive but little specific marker of infection, independent of CSF cellular reaction. Measuring this concentration makes it possible: 1) to evaluate the status of immune defences; 2) to predict that a meningitis will become chronic, and 3) to detect a possible parenchymal participation in a meningeal infection.

Methotrexate remains a commonly used drug in the chemotherapy of various malignancies. The known catabolites are 7-hydroxy-methotrexate, formed in the liver, and diamino-methyl-pteroic acid formed in the gut. We report for the first time evidence that 2,4-diamino-7-hydroxy-pteridine derivatives are present in the biological fluids of patients on high-dose methotrexate protocols. So far, two major derivatives have been identified as 2,4-diamino-6-hydroxymethyl-7-hydroxy-pteridine and 2,4-diamino-6-methyl-7-hydroxy-pteridine. In regard to the actual knowledge of the catabolism of pteridines, these compounds are presumably formed by intestinal bacteria during enterohepatic circulation of the drug. Their slow clearance from the body raises the question of possible interference of these compounds on pteridine-dependent enzymes, which might explain in part some of the toxic effects of methotrexate.

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.

D-erythro-neopterin, a biochemical marker of cell-mediated immune response, was found to be significantly (p less than 0.001) elevated in the serum of homosexual patients with acquired immunodeficiency syndrome and related lymphadenopathy syndrome. The most pronounced rise was observed in viral diseases causing a numerically measurable activation of the immune system such as EBV or CMV infections. In the latter patient groups, the increase in T8+, HLA-DR+ and monocytic antigen+ lymphocyte subsets correlated with the raised neopterin level. Thus, neopterin seems to be released under various conditions in viral diseases and may reflect a functional state of T-lymphocytes irrespective of the actual number expressing differentiation antigens on the cell surface.

Using the Crithidia assay 3.0 ng biopterin/ml blood was found, of which one third was present in the plasma. The erythrocyte fraction comprised 1.7 ng and the buffy coat 0.33 ng. After Ficoll separation 0.050 ng were found in the lymphocyte layer of 1 ml blood. During blast crisis of chronic myelocytic leukemias increased amounts of biopterin were found in the erythrocyte fraction and in the buffy coat. The high biopterin concentration per unit of protein in the white cell fraction indicated the presence of blasts. In Polycythemia vera increased amounts of biopterin in both the red cell fraction and in the buffy coat were also found but the percentage distribution within total cellular biopterin was markedly shifted toward the erythrocyte fraction. In cases of chronic and acute lymphocytic leukemias the low amounts of biopterin in the red cell fraction agreed with the current view of partial extinction of the erythropoietic line. The isolated lymphoblasts were characterized by high biopterin concentrations per unit of protein. During remission the biopterin patterns approached normal levels.

Serial plasma samples of 172 kidney transplant recipients and 42 chronic dialysis patients were evaluated retrospectively in a radioimmunoassay to determine the clinical relevance of plasma neopterin levels. Dialysis patients had a higher neopterin level 196 +/- 82 nmol/L (mean +/- SD) than 70 healthy controls (7 +/- 4 nmol/L). In 45 patients with a completely uneventful postoperative course, elevated pretransplant neopterin levels dropped rapidly within a week to a mean of 30 nmol/L and remained stable thereafter. In 22 outpatients with stable graft function there was a highly significant correlation between 170 paired serum creatinine and plasma neopterin values (r = 0.94). A group of 13 patients had experienced delayed graft function (ATN) without rejection. Their one-week mean neopterin level was 100 nmol/L and continued to drop in parallel with the serum creatinine. Another 15 patients rejected their kidneys irreversibly within 3 weeks-6 of them had extremely high neopterin levels during the rejection process (range 500-1000 nmol/L) that were not seen in other patients. A total of 169 rejection episodes in 43 patients were treated with bolus-dose cortisone. On the day of bolus therapy, both serum creatinine (P less than 0.002) and neopterin (P less than 0.005) were elevated. At 24 hours prior to bolus cortisone therapy, creatinine levels were not significantly elevated, whereas there was a significant rise in plasma neopterin (P less than 0.01). The overall sensitivity of neopterin increase was 86% with a 17% probability of false positives, and the sensitivity was 95% in biopsy-proved rejections. Plasma neopterin appears to be a useful marker for early detection of rejection and for identifying severe rejections that are not responsive to treatment.

This is a method for the simultaneous determination of neopterin, a product of interferon-gamma-activated macrophages, and creatinine in serum. Acidified, but not deproteinized serum is applied to a 4-propylbenzene sulfonic acid-modified silica sorbent cartridge, which quantitatively retains the analytes but not the serum proteins. The retained analytes are then eluted from the cartridge directly onto the liquid-chromatography column. Elution from the cartridge is facilitated by a pulse of 0.4 mol/L, pH 6.8 potassium phosphate buffer. On isocratic elution from an octadecylsilica column with potassium phosphate buffer (15 mmol/L, pH 6.0), neopterin is detected by its native fluorescence, creatinine by ultraviolet absorption. Detection limits are 0.5 nmol/L for neopterin, 1 mumol/L for creatinine at a sample volume of 100 microL. The standard curve is linear over the range of concentrations encountered in sera. For both neopterin and creatinine in serum, concentrations so measured agree well with results by established methods.

Plasma biopterin derivatives studied in 10 normal and 21 phenylketonuric children showed a significantly high concentration in the latter group. Biopterin derivatives correlated with plasma phenylalanine concentration, but in normal adults given an oral phenylalanine load the rate of increase with phenylalanine differed from that in phenylketonuric patients. A patient with hyperphenylalaninaemia, not due to phenylketonuria, had an abnormal biopterin derivatives response to phenylalanine distinct from that of patients with classical phenylketonuria. This may be a useful investigation to differentiate some variants of phenylketonuria.

To test the hypothesis that some of the neurologic sequelae of treatment for acute lymphoblastic leukemia (ALL) might be related to abnormalities in biopterin metabolism associated with methotrexate (MTX) therapy, total biopterin levels in cerebrospinal fluid (CSF) and plasma, and homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5HIAA) were measured in a cross-sectional study of 80 children with ALL. For comparison, biopterins were also measured in a group of children of similar age undergoing investigation for neurologic disease. In children with ALL studied before therapy, no significant difference was found between the means of plasma biopterin or CSF biopterin concentrations and the means in the control group. In children receiving MTX, plasma biopterin values were higher in the group given maintenance therapy than in children observed before treatment. CSF levels were significantly increased only in those patients who had completed 2 years of maintenance therapy. CSF concentrations of HVA and 5HIAA in patients with ALL who had received no treatment (median values 52 and 18 ng/ml, respectively) showed a wide scatter and were inversely related to age. In patients receiving MTX, concentrations of these metabolites were higher than in the untreated group, again reaching a peak in patients just completing 2 years of treatment (median HVA 110 ng/ml, 5HIAA 34 ng/ml). These results provide no support for the idea that neurotransmitter amine deficiency occurs in children with ALL receiving MTX, and indicate, rather, that amine and biopterin synthesis increases in such patients.

Plasma total biopterin concentration was measured by bioassay in 59 infants with hyperphenylalaninaemia and in 50 children with developmental regression and or movement disorder with normal plasma phenylalanine concentrations. In infants with raised phenylalanine concentrations plasma biopterin concentrations were significantly raised in proportion to the phenylalanine values. Five patients had plasma biopterin concentrations at the extremes of the range, and of these two had defective biopterin metabolism. One with low plasma biopterin concentration apparently had a partial defect of biopterin synthesis but died before investigations were complete. One with high plasma biopterin concentration, even when phenylalanine concentrations had fallen to the normal range, had dihydropteridine reductase deficiency. In this patient concentrations of homovanillic acid and 5-hydroxyindolacetic acid in the cerebrospinal fluid (CSF) were severely reduced. In children without hyperphenylalaninaemia plasma biopterin concentrations were normal. Twenty two patients were subjected to lumbar puncture, of whom six with developmental regression without movement disorder had normal CSF biopterin concentrations, and 11 with movement disorder other than torsion dystonia had significantly lower CSF biopterin concentrations. Five patients with torsion dystonia had normal biopterin concentrations.

We have recently shown that interferon-gamma is capable of activating the key enzyme of pterin biosynthesis in macrophages. This leads to excretion of the stable degradation product neopterin. In this article we present experimental evidence suggesting that stimulation of T cells by alloantigens is associated with release of interferon-gamma--which, in the case of rejection, is locally restricted and not always detectable in the bloodstream. Neopterin induced by this lymphokine, however, readily penetrates tissue barriers and is detectable in the serum. This conclusion is based on two different sets of observations: (1) If supernatants of MLCs are compared with sera from patients with documented acute rejection episodes for their interferon-gamma and neopterin levels, a marked gradient is observed to exist between interferon levels measured in vitro and in vivo; this is not the case for neopterin for which comparable levels were seen. (2) Detection of interferon-gamma in sera of allograft recipients invariably precedes an increase of neopterin; on the other hand, increasing neopterin counts are also seen in the absence of detectable interferon-gamma levels in the serum. It thus appears that although interferon-gamma release during allograft rejection is primarily restricted to the tissue, evaluation of certain metabolites of interferon-dependent metabolic pathways enables definition of its endogenous release. Whereas interferon gamma represents a less reliable marker in the monitoring of rejection episodes, it might offer an additional means to differentiate rejection from systemic infections. Such a discrimination can not be achieved with the neopterin marker.

Tetrahydrobiopterin metabolism was studied in nine Swedish patients with the Rett disease. Normal values were found for the serum biopterin level, urine biopterins level and dihydropterine reductase activity. These findings suggest that a primary disturbance of tetrahydrobiopterin metabolism is unlikely.

Luminol-dependent chemiluminescence of normal human monocytes activated by zymosan is demonstrated to be inhibited by tetrahydrobiopterin in a concentration-dependent manner. The reduced pterins tetrahydrobiopterin, dihydrobiopterin, and dihydroneopterin are all shown to be readily oxidized by the hydroxyl radical. The susceptibility of reduced pterins to free radical attack may explain the inhibition of chemiluminescence observed and an additional role of reduced pterins as free radical scavengers in tissues is considered.

Bone marrow aplasia was induced by fractionated whole body irradiation with 3000 R and restitution was started by autologous bone marrow transplantation. During the period of aplasia the amount of buffy coat biopterin clearly followed the decline of leukocytes and, vice versa, during reconstitution it largely paralleled their increase in number. The amount of red cell biopterin closely correlated with the number of reticulocytes rather than with the fairly constant values for hematocrit or of erythrocytic protein. Thereby it clearly followed the various periods of red cell recovery. The amount of cellular biopterin, its concentration with respect to cell number or to unit of protein and the percentage distribution of biopterin among the cell fractions are presented as characteristics of the activity of hemopoietic cell proliferation.

In order to determine free and conjugated pteridines, the pteridine-releasing procedure for biological materials (e.g., plasma, blood cells, bone marrow) was optimized and the parameters which influence pteridine releasing are discussed. Using reverse-phase high-performance liquid chromatography (rp-HPLC) on 5-micron ODS (C18; mobile phase, K2HPO4 buffer, pH 7.0-7.8), up to 10 pteridines (pterin-6-carboxylic acid, xanthopterin, neopterin, monapterin, isoxanthopterin, lumazine, biopterin, 6-hydroxymethylpterin, pterin) could be separated in the extracts of blood samples and determined fluorometrically (femtomolar range). In addition, three sepiapterins (sepiapterin, deoxysepiapterin, 3'-hydroxysepiapterin) could be separated with aqueous methanol as eluent (uv and fluorometric detection, respectively). The methods described are suited to compile pteridine patterns for plasma and individual cell fractions (erythrocytes, lymphocytes, buffy coat) from blood. Moreover, it was shown that the pteridine concentrations in dog blood (beagles) were in some cases substantially higher than in human blood, and that neopterin is lacking in the blood cell fractions of beagle dogs or occurs only in very low concentrations.

The levels of free phenylalanine and free tyrosine (and m-tyrosine) as well as of protein-bound tyrosine were determined by means of automated aminoacid analysis and by reaction with 1-nitrosonaphthol(2). Their absolute values as well as their percentages relative to total aminoacids of the blood were markedly increased in all tumor patients tested, as well as in experimental tumor rats. Although tetrahydrobiopterin could scarcely be detected by fluorometric methods in the blood of control persons, it is accumulated blood of all tumor patients tested. The major part was found in the erythrocyte fraction. The results are discussed with respect to decreased phenylalanine hydroxylation rate and to depressed tyrosine catabolism recently found by means of in vivo experiments in tumor-bearing rats.

After administration of tyrosine, total concentration of biopterin, the cofactor for tyrosine hydroxylase, was increased in the striatum, adrenal glands, and serum of rats, and in the serum of humans. Serum biopterin is lower in patients with Parkinson's disease than in normal controls. After oral administration of tyrosine, the increase in serum biopterin concentration was smaller in patients with Parkinson's disease (less than twofold) than in healthy controls (three-to sevenfold). These results suggest that tyrosine may have a regulatory role in biopterin biosynthesis and that patients with Parkinson's disease may have some abnormality in the regulation of biopterin biosynthesis.

During the last few years interesting results have been reported on alterations of folates and on pterins in tissues and body fluids from patients with malignant tumors. For some diseases defects in pterin biosynthesis was described. Plasma and blood cells from various tumor patients were analyzed to obtain information on the role of pteridine pattern and the pathological alterations of cells and on the relevance of some pteridine levels for diagnosis of cancer and other diseases. In the oxidized and deproteinized blood fractions (plasma, white cells, and erythrocytes) nine pteridines (pterin-6-carboxylic acid, xanthopterin, neopterin, monapterin, isoxanthopterin, lumazine, biopterin, 6-hydroxymethylpterin, and pterin) were quantified after their separation by reverse-phase high-performance liquid chromatography with fluorescence detection at exc/em = 360/460 nm.

Mean serum biopterin derivative levels in adult males and females were 1.75 microgram/l and 1.53 microgram/l respectively, increased with age and followed a cyclic pattern during menstruation. Foetal brain and liver levels at 16 weeks approximated adult concentrations. Wide distribution in adult tissues and variation in concentration independent of serum levels demonstrated local synthesis. Serum levels were significantly decreased in senile dementia, lead poisoning, coeliac disease and malignant carcinoid disease. Abnormal metabolism of phenylalanine in senile dementia suggest lowered levels of tetrahydrobiopterin in tissue. Methotrexate increased serum biopterin levels by inhibition of dihydropteridine reductase; a saturation effect was noted.

In 518 sera from blood donors below 30 years of age Neopterin was determined by radioimmunoassay. 21 of these patients (= 4.05%) showed elevated serum levels for Neopterin. By clinical investigation of these cases viral infections of the upper airways were found. Furthermore after elimination of elevated values significant differences in normal Neopterin serum levels could be demonstrated for female and male blood donors (p less than 0.01). Because elevated Neopterin serum levels indicate immune responses to several antigens, determination of Neopterin from serum may be useful for detection of infectious blood samples.