Search

Some results from storage experiments at +4 degrees C with adenine and purine nucleosides in this laboratory are compared with incubation experiments at 37 degrees C with guanosine at pH 7.0 and 7.4 and different levels of inorganic phosphate (iP). At high iP concentration the synthesis of GTP was extremely rapid. Replacement of ATP by and ATP-GTP mixture has metabolic effects on several enzymic steps e.g. phosphofructose kinase and phosphoglycerate kinase explaining the reported beneficial effect on human erythrocytes during preservation. In order to explore the role of erythrocytes in the physiological transport of purines and purine nucleosides in vitro experiments were designed in which the substances were continuously added at the appropriate concentrations by means of a dialysis tubing. Analyses of purines, nucleosides, glucose, lactate and iP in the efferent dialysis solution and the extra- and intracellular volumes reveal the metabolic processes in the erythrocytes if the transport across the dialysis tubing is accounted for. Prior to incubation the erythrocytes were stored with NaF in order to reduce the total adenylate concentration. The rate of synthesis of adenylates was about 0.05 mmol x h(-1) (calculated per litre of erythrocytes) when adenine was supplied as source for the adenine moiety of adenylates but reached a five times higher substance rate when adenosine was supplied at a sufficiently high rate. Adenosine deaminase played an important role also at "sub-physiological" concentrations. The results indicate that the erythrocytes take part in the transport of purines between organs.

Cow red cell glycolysis, which can be stimulated by a variety of purines and pyrimidines, was also found to be elevated by its own plasma. Dialyzed or charcoal-treated plasma could no longer stimulate glycolysis, suggesting that the stimulating factors may be purines or pyrimidines. Determination of purines or pyrimidines in plasma revealed the presence of xanthine (0.31 muM), hypoxanthine (0.60 muM), and adenosine (0.05 muM), as well as unknown compounds. A physiologic level of hypoxanthine, with or without xanthine and adenosine approximating their concentrations in plasma, resulted in the stimulation of cow red cell glycolytic rate by 16% (P less than 0.01). These findings suggest that plasma-borne purines may act on cow red cells in concert with as yet unidentified factors. Moreover, exchanging calf and cow plasmas produced no stimulatory effect on either calf or cow red cell glycolysis, suggesting that a) calf red cells lack some of the cellular components that respond to this stimulator and, b) only cow plasma contains this specific stimulator. In other species, including dog, cat, rabbit, rat, guinea pig, and human, stimulation of glycolysis by plasma was not observed.

Adenine analogue binding protein of rabbit erythrocytes reversibly binds [3H] adenosine with a KD of 5.3 X 10(-9) M, an association rate constant of 1.4 X 10(-12) M-1 min-1 and a dissociation rate constant of 7.5 X 10(-3) min-1, as estimated by a nonlinear curve-fitting program applied to data on the time course of the binding reaction. Independent estimates of KD by Scatchard plots and of the dissociation rate constant by dilution or adsorption of free [3H] adenosine on charcoal or by the addition of excess adenosine agreed closely with the estimates from the curve-fitting program. Inhibition of [3H] adenosine binding by a series of 77 adenosine analogues was used to define the factors determining the binding affinity of this nucleoside. These are: (1) the size and aromaticity of the purine base; (2) a glycosylic torsion angle of approximately -120 degrees; (3) the ribo configuration of the 2'-and 3'-hydroxyls and also the 5'-hydroxyl. Bulky substituents in the region of C-2' and to a lesser extent in the region of C-3' reduce binding affinity.

The influence of 1-phenyl-2-(p-hydroxyphenyl)-3,5-dioxo-4n-butylpyrazolidine-monohydrate (oxyphenbutazone) and N-(alpha,alpha,alpha-trifluoro-m-tolyl)-anthranilic acid (flufenamic acid) on the purine salvage pathway of erythrocytes and lymphocytes following incubation of heparinized whole blood with the drugs was investigated. As described in earlier experiments in vitro with enzymes derived from cell homogenates, the enzyme activities of whole cells were also decreased. Lymphocytes exhibited a reduction of purine-phosphoribosyltransferase (purine-PRT) activities (adenine-, guanine- and hypoxanthine-PRT) of about 20%. The effect on the enzyme activities in red blood cells was different. The A-PRT-activity was decreased to about 33%, the GH-PRT on the other hand, to about 10%. The possible significance of these results in vivo is discussed.

It appeared possible that abnormal purine or pyrimidine metabolism could cause cyclic hematopoiesis by analogy with the defective lymphopoiesis associated with inherited deficiencies of adenosine deaminase and purine nucleoside phosphorylase. Therefore, we examined erythrocyte purine and pyrimidine nucleotide levels, as well as plasma purine and pyrimidine nucleosides and bases in three patients and in normal controls. These studies showed that during neutropenia there was a significant elevation in the levels of guanosine triphosphate (P = 0.005) and adenosine triphosphate (P less than 0.001) in the patients' red cells not attributable to reticulocyte variation. Serial analysis of a patient's plasma showed a fivefold elevation of hypoxanthine (10.6 mumol/L) during neutropenia, with a return to normal values (1.4 mumol/L) as neutrophil numbers increased. Plasma inosine was also significantly elevated in comparison with normal control values (2.0 mumol/L vs. 0.8 mumol/L), whereas plasma and urinary uric acid were within the normal range. Serial analysis of red cells and plasma from two patients with chronic neutropenia showed no elevations of purine or pyrimidine metabolites. These results provide evidence of a link between abnormal concentrations of purine metabolites and cyclic hematopoiesis, and permit the speculation that aberrant purine metabolism is primarily related to the defective hematopoietic cell proliferation that is characteristic of this disease.

Uric acid assays were performed in the blood serum from 20 obese patients (19 women and 1 man, with a mean age of 39.75 +/- 8.16 years), before and after an average period of three weeks of hypocaloric diet (400-500 cal/day) during which they lost about 8 kg of body weight each. The variations of uricemia were followed up in relation with the weight loss and with the presence or absence of changes in the plasma lipid values. The mean rise of uricemia in the whole lot of subjects, as well as in those who lost more than 8 kg b.w. were weakly significant, as compared to the initial values. In the patients distributed according to the changes of lipid values, the increase was not significant. In conditions of rapid weight loss, even when the diet was supplemented by 70 g proteins/day, the serum uric acid values rose up to the limit of plasma urate solubility. This imposes a periodical check-up of uricemia, at the beginning, in the course and at the end of the periods of rapid weight loss, in view of detecting and correcting the eventual disorders which might occur in such conditions.

The hemodynamic effects of ATP and adenosine (i.v. infusions) were studied in dogs in parallel with quantitative determination of purines in plasma by HPLC. In two experiments, infusion were performed during treatment with dipyridamole, an uptake inhibitor of adenosine. A 50-60% reduction of mean arterial blood pressure (MABP) was induced by both ATP and adenosine at infusion rates ranging between 17-290 mumoles/min. Cardiac output was unaffected by the purine infusions, indicating that the reduction of MABP was caused by a reduction of the systemic vascular resistance. Elevated ATP and adenosine concentrations were seen in venous plasma (pulmonary artery) during infusion, while only approximately 10% recovered ATP had been degraded to adenosine. On the other hand, in arterial plasma, virtually all nucleotides had been eliminated whereas the adenosine concentrations in plasma ranged between 5 and 20 microM. The magnitude of the vasodilatation was strictly related to the arterial plasma adenosine level irrespective of whether ATP or adenosine was infused. Thus, adenosine probably mediates the vasodilatory effect of ATP.

Human red cells were incubated at 37 degrees C in physiological saline at pH 7.4 with 8-3H-adenine to label the adenine nucleotides. After washing, the cells were incubated in similar salines which were buffered to different values of pH. The loss of radioactivity from cells into medium was very slow below pH 7.4 and accelerated greatly with increasing pH up to about 7.8. At that pH the loss increased for at least 3 h when it represented more than 30% of the initial radioactivity of the cells. The loss could not be accounted for by haemolysis which remained minimal. The extracellular radioactivity was present mostly in hypoxanthine and some inosine. The loss of purine did not depend on oxygenation of the cells. These results, together with the high-affinity uptake system for adenosine, could provide a partial explanation of the turnover of adenine nucleotide purines of red cells which is many times faster than that of the cells themselves.

Significant and often substantial enhancement of the antitumor properties of several individual phleomycins , by co- administration via intraperitoneal injection of a number of purine analogues, is demonstrated in rats and mice having three diverse tumors. It is evident that the dose levels of both the phleomycin and the amplifier are very significant and that optimal levels vary widely with the actual agents used. Constant serum levels of amplifier can be maintained for several days by administration via silastic-pellet implantation rather than injection, and this route of administration is an effective alternative for amplifiers of low solubility.

The regulation of ATP metabolism by inorganic phosphate (Pi) was examined in five normal volunteers through measurements of ATP degradation during relative Pi depletion and repletion states. Relative Pi depletion was achieved through dietary restriction and phosphate binders, whereas a Pi-repleted state was produced by oral Pi supplementation. ATP was radioactively labeled by the infusion of [8(14)C]adenine. Fructose infusion was used to produce rapid ATP degradation during Pi depletion and repletion states. Baseline measurements indicated a significant decrease of Pi levels during phosphate depletion and no change in serum or urinary purines. Serum values of Pi declined 20 to 26% within 15 min after fructose infusion in all states. Urine measurements of ATP degradation products showed an eightfold increase within 15 min after fructose infusion in both Pi-depleted and -supplemented states. Urinary radioactive ATP degradation products were fourfold higher and urinary purine specific activity was more than threefold higher during Pi depletion as compared with Pi repletion. Our data indicate that there is decreased ATP degradation to purine end products during a relative phosphate repletion state as compared to a relative phosphate depletion state. These data show that ATP metabolism can be altered through manipulation of the relative Pi state in humans.

1. Enzymological and metabolic data in a patient with nucleoside phosphorylase (NP) deficiency are described. 2. Incubation of intact NP-deficient red cells with [14C]adenosine showed a rapid uptake and conversion to inosine. Almost no radioactivity was incorporated in the adenosine nucleotides and no hypoxanthine labeling could be detected. 3. Incubation with [14C]inosine resulted in a rapid conversion to IMP in the normal intact red cells but in an accumulation of inosine in the medium with the erythrocytes of the patient, proving again that a NP deficiency is present. 4. The high PRPP level found may result from impaired consumption due to lack of substrates for the salvage enzyme HGPRT. 5. Incubation with [14C]hypoxanthine and [14C]adenine showed that normal HGPRT and APRT activities were present in the NP-deficient red cells. 6. In serum and urine of the patient the levels of inosine and guanosine were considerably increased, while the serum and urinary levels of uric acid were very low. In the two deceased sisters NP deficiency was also strongly suggested by analyses of the serum purines, of stored deep frozen samples.

The effect of adenosine and some related compound was studied on blood flow in feline oral mucosa. Changes in the rate of disappearance of 125Iodine (k-value) from a local depot in the oral mucosa was used to determine changes in the mucosal blood flow. Infusion of adenosine caused a dose-dependent increase of blood flow. Two stable adenosine analogues, adenosine 5'- ethylcarboxamide ( NECA ) and L-phenylisopropyl-adenosine (L-PIA), were 20 and 10 times more potent than the parent compound. Dipyridamole (2 mg/kg), which blocks adenosine uptake, significantly enhanced the potency of adenosine. Theophylline (10 mg/kg) inhibited the vasodilatory effect of adenosine and of the adenosine analogues. The result suggest that adenosine may be involved in the regulation of blood flow in the oral mucosa.

A reverse phase partition mode of high pressure liquid chromatography was developed for the estimation of purine, pyrimidine and pyrazolopyrimidine nucleosides and bases. This method has been applied to the investigation of purine and pyrimidine metabolism in subjects with gout and/or renal failure during allopurinol therapy. Plasma levels of hypoxanthine and xanthine were quantitated in all subjects. No significant differences were observed between healthy males and gouty subjects with or without allopurinol medication. There was a significant increase in xanthine levels in patients with gout and renal failure, or renal failure on allopurinol, and plasma levels of oxipurinol were increased, but no correlation with glomerular filtration rate (GFR) was observed. The separation method was also used to examine urines from patients with known abnormalities of purine and pyrimidine metabolism in an attempt to evaluate its usefulness as a urine screening technique.

Studies on purine de novo-synthesis and cyclic AMP in normal and pathologic leukocytes. The destruction of neoplastic cells by cytostatic agents depends on the existence of metabolic reactions within the cells which can be specifically blocked by these agents. Concerning the action of purine antagonists there is, as yet, no clear evidence whether de novo synthesis of purines takes place in normal as well as in leukemic leukocytes. Therefore studies were performed to evaluate the capacity of these cells to incorporate the purine precursor, glycine, into their acid-soluble adenine nucleotides. The results demonstrate that "resting" human lymphocytes are not capable of building purines de novo. However, stimulation of the cells with a mitogen (Phytohemagglutinin) leads to the induction of de novo purine synthesis. Leukemic cells do not show uniform behaviour. In order to get more information about the mechanisms by which synthesis of purines as well as the morphologic transformation of lymphocytes may be induced, the metabolism of cyclic AMP was investigated. According to our results the nucleotide does not play an essential role in initiating the transformation process, but it seems to have regulating function in the further development of cell transformation. The studies furthermore indicate an important role of cyclic AMP during proliferation of leukemic cells.

Immune deficiency is characteristic of alcoholic subjects. These subjects usually show altered lymphocyte function. We determined the activities of adenosine deaminase (ADA) and ecto-5'-nucleotidase (ecto-5'N) in lymphocytes from 54 subjects: 15 healthy controls, 28 non-cirrhotic alcoholics, 8 alcoholic cirrhotics and 3 non-alcoholic cirrhotics. Whereas ADA activity was the same for all 54 subjects, ecto-5'N activity was in general lower in alcoholic subjects after cessation of alcohol intake. Following alcohol intoxication, however, ecto-5'N activity increased. The decrease of ecto-5'N activity in alcoholic subjects might be explained by shedding of the ecto-enzyme and alteration of lymphocyte subpopulations. We observed decreased mitogenic-induced lymphoblastic transformation in 3 patients with cirrhosis. All other subjects (including healthy controls) had normal mitogenic-induced blastogenesis. Interestingly, following alcohol intake, non-stimulated lymphoblastic transformation increased, leading to an apparently decreased stimulation index.

Some results from storage experiments at +4 degrees C with adenine and purine nucleosides (cf. [1]) are compared with incubation experiments at 37 degrees C with guanosine at pH 7.0 and 7.4 and three different levels of inorganic phosphate (iP). At the lower pH and 20 mmol . l-1 iP the synthesis of GTP was extremely rapid; the intracellular concentration reached 0.5 mmol . l-1 within 25 min constituting 1/3 of the purine nucleoside triphosphate pool. Replacement of ATP by an ATP-GTP mixture has metabolic effects on several enzymic steps, e.g., phosphofructose kinase and phosphoglycerate kinase explaining the reported beneficial effect on human erythrocytes during preservation. In order to explore the role of erythrocytes in the physiological transport of purines and purine nucleosides in vitro experiments were designed in which the substances were continuously added to the appropriate concentrations by means of a dialysis tubing. Prior to incubation the erythrocytes were stored with NaF in order to reduce the total adenylate concentration. The rate of synthesis of adenylates was about 0.05 mmol . h-1 (calculated per litre of erythrocytes) both when adenosine and adenine were supplied as source for the adenine moiety of the adenylates. Analyses of purines, nucleosides, glucose, lactate and iP in the efferent dialysis solution and the extra- and intracellular volumes reveal the metabolic processes in the erythrocytes if the transport across the dialysis tubing is accounted for. The low capacity of the erythrocyte adenylate synthesis indicates that only a minor part of the adenine compounds physiologically exported by the liver could be delivered as intraerythrocytic adenylates.

In order to study if enzymes of purine metabolism could be used as cell markers in B-chronic lymphocytic leukemia (B-CLL), the activities of adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP), and 5'-nucleotidase (5'N) were repeatedly measured in blood mononuclear cells from B-CLL patients and were compared to those obtained in normal controls. Enzyme activities in patients were also compared to other biological parameters indicative of B-CLL to activities of ADA and PNP in erythrocytes. Results show that B-leukemic cells display abnormal enzyme patterns: subnormal ADA activity is characteristic; 5'N activity is depressed in 60% of the cases but increased in 15%. An inverse relationship between PNP activity and corresponding lymphocytosis is observed in leukemic but not in normal cells. The enzymatic anomalies seem to be linked to the presence of an unusual peripheral lymphocytic population, induced by the leukemic process. Indeed, ADA and PNP are not abnormal in erythrocytes. In untreated nonevolutive patients, the enzyme profile tends to remain stable throughout the course of the illness; normalization of enzyme patterns in treated patients occurs only when therapy induces improvement in T and B cell distribution.

We tested the hypothesis that there is an enhanced rate of hypoxanthine salvage in two siblings with hereditary xanthinuria. We radiolabeled the adenine nucleotide pool with [8-14C]adenine and examined purine nucleotide degradation after intravenous fructose. The cumulative excretion of radioactivity during a 5-d period was 9.7% and 9.1% of infused radioactivity in the enzyme-deficient patients and 6.0 +/- 0.7% (mean +/- SE) in four normal subjects. Fructose infusion increased urinary radioactivity to 7.96 and 9.16 X 10(6) cpm/g creatinine in both patients and to 4.73 +/- 0.69 X 10(6) cpm/g creatinine in controls. The infusion of fructose increased total urinary purine excretion to a mean of 487% from low-normal baseline values in the patients and to 398 +/- 86% in control subjects. In the enzyme-deficient patients, the infusion of fructose elicited an increase of plasma guanosine from undetectable values to 0.7 and 0.9 microM. With adjustments made for intestinal purine loss, these data support the hypothesis that there is enhanced hypoxanthine salvage in hereditary xanthinuria. Degradation of guanine nucleotides to xanthine bypasses the hypoxanthine salvage pathway and may explain the predominance of this urinary purine compound in xanthinuria.

Storage of erythrocyte units from donors with ITP pyrophosphohydrolase deficiency have been studied and compared with units from normal donors. Verifying other investigations the incidence of this genetic disorder was found to be as high as about 3%. Hemolysis in the units was higher than in other units and there was a tendency to low total adenylate concentration. It is suggested that blood centers should organize a quality assurance program where one of the aims should be to detect genetic disorders that make the erythrocytes from the donors less suitable for long term liquid storage.

A survey is given of our present investigations of the pharmacokinetics of 6MP and the effects of the combination of MTX and 6MP on purine and pyrimidine metabolism. Both drugs are used in the oral maintenance therapy of ALL in children. The very low serum-concentrations after oral administration of 6MP and the short serum-half-life-times after intravenous administration point to more efficacy after prolonged intravenous infusion. The penetration of 6MP in the cerebrospinal fluid after intravenous administration could account for (prophylactic) treatment of the CNS in ALL. Pretreatment of leukemic lymphoblasts with MTX results in an increase of intracellular PRPP due to inhibition of purine de novo synthesis. This can be used for an increased incorporation and conversion of 6MP. Thus, increased incorporation and conversion of 6MP can be obtained when 6MP is added to MTX-pretreated leukemic lymphoblasts at that point of time where MTX causes maximal PRPP accumulation. This will result in increased incorporation of 6MP into nucleic acids and thus in enhanced cytotoxicity. Pharmacokinetic and metabolic investigations of 6MP and MTX could lead to a more rational and more effective use of both agents in patients with ALL.

Inherited purine and pyrimidine disorders may be associated with serious, sometimes life-threatening consequences. Early and accurate diagnosis is essential. Difficulties encountered when using existing high pressure liquid chromatographic (HPLC) methods led to the development of an improved method based on prior fractionation of urine. The advantages are as follows. 1. Production of fingerprints demonstrating altered urinary excretion patterns characteristic of any one of ten different disorders, in 30 minutes. 2. Positive identification and quantification by comparison with established methods (using conventional chromatography, electrophoresis and UV spectrophotometry) in addition to specific retention times and characteristic UV absorbance ratios at two separate wavelengths (245 and 280 nm) by HPLC. 3. Direct analysis of all the purines and pyrimidines normally found in human body fluids as well as identification of abnormal compounds. 4. Short time between successive analyses while maintaining excellent resolution between compounds of interest and column longevity. 5. Improved separation of the different adenine-based compounds encountered in some disorders, plus demonstration of potential interference by dietary or drug metabolites. 6. Applicability to the monitoring of therapy involving a variety of different purine and pyrimidine analogues. Particular attention should be paid to sample preparation. Plasma profiles will confirm the diagnosis in some, but not all, of these disorders.

The contribution of enhanced purine salvage to the decreased total purine excretion associated with allopurinol therapy was measured by the intravenous administration of tracer doses of [8-(14)C] adenine to four patients with gout and normal purine salvage enzyme activity and four patients with the Lesch-Nyhan syndrome and absent purine salvage activity. The mean cumulative excretion of radioactivity 5 days after the adenine administration to patients not receiving and receiving (off and on) allopurinol therapy was 6.1% and 3.6% of infused radioactivity for gouty subjects and 15.9% and 20.8% for the Lesch-Nyhan patients. Urate pool size and urate turnover, as measured by pool labeling with [2-(14)C]uric acid, were substantially decreased in both groups of patients during allopurinol therapy. The intestinal loss of uric acid was estimated from these pool measurements on and off allopurinol. With a correction for this extrarenal purine loss, the mean cumulative excretions of radioactivity 5 days after adenine administration to patients off and on allopurinol therapy were 11.9% and 4.8% for the gouty subjects and 31.7% and 24.5% for the Lesch-Nyhan patients. In vitro studies demonstrated no alteration of the synthesis or degradation of adenine nucleotides by allopurinol in cultured human diploid fibroblasts. These observations suggest that enhanced purine salvage is an important component leading to decreased purine excretion during allopurinol therapy.

A method is described for the separation of adenine and guanosine from whole blood and plasma fractions, containing purinderivatives as anticoagulant-additives. The highest concentration determined per gram human albumin or gamma-globulin was 0.5 mg. Due to interactions with blood cells guanosine could only be detected in plasma which was stored for more than one week as whole blood. The amount of adenine and guanosine to be expected in therapeutical products is uncritical according to published pharmacological investigations.

We have studied purine metabolism in mononuclear and polymorphonuclear cells from uraemic patients using microradiochemical enzyme assays and high-pressure liquid chromatography. In mononuclear cell lysates the mean activities of adenosine deaminase (EC 3.5.4.4) and 5'-nucleotidase (EC 3.1.3.5) were significantly diminished. The activities of adenylate kinase (EC 2.7.4.3), purine nucleoside phosphorylase (EC 2.4.2.1), adenine phosphoribosyltransferase (EC 2.4.2.7), and hypoxanthine phosphoribosyltransferase (EC 2.4.2.8) were not significantly different in the two groups. The activities of adenosine deaminase and adenine phosphoribosyltransferase were reduced in the polymorphonuclear cell lysates. No clear differences emerged in the concentration of adenine nucleotides in the mononuclear cells. The significance of these changes, which are less marked than those in erythrocytes, is discussed with reference to the immunodeficiency associated with uraemia.

Deficiency of purine nucleoside phosphorylase (PNP) was detected in a 3-yr-old boy who was admitted for investigation of a behavior disorder and spastic diplegia. The urinary excretion of purines, analyzed by high-performance liquid chromatography, showed the presence of large amounts of (deoxy)inosine and (deoxy)guanosine and low uric acid levels. Analysis of the (deoxy)nucleotide pools of erythrocytes showed elevated levels of deoxyguanine nucleotides and NAD and decreased guanine nucleotides. PNP activity in red blood cells was 0.1-0.5% of normal on two occasions and undetectable on four later measurements. Furthermore no immunoreactive material could be detected in his red cell lysate using an anti-PNP antiserum. PNP activities in the red cells of the patient's parents were 35 and 50% of normal. The presence of (minor) residual PNP activity in the patient enabled the investigation of some enzyme properties after partial purification. No abnormalities could be detected in substrate affinity for inosine, heat stability, and electrophoretic properties. In the heterozygous parents no signs of a mutant enzyme could be found. The molecular specific activities of the parental enzymes were also normal, indicating that no immunoreactive material attributable to inactive-mutant enzyme subunits was present. A striking feature of the patient is the prevailing neurologic abnormalities presumably caused by the metabolic disorder. A severe lymphopenia exists; however, clinical symptoms of an immune deficiency did not become apparent until the age of 4 yr.

Adenine, adenosine, inosine, hypoxanthine, xanthosine, xanthine, guanine and guanosine blood levels in 11 Duchenne muscular dystrophy patients treated with allopurinol, 10 untreated patients and 8 healthy controls, were determined by HPLC. Serum ADA, PNP and 5'-NT were also determined. Untreated patients showed lower adenine (p less than 0.001) and higher adenosine, xanthine, ADA and PNP levels (p less than 0.01) than controls. Treated patients had lower adenine and higher xanthine levels (p less than 0.001), but higher hypoxanthine, xanthosine and guanine levels (p less than 0.001), than controls, with normal ADA and PNP. The changes observed in ADA and PNP levels suggest an involvement of these enzymes in accelerated degradation of purines in Duchenne dystrophy.

A comprehensive investigation of the high-performance liquid chromatographic separation of nucleosides, their bases and other low-molecular-weight UV-absorbing compounds that might be found in serum is reported. A buffer-methanol gradient was used in conjunction with chemically bonded, microparticulate columns to separate many of the biologically important compounds under study in minimal time with maximal resolution. Retention data, absorbance ratios (280/254 nm) and fluorescence responses are reported for 86 nucleosides, bases, nucleotides and other UV-absorbing compounds commonly encountered in biological studies.