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Background: There are limited data on the effect of body mass index (BMI) on the actions of fixed-dose regadenoson. The purpose of this study was to determine the effect of BMI on the efficacy, side effects, and plasma concentration of regadenoson for Myocardial Perfusion Imaging (MPI)., Methods and Results: The study included 2,015 subjects from the ADVANCE MPI trials. Initial adenosine MPI was followed by randomization to regadenoson (400-μg bolus injection) or adenosine (6-minute infusion) MPI. Subjects were classified according to BMI into six categories from underweight (<20 kg/m(2)) to extremely obese (≥40 kg/m(2)). PK modeling was used to predict the effect of BMI on plasma regadenoson concentration (PRC). Adenosine-regadenoson agreement rates for the presence and extent of reversibility were similar across BMI categories (P > .05). The incidence of side effects was also similar across BMIs (P ≥ .06). Subjects were less likely to feel very or extremely uncomfortable after regadenoson vs adenosine in all groups with BMI ≥ 25 kg/m(2), but this trend was not statistically significant in subjects with BMI 20-24 kg/m(2) (P > .05). PRC was inversely related to BMI with 19% higher PRC in the underweight and 36% lower PRC in the extremely obese compared with a normal weight subject., Conclusions: BMI does not alter the efficacy of regadenoson MPI despite lower PRC in high BMI subjects, or its side effect profile despite higher PRC in low BMI subjects. Regadenoson is better tolerated than adenosine but this benefit seems to lose statistical significance in subjects with BMI < 25 kg/m(2).

An oleic acid-based microemulsion system with a member of the Tween series or Cremophor EL as the surfactant and a short-chain alcohol as the cosolvent was developed for rapid-onset intranasal delivery of sildenafil. The phase behaviour and solubilization capacity of the microemulsion system were characterized, and nasal absorption of sildenafil from the microemulsion formulations was investigated in rabbits. Sildenafil displayed a high solubility of 124 mg/mL in the microemulsion consisting of 40% oleic acid, 10% H(2)O, and 50% Tween 80:ethanol (EA) (at a 1:4 weight ratio). Nasal absorption of sildenafil from this microemulsion was found to be fairly rapid. With a 10-mg dose, the onset of action was arrived instantly following intranasal administration and the duration was over 3 h using an in vivo rabbit studies. In addition, nasal ciliotoxicity studies were carried out using in vivo rat nasal mucosa model and showed no ciliotoxicity. Therefore, the prepared systems are no serious nasal ciliotoxicity for intranasal administration. The microemulsion system composed of oleic acid, Tween 80, EA, and water may be a practical approach for the rapid-onset delivery of sildenafil for the treatment of erectile dysfunction.

Objective: To determine the effects of chronic administration of sildenafil citrate on healthy pregnant rats., Design: In vivo animal experimental study., Setting: Fundación IVI-Instituto Universitario IVI, Valencia, Spain., Sample: Pregnant and non-pregnant Wistar rats exposed to chronic administration of sildenafil., Methods: Placental cross-barrier and feto-maternal relationship levels, maternal blood pressure, and haemodymamic effects on uterine arteries were evaluated. The effect of growth on weight and fetal tissues, and on perinatal outcome, was investigated., Main Outcome Measures: Maternal blood pressure, blood viscosity, vascular indices of uterine arteries and fetal ductus venosus, plasmatic levels of sildenafil, embryo/fetal and litter weights, perinatal/postnatal survival rates., Results: Sildenafil citrate crossed the placenta. The maternal and fetal levels of sildenafil, and its metabolite desmethyl-sildenafil, demonstrated a positive linear correlation in treated pregnant animals versus controls; a selective maternal hypotensive effect without changes in uterine vascular resistance was noted on days E8 and E11 (embryonic day). The lower pulsatility index of the ductus venosus on day E18 suggests fetal overflow at the end of the pregnancy. Effects on offspring were placental and liver enlargement, and increased fetal weight gain in the second half of pregnancy (irrespective of liver enlargement) and at birth. Perinatal and postnatal survival rates in the sildenafil group remained unaltered. No haemodynamic effects were evident in non-pregnant animals., Conclusions: In normotensive rats, sildenafil appears to have a selective effect at the onset of pregnancy, implying increased fetal blood supply, and increased fetal weight, and placental and liver enlargement, but no increased perinatal mortality., (© 2011 The Authors Journal compilation © RCOG 2011 BJOG An International Journal of Obstetrics and Gynaecology.)

Background: Pulmonary hypertension as a frequent complication of left heart disease (PH-LHD) is characterized by lung endothelial dysfunction and vascular remodeling. Although PH-LHD contributes to morbidity and mortality in heart failure, established therapies for PH-LHD are lacking. We tested the effect of chronic sildenafil treatment in an experimental model of PH-LHD., Methods and Results: In Sprague-Dawley rats, PH-LHD was induced by supracoronary aortic banding. Oral sildenafil treatment (60 mg/kg daily) was initiated after 7 days, and lung endothelial function (n=5), vascular remodeling, and right ventricular function (n=11 each) were analyzed 9 weeks after banding. As compared with sham-operated controls, aortic banding induced pulmonary hypertension and lung endothelial dysfunction evident as lack of endothelial nitric oxide production and endothelium-dependent vasodilation. These changes were associated with an increased pulmonary vascular resistance, medial thickening, and biventricular cardiac hypertrophy. Sildenafil treatment largely attenuated these pathological changes and was not associated with detectable adverse effects pertinent to lung vascular barrier function, edema formation, or systemic hemodynamics., Conclusions: Our data identify sildenafil as a promising therapy for PH-LHD. In light of its documented protective effects at the myocardial level in heart failure, sildenafil presents a particularly attractive strategy in that it simultaneously targets cardiac remodeling and secondary PH-LHD.

To develop sildenafil lactate, a salt form of sildenafil with improved solubility and bioavailability of poorly water-soluble sildenafil base, this salt form was prepared using a spray dryer. Its solubility and pharmacokinetics in rabbits were evaluated compared with sildenafil base and sildenafil citrate. Sildenafil lactate improved the solubility of sildenafil in various solvents including distilled water compared with sildenafil citrate. It provided higher AUC and C(max) and, shorter t(1/2) values than did the other materials, indicating that it improved the oral bioavailability of sildenafil in rabbits. Our results suggest that sildenafil lactate would be useful to deliver sildenafil in a pattern that allows fast absorption and late metabolism. Furthermore, the plasma concentration at 0.25 h in sildenafil lactate was similar to the C(max) value at T(max) (0.5 h) in sildenafil citrate. Thus, sildenafil lactate might provide a faster onset of action and immediate erection compared with sildenafil citrate, the conventional drug.

This study was designed to examine the influence of sub-maximal exercise on purine and pyrimidine catabolism in horses. Ten horses were initially trained for 12 weeks at the end of which they underwent a standardized exercise test (SET); venous blood samples were taken at rest, 5 and 30 min after the SET. Six untrained healthy horses, from which a blood withdrawal was taken at rest, were used as the control group. Samples were analyzed by HPLC for the simultaneous determination of uric acid, uridine, β-pseudouridine and creatinine in plasma. Glucose and lactate were measured in blood. Trained horses had basal uridine levels significantly lower than sedentary horses. The SET caused significant increase in plasma uric acid, uridine, β-pseudouridine and creatinine. Following the SET, a significant negative correlation was found between plasma uridine and glucose, whilst a significant positive correlation was observed between plasma uric acid and creatinine. These results indicate that increase in energy demand during exercise in the horse causes not only the degradation of purine but also of pyrimidine compounds, the latter possibly exerting a control on glucose uptake as also demonstrated in human beings.

Aims: To assess pharmacokinetics and pharmacodynamics of a 10 mg intravenous sildenafil bolus in pulmonary arterial hypertension (PAH) patients stabilized on 20 mg sildenafil orally three times daily., Methods: Pharmacokinetic parameters were calculated using noncompartmental analysis., Results: After an acute increase, plasma concentrations stabilized within the range reported previously for a 20 mg oral tablet. At 0.5 h, mean ± SD changes from baseline were -8.4 ± 11.7 mmHg (systolic pressure), -2.6 ± 7.3 mmHg (diastolic pressure) and -3.5 ± 10.4 beats min(-1) (heart rate). There was no symptomatic hypotension., Conclusions: Although further research is warranted, a 10 mg sildenafil intravenous bolus appears to provide similar exposure, tolerability and safety to the 20 mg tablet., (© 2011 The Authors. British Journal of Clinical Pharmacology © 2011 The British Pharmacological Society.)

Pharmacokinetics of sildenafil and its metabolite, N-desmethylsildenafil, in humans and rats with liver cirrhosis (LC) and diabetes mellitus (DM), alone and in combination (LCD) did not seem to be reported. Sildenafil was administered intravenously (10 mg/kg) and orally (20 mg/kg) to control, LC, DM, and LCD rats. Expression of intestinal CYP isozymes in those rats was also measured. In LC, DM, and LCD rats, the areas under the curve (AUCs) of intravenous sildenafil were significantly greater (by 195%, 54.2%, and 127%, respectively) than controls. In LC and LCD rats, AUCs of oral sildenafil were significantly greater (3010% and 2030%, respectively) than controls. In LC, DM, and LCD rats, significantly greater AUCs of intravenous sildenafil were due to the slower hepatic extraction of sildenafil (because of decrease in the protein expression of hepatic CYP2C11 and 3A subfamily in LC and LCD rats, and CYP2C11 in DM rats). In LC and LCD rats, greater magnitude of increase in AUCs of oral sildenafil than those after the intravenous administration could be mainly due to the decrease in the intestinal extraction of sildenafil (because of decrease in the protein expression of intestinal CYP2C11 in LC and LCD rats).

Both platelet aggregation and high blood pressure are associated with development of atherosclerosis. Among other factors that modulate platelet aggregation and blood pressure, extracellular purines (e-purines) influence these processes via purinoceptors P1 and P2 for which they are natural ligands. We hypothesized that ecto-enzymes such as nucleoside triphosphate diphosphohydrolases (NTPDases), adenylate kinase, 5'-nucleotidase, and adenosine deaminase that regulate the level of e-purines may be involved in the development of atherosclerosis. The enzymatic assays were performed either on the fragments of human abdominal aortas obtained after death or on abdominal aneurysm samples collected during surgery. The substrates and products such as adenine nucleosides and nucleotides were analyzed using reverse phase high-performance liquid chromatography (HPLC) method. Here, we estimated and demonstrated the activities of these ecto-enzymes in the patients with atherosclerosis or atherosclerosis-like diseases such as abdominal aneurysm, myocardial infarction, or Leriche syndrome (LS) with worse thrombosis of extremities. In particular, we noticed reduction in activity of NTPDase1(app), NTPDase2(app), ecto-adenylate kinase( app), and ecto-adenosine deaminase(app); however, ecto-5'-nucleotidase(app) that hydrolyzed e-adenosine monophosphate (e-AMP) into e-adenosine did not show any significant changes. This led us to suggest that alteration of the activity of examined ecto-enzymes is responsible for the development of atherosclerosis or atherosclerosis-like diseases.

This study investigated the effect of sildenafil on uterine volumetric blood flow (UVF) and vascular impedance in nonpregnant, nulliparous women. Fifteen women were randomized in a double-blind fashion to receive either placebo or sildenafil (25 or 100 mg) during the luteal phase of the menstrual cycle. Color Doppler ultrasound of both uterine arteries was performed at baseline and at 1 and 3 hours postdosing to calculate resistance index (RI) and UVF. Those who received sildenafil significantly increased UVF and decreased RI over the 3-hour monitoring period. When UVF responses to sildenafil were examined as a function of baseline UVF, a significant increase in UVF was observed in only those participants with higher baseline UVF. Overall, women in the luteal phase demonstrated a significant increase in UVF in response to sildenafil. However, this increase appears to be directly associated with basal UVF.

The purpose of the current study was to investigate findings, peculiarities, parameters and indicators of protein, carbohydrate, lipid and purinic metabolism in the blood, erythrocytes and homogenate of femur muscles in case of moderate and maximal physical exertion. 24 male rats were studied. The above rats underwent muscular exertion in the form of loads during swimming. The load accounted for 10% of the body weight. Specific changes of metabolism due to the influence of moderate and maximal physical muscular exertion were revealed. Mechanisms of atherogenic effects in maximal muscular exertion are discussed.

Purpose: The main purpose of this work was to investigate the effect of pummelo juice on the pharmacokinetics of sildenafil after oral administration., Methods: This was a comparative, randomized, two-period, two-treatment, two-sequence, single dose, crossover study investigating the effect of pummelo juice on the pharmacokinetics of sildenafil citrate tablets (equivalent to 50 mg sildenafil) in healthy male participants under fasting conditions compared with water as a control. Six healthy normal adult males were administered 50 mg sildenafil with pummelo juice or water at two different periods in a crossover study., Results: Study results showed that pummelo juice reduced the rate and extent of sildenafil bioavailability to around 60% [maximum plasma concentration (C(max)); from 212.44 ng/ml to 134.07 ng/ml, 90% confidence interval (90% CI) 44.70-89.11, and area under the plasma concentration time curve from zero to infinity (AUC(infinity)); from 564.51 ng.hr/ml to 336.87 ng.hr/ml, 90% CI 39.17-90.92]. This interaction was opposite to that expected and is speculated to be due to either an effect on transporters or a physicochemical interaction between sildenafil and some components of the juice., Conclusion: Patients should be advised not to drink pummelo juice before or immediately after taking sildenafil.

Sildenafil is used to treat pulmonary hypertension in neonatal and pediatric patients. Pharmacokinetic studies in these patients are complicated by the limited sample volume. We present the validation results of an assay method to quantitate sildenafil and desmethylsildenafil simultaneously in 50 microL of plasma. Deuterated sildenafil was used as an internal standard. After liquid-liquid extraction, analytes were separated on an ultra-performance liquid chromatography (UPLC)-column and quantified via tandem mass spectrometry. The calibration range was linear, with acceptable accuracy and a precision of <15% for both compounds. The lower limits of quantification were 1 ng/mL. Matrix effects were present, but inter-plasma batch variability was under 12%. The method was successfully applied to samples from a pharmacokinetic study into sildenafil pharmacokinetics in neonates, making maximum use of the limited number and amount of plasma samples available., ((c) 2009 John Wiley & Sons, Ltd.)

Novel sample preparation approaches for HPLC bioanalysis based on the phenomenon that acetonitrile can be separated from water by adding salts or cooling at subzero temperatures have been reported. These two methods are superior to conventional liquid-liquid extraction since the separated acetonitrile phase can be directly injected to the RP-LC system. However, the salting-out method suffers from a potential problem that the remained salt in the acetonitrile phase may harm the MS detector, while the subzero-temperature method is troublesome to operate. Here, we have reported a similar phase separation phenomenon that the acetonitrile aqueous mixture can be separated by adding a hydrophobic solvent; and capitalising on this phase transition phenomenon, we have proposed an alternative approach, named solvent induced phase transition extraction (SIPTE), to extract drug from plasma for HPLC-MS analysis. The proposed SIPTE method is much simpler and avoids contaminating the MS detector. Three structurally diverse drugs were selected as test compounds to design the SIPTE method and to validate the efficiency of this method. The four goals of plasma sample pretreatment for HPLC-MS analysis, i.e. removal of proteins, removal of other low-molecular interferences, preconcentration of the analytes of interest, and matching the sample solvent with the HPLC-MS system, can be rapidly performed in a very simple step by using the SIPTE method., (2009 Elsevier B.V. All rights reserved.)

An ultrafast and efficient high-performance liquid chromatographic (LC) method was developed to purify positron emission tomography (PET) radiopharmaceuticals as well as for metabolite analysis of the plasma sample. Chromatographic separation was achieved on a short (60 mm length) semipreparative (10 mm I.D.) column packed with 2.5-mum particles using a mixture of acetonitrile and sodium phosphate buffer as the mobile phase at a flow rate of 8-10 ml/min. Under the optimum conditions, excellent separation of the target PET probe was obtained from chemical/radiochemical impurities or radioactive metabolites with a very short run time of 2 min. This characteristic enabled significant shortening of the purification and evaporation processes in the production of short-lived radiopharmaceuticals and highly sensitive radiometric analysis with good temporal resolution during the metabolism study., (Copyright 2010 Elsevier Inc. All rights reserved.)

A simple, precise, rapid and accurate HPLC method for the determination of sildenafil citrate (SLD) in human plasma has been developed based on previous reports, to offer an alternative way to detect and quantify SLD and also to improve some characteristics of the validation process. Chromatography was carried out on a C18 reversed-phase column, using a mixture of acetonitrile: ammonium acetate (0.3 M pH 6.8) (1:1 v/v) as the mobile phase at a flow of 0.750 mL/min. Diazepam was used as an internal standard (IS) and detection was by UV at 240 nm. Samples were extracted with 200 microl of 0.1 M Na2CO3 and 5 mL of diethyl ether: dichloromethane (60:40). Retention times of SLD and IS were 4.8 and 7.1 min, respectively; total run time was 10 min. The linear range of SLD was found to be 20 - 1000 ng/mL. Limit of quantitation (LOQ) and limit of detection (LOD) were calculated to be 10 and 20 ng/mL, respectively. An improved percentage recovery of analyte is reported, showing a fast and reproducible approach to detect SLD in plasma human sample. The method was validated for its linearity, precision, accuracy, recovery, stability and specificity.

Sildenafil is a selective inhibitor of phosphodiesterase type 5 enzyme. A simple and sensitive LC-MS assay was developed to determine the concentration of sildenafil in human plasma. Sildenafil and omeprazole (internal standard) were extracted from the plasma with diethyl ether. The extract was evaporated under nitrogen and the residue was constituted with ACN and injected onto Novapak C(18) column (75x3.9 mm, 4 microm). The mobile phase consisted of 90% ACN plus 10% ammonium acetate (20 mM) containing 0.02% formic acid and was delivered isocratically at a flow rate of 0.2 mL/min. Sildenafil and omeprazole were monitored using a positive electrospray mode with single-ion recording set at m/z 475 and 346, respectively, which are consistent with [M+H](+) molecular ions, and the run time was less than 5 min. The detection limit of sildenafil was 0.5 ng/mL, and the calibration curve was linear between 0.5 and 2000 ng/mL (R(2)>0.99). Within- and between-day coefficients of variation were less than 7%. This method has been successfully used to measure sildenafil plasma concentrations in a beagle dog model following an oral administration of the drug.

The purpose of the study was to characterize the changes in purine metabolism in long-distance runners in the main phases of their 1-year training cycle. Nine male athletes competing in distances 5 and 10 km at national/regional level, mean age 22.9 +/- 0.6 years, practising sport for 8.6 +/- 0.3 years, participated in the study. The changes in plasma concentrations of hypoxanthine (Hx), xanthine (X) and uric acid (UA) and the activity of the enzyme HGPRT in red blood cells haemolysate were followed in four characteristic points of the annual training cycle: preparatory phase (specific subphase), competition period, transition period and preparatory phase (intermediate subphase). Resting and postexercise plasma concentrations of X and, Hx and HGPRT activity changed significantly during 1-year training cycle. Significant changes in postexercise Hx values between training phases were found, from 9.3 micromol l(-1) in competition period to 22.9 micromol l(-1) in transition period (Friedmann's ANOVA, P < 0.01). Postexercise UA values ranged from 371 to 399 micromol l(-1) and did not change significantly between training phases. An increase in resting (from 52.0 to 58.4 IMP mg(-1) Hb min(-1), P < 0.05) and postexercise (from 70.7 to 76.2 IMP mg(-1) Hb min(-1), not significant) HGPRT activity between the specific preparation and competition period was observed. In the transition period, Hx postexercise concentration increased (22.9 micromol l(-1), P < 0.01) and HGPRT postexercise activity decreased (58.8 IMP mg(-1) Hb min(-1), P < 0.01) significantly. The results indicate that the level of plasma Hx at rest and after standard exercise may be a useful tool for monitoring the adaptation of energetic processes in different training phases and support the overload/overtraining diagnosis.

Objectives: In this study, the concentrations of uric acid, purine profile and creatinine in samples of cerebrospinal fluid and serum of multiple sclerosis (MS) patients were measured by HPLC and compared with corresponding values recorded in patients without MS (cerebrospinal fluid) and healthy subjects (serum)., Design and Methods: All samples were deproteinized with ultrafiltration (which ensures minimal sample manipulation and efficient protein removal) and then assayed for the synchronous HPLC separation of uric acid, hypoxanthine, xanthine, inosine, adenosine, guanosine and creatinine., Results: The values of all compounds assayed were significantly higher in both biological fluids of MS patients with respect to values measured in controls. In particular, serum hypoxanthine, xanthine, uric acid and sum of oxypurines were, respectively, 3.17, 3.11, 1.23 and 1.27-fold higher in these patients than corresponding values recorded in controls (p<0.001)., Conclusions: Differently from what previously reported, we here demonstrate that all purine compounds, including uric acid, are elevated in biological fluids of MS patients. Reinforced by the trend observed for creatinine, this corroborates the notion of sustained purine catabolism, possibly due to imbalance in ATP homeostasis, under these pathological conditions. These results cast doubt on the hypothesis that uric acid is depleted in MS because of increased oxidative stress, rather suggesting that this disease causes a generalized increase in purine catabolism. As observed in other pathological states, uric acid, purine compounds and creatinine, can be considered markers of metabolic energy imbalance rather than of reactive oxygen species, even in MS.

Purpose: To assess, in dogs with naturally occurring non-Hodgkin's lymphoma, pharmacokinetics, safety, and activity of GS-9219, a prodrug of the nucleotide analogue 9-(2-phosphonylmethoxyethyl) guanine (PMEG), which delivers PMEG and its phosphorylated metabolites to lymphoid cells with preferential cytotoxicity in cells with a high proliferation index such as lymphoid malignancies., Experimental Design: To generate proof-of-concept, a phase I/II trial was conducted in pet dogs (n = 38) with naturally occurring non-Hodgkin's lymphoma using different dose schedules of GS-9219. A subset of dogs was further evaluated with 3'-deoxy-3'-(18)F-fluorothymidine positron emission tomography/computed tomography imaging before and after treatment., Results: The prodrug had a short plasma half-life but yielded high and prolonged intracellular levels of the cytotoxic metabolite PMEG diphosphate in peripheral blood mononuclear cells in the absence of detectable plasma PMEG. Dose-limiting toxicities were generally manageable and reversible and included dermatopathy, neutropenia, and gastrointestinal signs. Antitumor responses were observed in 79% of dogs and occurred in previously untreated dogs and dogs with chemotherapy-refractory non-Hodgkin's lymphoma. The median remission durations observed compare favorably with other monotherapies in dogs with non-Hodgkin's lymphoma. High 3'-deoxy-3'-(18)F-fluorothymidine uptake noted in lymphoid tissues before treatment decreased significantly after treatment (P = 0.016)., Conclusions: GS-9219 was generally well tolerated and showed significant activity against spontaneous non-Hodgkin's lymphoma as modeled in pet dogs and, as such, supports clinical evaluation in humans.

The purpose of the present study was to determine sildenafil and a novel PDE-5 inhibitor, mirodenafil in the plasma and corpus cavernosum tissue of rats to compare their pharmacokinetic properties. The concentrations of mirodenafil and sildenafil in the rat plasma and corpus cavernosum tissue samples were analyzed using LC-MS/MS after a single oral administration at a dose of 40mg/kg to rats. Although the T(max), Tlambda(1/2) and MRT were not different between mirodenafil and sildenafil, the C(max) and AUC of mirodenafil were significantly higher than those of sildenafil in the plasma and corpus cavernosum tissue. Consequently mirodenafil remained longer than sildenafil in the plasma and tissue. This may provide pharmacokinetic evidence for assessment of the in vivo efficacy of mirodenafil and sildenafil.

Purpose: Cell cycle dysregulation resulting in expression of antiapoptotic genes and uncontrolled proliferation is a feature of undifferentiated nasopharyngeal carcinoma. The pharmacodynamic effects of seliciclib, a cyclin-dependent kinase (CDK) inhibitor, were studied in patients with nasopharyngeal carcinoma., Experimental Design: Patients with treatment-naïve locally advanced nasopharyngeal carcinoma received seliciclib at 800 mg or 400 mg twice daily on days 1 to 3 and 8 to 12. Paired tumor samples obtained at baseline and on day 13 were assessed by light microscopy, immunohistochemistry, and transcriptional profiling using real-time PCR low-density array consisting of a panel of 380 genes related to cell cycle inhibition, apoptosis, signal transduction, and cell proliferation., Results: At 800 mg bd, one patient experienced grade 3 liver toxicity and another had grade 2 vomiting; no significant toxicities were experienced in 13 patients treated at 400 mg bd. Seven of fourteen evaluable patients had clinical evidence of tumor reduction. Some of these responses were associated with increased tumor apoptosis, necrosis, and decreases in plasma EBV DNA posttreatment. Reduced protein expression of Mcl-1, cyclin D1, phosphorylated retinoblastoma protein pRB (T821), and significant transcriptional down-regulation of genes related to cellular proliferation and survival were shown in some patients posttreatment, indicative of cell cycle modulation by seliciclib, more specifically inhibition of cdk2/cyclin E, cdk7/cyclin H, and cdk9/cyclin T., Conclusions: Brief treatment with this regimen of seliciclib in patients with nasopharyngeal carcinoma is tolerable at 400 mg bd and associated with tumor pharmacodynamic changes consistent with cdk inhibition, and warrants further efficacy studies in this tumor.

Objectives: The assessment of the clinical significance of adenosine, adenine, inosine, xanthine, hypoxanthine and uric acid concentrations in patients with type 2 diabetes mellitus (T2DM) and diabetic nephropathy (DN) for the detection of the relationship between purine metabolites and disease., Design and Methods: The study group consisted of 119 subjects which were divided into three groups: control (n=31), type 2 diabetes without nephropathy (DM, n=23) and with nephropathy (DN, n=65). Levels of related metabolites were measured in plasma of all participants., Results: There is a significant increase of levels of adenosine (P<0.001), inosine (P<0.001), xanthine (P=0.012) and uric acid (P=0.016) with DN compared to DM. The level of xanthine oxidase (reflected by the uric acid: xanthine) did not change., Conclusion: The levels of adenosine, inosine, uric acid and xanthine may be useful for monitoring the progression of DM and evaluating the treatment.

The phosphodiesterase 5 inhibitor sildenafil is a potential therapeutic option in the treatment of persistent pulmonary hypertension of the newborn (PPHN) and neonatal hypoxemia. In this open-label trial, 36 term neonates with PPHN or hypoxemia were administered intravenous sildenafil for up to 7 days starting within 72 h of birth. A mixed-effects pharmacokinetic model that included two-compartment disposition of sildenafil and its metabolite and an effect of postnatal age on sildenafil clearance was used to describe plasma concentration-time data of parent and metabolite. Allometrically scaled sildenafil clearance increased threefold from the first day after birth to values similar to those in adults by the first week. Volume of distribution of sildenafil in neonates was fourfold higher than in adults, resulting in a longer terminal half-life in neonates (48-56 h) compared to adults. Increase in sildenafil clearance in the early postnatal period likely reflects maturation of CYP-mediated N-demethylation.

A simple, quick, accurate, and sensitive colorimetric method is described for the determination of sildenafil citrate (SLD). The method is based on the reaction of SLD with Congo Red, Sudan II, and Gentian Violet in buffered aqueous solutions at pH 2.5, 6.5, and 11.0, respectively, to give highly colored soluble ion-associate complex species; the colored products are quantitated colorimetrically at 523, 554, and 569 nm, respectively. The various experimental conditions were optimized. The stoichiometric ratio was found to be 1:1 for all ion associates; the calculated logarithmic stability constants were 8.51, 7.79, and 5.58, respectively. Beer's law was obeyed over the concentration range of 0.2-7.0 microg/mL, whereas the Ringbom optimum concentration range was 0.4-6.5 microg/mL. Values for molar absorptivity, Sandell sensitivity, and detection and quantification limits were also calculated. The proposed method was successfully applied to the determination of SLD in Viagra tablets and in serum samples by using the technique of standard additions with mean accuracy values of 100.06 +/- 1.14, 99.87 +/- 0.70, and 99.86 +/- 0.97% for Viagra tablets and 99.88 +/- 0.60, 99.90 +/- 0.90, and 100.24 +/- 0.80% for serum samples, respectively.

A simple, sensitive and specific liquid chromatography tandem mass spectrometry method with minimal sample pretreatment was developed for the simultaneous analysis of sildenafil and its metabolite desmethylsildenafil in human serum. Sample pretreatment consisted of adding a methanolic solution of the internal standard vardenafil to the samples. After vortexing and centrifugation the samples were directly injected onto the C18 column using gradient elution. The aqueous and organic mobile phases were ammonium acetate 2 mM supplemented with 0.1% formic acid in water and methanol, respectively. The detection by a triple quadrupole mass spectrometer in positive ESI ionization mode was completed within 5 min. The lower limits of quantification for sildenafil and desmethylsildenafil are 1.0 ng/ml. The intra- and inter-day precisions measured as relative standard deviation were within 10% for both compounds over the linear range. Intra- and inter-day accuracy of sildenafil and desmethylsildenafil ranged from 92 to 103%. This method has been used in a clinical pharmacokinetic study of sildenafil in intensive care patients.

The pharmacokinetic interaction between sildenafil, a phosphodiesterase type 5 (PDE-5) inhibitor, and ambrisentan, an ET(A)-selective, propanoic acid-based endothelin receptor antagonist (ERA), was studied in a 2-period crossover study in 19 healthy volunteers, with ambrisentan exposure (AUC(0-infinity)) and maximum plasma concentration (C(max)) determined over 24 hours for a 10-mg dose of ambrisentan alone and again after 7 days of sildenafil 20 mg 3 times daily. The AUC(0-infinity) and C(max) for sildenafil and N-desmethyl sildenafil (active metabolite) were determined over 24 hours for a 20-mg dose of sildenafil alone and again after 7 days of dosing with ambrisentan 10 mg once daily. There was no clinically relevant pharmacokinetic interaction between ambrisentan and sildenafil or N-desmethyl sildenafil. Ambrisentan C(max) was unchanged (96.3% [90% confidence interval: 86.0%-107.8%]), with a minor increase in AUC(0-infinity) (108.5% [102.6%-111.7%]) with sildenafil coadministration. Sildenafil C(max) was increased slightly (113.4% [99.6%-129.1%]), and AUC(0-infinity) was unchanged (98.7% [91.2%-110.5%]) with ambrisentan coadministration. N-desmethyl sildenafil was unaltered. Dose adjustment of either drug is not necessary compared with administration alone.

Aim: To investigate if sildenafil increases splanchnic blood flow and changes the hepatic venous pressure gradient (HVPG) in patients with cirrhosis. Phosphodiesterase type-5 inhibitors are valuable in the treatment of erectile dysfunction and pulmonary hypertension in patients with end-stage liver disease. However, the effect of phosphodiesterase type-5 inhibitors on splanchnic blood flow and portal hypertension remains essentially unknown., Methods: Ten patients with biopsy proven cirrhosis (five females/five males, mean age 54 +/- 8 years) and an HVPG above 12 mmHg were studied after informed consent. Measurement of splanchnic blood flow and the HVPG during liver vein catheterization were done before and 80 min after oral administration of 50 mg sildenafil. Blood flow was estimated by use of indocyanine green clearance technique and Fick's principle, with correction for non-steady state., Results: The plasma concentration of sildenafil was 222 +/- 136 ng/mL 80 min after administration. Mean arterial blood pressure decreased from 77 +/- 7 mmHg to 66 +/- 12 mmHg, P = 0.003, while the splanchnic blood flow and oxygen consumption remained unchanged at 1.14 +/- 0.71 L/min and 2.3 +/- 0.6 mmol/min, respectively. Also the HVPG remained unchanged (18 +/- 2 mmHg vs 16 +/- 2 mmHg) with individual changes ranging from -8 mmHg to +2 mmHg. In seven patients, HVPG decreased and in three it increased., Conclusion: In spite of arterial blood pressure decreases 80 min after administration of the phosphodiesterase type-5 inhibitor sildenafil, the present study could not demonstrate any clinical relevant influence on splanichnic blood flow, oxygen consumption or the HVPG.

A simple HPLC method has been validated for the determination of sildenafil and its active metabolite (N-desmethylsildenafil) in human blood, using an octadecyl silica (ODS) hypersil column. The chromatographic run time is less than 25 min using a mobile phase of 35:65 (v/v) acetonitrile-0.015 M disodium hydrogen phosphate (Na(2)HPO(4)), triethylamine 0.1%, pH 7.4 at 1 mL/min flow rate and UV-vis detection at 230 nm. The method is linear in the concentration range of 10-500 ng/mL (r>0.999, n=5) for each analyte, with relative standard deviation (R.S.D.) less than 5.05%. Interday and intraday errors were found to be < or =11.94%. The limits of detection and quantitation for both analytes were 5.0 ng/mL (s/n>3) and 10.0ng/mL (s/n>10), respectively. The method was applied in two post-mortem human blood samples, concerning two fatal cases from sildenafil citrate use, reported for the first time in Greece, and the results were further confirmed with LC-MS. The method is proposed as supplementary to LC-MS when inadequate mass fragmentation does not provide information appropriate to meet confirmation criteria.

Objective: To examine the effect of mode of birth on plasma purine and malondialdehyde levels in normal term infants., Study Design: Umbilical arterial cord blood was obtained immediately after birth from a convenience sample of 119 normal term newborns born by vaginal delivery, with or without oxytocin augmentation or by elective cesarean delivery. Plasma was analyzed for purine and/or malondialdehyde levels. Numeric data were analyzed utilizing independent samples t-test and ordinal data were analyzed using Mann-Whitney test. Correlation coefficients were obtained using Spearman's rho., Result: Uric acid levels were significantly elevated (P<0.001) in neonates undergoing vaginal birth, compared to neonates born by elective cesarean delivery. When the effect of oxytocin and length of labor was analyzed, neonates born to mothers on oxytocin had lower hypoxanthine, significantly lower xanthine (P=0.05) and higher uric acid levels. In addition, malondialdehyde levels were significantly higher (P<0.006) in neonates born to mothers who received oxytocin compared to neonates born to mothers without oxytocin augmentation. We also found significant correlations between malondialdehyde (MDA) and hypoxanthine (r=-0.465, P<0.039) and between MDA and xanthine (r=-0.753, P=0.003) in neonates born via oxytocin-augmented birth. Mode of birth had no statistically significant effect on clinical outcomes, although infants born by elective cesarean had higher incidence of acute respiratory distress and transient tachypnea of the newborn compared to those born vaginally., Conclusion: Neonates born by elective cesarean had the lowest purine levels in cord blood compared to neonates born vaginally. Oxytocin augmentation is associated with some degree of uterine hyperstimulation which may enhance the ATP degradation pathway resulting in the rapid conversion of hypoxanthine to xanthine and xanthine to uric acid. Significantly higher MDA levels in neonates whose mothers received oxytocin as well as significant correlation between MDA and the purines hypoxanthine and xanthine, suggest free-radical production, most likely due to xanthine oxidase activation. However, despite differences in plasma purine and malondialdehyde levels, no significant differences were seen in neonatal outcome. Further studies are required to fully characterize the effect of mode of birth on purine metabolism and free-radical production.

Roscovitine is a cyclin-dependent kinase (Cdk) and signal-regulated kinase (Erk1/2) inhibitor that has been shown to be effective against several cancer types including brain tumors. We have shown previously that roscovitine crosses the blood brain barrier (BBB) and is rapidly eliminated from both plasma and brain in adult rats. However, age-dependent kinetics and its effects on the brain have not been reported. In the present study, we investigated the pharmacokinetics of roscovitine in adult and in 14 days old rats after the administration of a single dose of 25 mg/kg. Moreover, we studied the effect of the drug on Cdk5 and Erk1/2 activities in three brain regions, hippocampus, frontal cortex and cerebellum. The pharmacokinetics of roscovitine followed a two-compartment model in both plasma and brain in both adult and young rats. The terminal elimination half-life was 7 h in brain as well as in plasma in rat pups compared to < 0.5 h observed in adult rats. Brain exposure expressed as AUC brain/AUC plasma was 100% in rat pups compared to 20% found in adult rats. Roscovitine induced a significant Cdk5 inhibition and significant Erk1/2 activation in all studied pups brain regions at 2 h. This is the first study describing age-dependent pharmacokinetics of roscovitine and showing the high brain exposure of infant rats to the drug. Thus, roscovitine may be a promising candidate for the treatment of brain tumors in children.

Female carriers of hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency have somatic cell mosaicism of HPRT activity and are healthy. We report a 50-year-old woman without gout or nephrolithiasis. She was never on allopurinol. Normal serum uric acid concentrations, increased plasma hypoxanthine, and xanthine were found. HPRT activity in erythrocytes was surprisingly low: at 8.6 nmol h(-1) mg (-1) haemoglobin. Mutation analysis revealed a heterozygous HPRT gene mutation, c.215A > G (p.Tyr72Cys). Assessment of X-inactivation ratio has shown that > 75% of the active X-chromosome bears the mutant allele and could explain these unusual, previously undescribed findings.

Sucrose is converted fructose and glucose, which may increase plasma uric acid concentration (pUA) through increased purine degradation and/or decreased uric acid (UA) excretion. To investigate effects of acarbose, an inhibitor of alpha-glucosidase, on the increased pUA from sucrose administration, we measured pUA and urinary UA excretion in 6 healthy subjects before and after administering sucrose, with and without co-administration of acarbose. Sucrose raised pUA by 10% (p < 0.01). However, excretion and fractional clearance of UA were unchanged. Sucrose and acarbose coadministration also increased pUA, but less than did sucrose alone (sucrose: 4.9 to 5.4 mg/dl; sucrose + acarbose, 4.7 to 4.9 mg/dl, p < 0.05) without changes in urinary excretion and fractional clearance of UA. Acarbose appears to attenuate the rise in pUA by sucrose ingestion by inhibiting sucrose absorption.

We investigated the effects of allopurinol on beer-induced changes in the plasma concentration and urinary excretion of purine bases. Five healthy subjects underwent three studies: ingestion of beer after taking 300 mg allopurinol (combination study); ingestion of beer alone; ingestion of allopurinol alone. Increased plasma concentrations and urinary excretion of hypoxanthine were greater in the combination study than the beer alone study. However, increases in total plasma purine base concentrations were greater in the beer alone study, even though increases in plasma uridine concentrations did not differ. Beer-induced increases in plasma concentrations of purine bases appear partially offset by increased urinary excretion of hypoxanthine after allopurinol, which also controls increases in plasma uric acid levels caused by alcoholic beverage ingestion.

Whole body exposure of wild type control littermates and A2A adenosine receptor (A2AR) gene deleted mice to low oxygen containing inspired gas mixture allowed the investigation of the mechanism that controls inflammatory liver damage and protects the liver using a mouse model of T cell-mediated viral and autoimmune hepatitis. We tested the hypothesis that the inflammatory tissue damage-associated hypoxia and extracellular adenosine --> A2AR signaling plays an important role in the physiological anti-inflammatory mechanism that limits liver damage during fulminant hepatitis. After induction of T cell-mediated hepatitis, mice were kept in modular chambers either under normoxic (21% oxygen) or hypoxic (10% oxygen) conditions for 8 h. It was shown that the whole body exposure to hypoxic atmosphere caused tissue hypoxia in healthy animals as evidenced by a decrease in the arterial blood oxygen tension and increase of the plasma adenosine concentration (P < 0.05). This "hypoxic" treatment resulted in significantly reduced hepatocellular damage and attenuated levels of serum cytokines in mice with acute liver inflammation. The anti-inflammatory effects of hypoxia were not observed in the absence of A2AR in studies of A2AR gene-deficient mice or when A2AR have been pharmacologically antagonized with synthetic antagonist. The presented data demonstrate that total body hypoxia-triggered pathway provides protection in acute hepatitis and that hypoxia (upstream) and A2AR (downstream) function in the same immunosuppressive and liver tissue-protecting pathway.

Background: Since grape juice contains considerable amounts of fructose, which may increase the plasma concentration of urate, the combination of exercise and grape juice may increase the plasma concentration of urate to a greater degree than grape juice or exercise alone., Methods: We performed 3 experiments with 6 healthy male Japanese. The first was exercise alone (exercise alone experiment), the second was grape juice ingestion alone (grape juice alone experiment), and the third was a combination of exercise and grape juice ingestion (combination experiment)., Results: In the exercise alone experiment, the concentrations of purine bases and uridine in plasma, and lactate in blood, as well as the urinary excretion of oxypurines were increased, whereas the urinary excretion of uric acid and fractional excretion of purine bases were decreased. In the grape juice alone experiment, the concentrations of purine bases and uridine, as well as lactate in blood were increased, whereas the fractional excretion of uric acid was decreased. In the combination experiment, the concentrations of purine bases and uridine in plasma, and lactate in blood, as well as the urinary excretion of oxypurines were increased, whereas the urinary excretion of uric acid and fractional excretion of hypoxanthine, xanthine, and uric acid were decreased. The increase in plasma concentration of urate by the combination of exercise and grape juice was greater than that by each alone, though it was not significantly different from the sum of increases in those 2 experiments., Conclusion: Increases in adenine nucleotide degradation and lactic acid production caused by both exercise and grape juice ingestion play an important role in the increase in plasma concentration of urate, while those in combination have an additive effect on that concentration.

Background and Objectives: Darunavir (DRV, TMC114) is a novel protease inhibitor administered in combination with low-dose ritonavir (DRV/r) and is highly active against both wild-type and multidrug-resistant HIV-1 strains. Sildenafil is an oral therapy for erectile dysfunction. Concomitant administration of protease inhibitors and sildenafil increases sildenafil plasma concentrations. The potential for a pharmacokinetic drug interaction exists when sildenafil and DRV/r are co-administered, as these drugs are primarily metabolized by cytochrome P450 (CYP) 3A, and darunavir and ritonavir are CYP3A inhibitors. The primary objective of this open-label, crossover, phase I study was to assess the effect of multiple doses of DRV/r on the pharmacokinetics of sildenafil and its active metabolite N-desmethyl sildenafil. The secondary objective was to assess the short-term safety and tolerability of co-administration of sildenafil and DRV/r., Methods: Sixteen HIV-negative healthy male subjects were randomized to one of two sequences. In two sessions each subject received treatments A and B. In treatment A, a single dose of sildenafil 100 mg was administered. In treatment B, the subjects received DRV/r 400/100 mg twice daily for 8 days and on day 7 a single dose of sildenafil 25 mg was co-administered. Full pharmacokinetic profiles of sildenafil, N-desmethyl sildenafil, darunavir and ritonavir were determined. Safety and tolerability were also assessed., Results: Sildenafil exposure (area under the plasma concentration-time curve [AUC]) was comparable between the two treatments despite administration of a lower dose of sildenafil (25 mg) with DRV/r than when sildenafil (100 mg) was administered alone. When sildenafil 25 mg was co-administered with DRV/r, the sildenafil maximum plasma concentration (Cmax) was 38% lower compared with Cmax after administration of sildenafil alone at a dose of 100 mg. N-desmethyl sildenafil Cmax and AUC from the time of administration until the last time point with a measurable concentration after dosing (calculated by linear trapezoidal summation [AUClast]) values decreased by approximately 95% when sildenafil 25 mg was co-administered with DRV/r compared with sildenafil 100 mg alone. Combined treatment with DRV/r and sildenafil was generally safe and well tolerated., Conclusion: Sildenafil exposure is increased in the presence of DRV/r. In this setting, a dose adjustment for sildenafil is warranted; no more than 25 mg of sildenafil is recommended over a 48-hour period when co-administered with DRV/r.

A rapid, sensitive and reproducible HPLC method was developed and validated for the analysis of pantoprazole (CAS 102625-70-7) in human plasma. The separation was achieved on a monolithic silica column using acetonitrile-potassium dihydrogen phosphate buffer (25:75,v/v), pH 3.0, as the mobile phase at a flow rate of 1.5 ml min(-1). The wavelength was set at 290 nm. The assay enables the measurement of pantoprazole for therapeutic drug monitoring with a minimum quantification limit of 20 ng ml(-1). The method involves a simple protein precipitation procedure. Analyticil recovery was complete. The calibration curve was linear over the concentration range 20-3500 ng ml(-1) The coefficients of variation forthe inter-day and intra-day assay were found to be less than 7%.

Background: Sildenafil citrate (Viagra) is a selective inhibitor of cyclic guanosine monophosphate (cGMP)-specific phosphodiesterase type 5, which might enhance the vasorelaxant and natriuretic actions of atrial natriuretic peptide (ANP) in patients with heart failure. The objective of this study was to examine the combined effect of Viagra on hemodynamic changes during infusion of exogenous ANP., Methods and Results: Healthy male beagles were used to assess systemic blood pressure, pulmonary artery pressure (PAP), and plasma levels of cGMP. After hemodynamic variables were measured, 0.1 microg.kg(-1).min(-1) of ANP was given during this study. One hour after initiating infusion of ANP, 2 mg/kg of sildenafil citrate or vehicle was given orally via a nasogastric tube. Hemodynamic changes were measured before and 1 h after these administrations. Mean systemic and PAP decreased during infusion of ANP, and further decreased after sildenafil citrate administration, however, mean systemic blood pressure decreased within 10 mmHg. Plasma levels of cGMP also increased after sildenafil citrate administration., Conclusion: In normal dogs, sildenafil citrate enhances the vasodilator effect of ANP by increasing the cGMP level, however, the concomitant use of sildenafil citrate with ANP will not induce severe hypotension.

Introduction: Little is known about the toxicity of sildenafil overdose in the pediatric population. We present a case of prolonged priapism and tachycardia due to unintentional sildenafil overdose in a child., Case Report: A 19-month-old male ingested up to six 50 mg Viagra tablets 45 minutes prior to presentation at the emergency department. Initial vital signs included temperature 98.2 degrees F, blood pressure 90/58 mmHg, heart rate 140, respirations 20, and oxygen saturation of 100% on room air. The child weighed 10.4 kg. Physical exam revealed a happy, smiling, laughing toddler who was cooperative with all aspects of his exam. He had mild facial flushing and an erect penis which was normal in color and had a capillary refill of two seconds. Precordial palpation did not show evidence of increase dynamic force, his heart sounds were regular, and no ectopy was noted. His peripheral pulses were strong and regular in all four extremities. No gastrointestinal decontamination was performed. The patient was started on maintenance IV fluids and admitted to the pediatric floor for observation. The patient had a non-painful tumescent penis and mild tachycardia for about 24 hours post-ingestion. The child never had pain from the constant erection. Sildenafil concentration drawn approximately seven hours after ingestion was 3900 ng/ml (reporting limit 24 ng/ml) and N-desmethylsildenafil level was 1700 ng/ml (reporting limit 24 ng/ml)., Conclusion: This case of pediatric sildenafil ingestion (up to 30 mg/kg) initially resulted in facial flushing and priapism. The child had asymptomatic tachycardia and prolonged priapism that persisted until hospital discharge approximately 24 hours after ingestion. The erection was non-painful and required no urologic intervention, most likely representing a high flow state. In this ingestion, only supportive care was required.

The authors have investigated the pharmacokinetics and tolerability of regadenoson, a selective A2A adenosine receptor agonist for use in drug-stressed myocardial perfusion imaging in subjects with varying degrees of renal function. Sixteen subjects with different creatinine clearance values (range: 15-132 mL/min) received a single intravenous bolus dose of 400 microg regadenoson. A population pharmacokinetic model was developed to describe the pharmacokinetics of regadenoson in these subjects. Regadenoson elimination half-life was prolonged with decreasing renal function. However, maximum plasma concentrations, number, or severity of adverse events did not differ significantly between the subjects. Heart rate increased in all subjects after regadenoson injection but returned to normal within 150 minutes. There were no blood pressure pattern differences with respect to renal function. Results from this study do not indicate that dose adjustments are necessary when subjects with decreased renal function are administered the clinically relevant dose of 400 microg regadenoson.

The evaluation of molecularly imprinted polymers (MIPs) as selective sorbents for the solid-phase extraction of sildenafil and its principal metabolite, desmethylsildenafil, was investigated. Two MIPs were synthesised using structural analogues of sildenafil as templates, and a comparison of the performance of the two MIP sorbents in organic and aqueous media was performed. Additionally, the feasibility of applying molecularly imprinted solid-phase extraction (MISPE) to the clean-up of plasma samples containing sildenafil and desmethylsildenafil was investigated. A preliminary, quantitative MISPE for the determination of both compounds in plasma was also performed. The results showed that the MIPs used for the selective extraction of sildenafil gave better compound recovery when aqueous samples were used in comparison to organic-based samples. A preliminary, quantitative MISPE of sildenafil and desmethylsildenafil indicated that the imprinted materials could be used successfully as SPE sorbents for sample pre-treatment for the determination of sildenafil, and related compounds, in plasma.

Effects of caffeine on regadenoson-induced coronary vasodilation and changes in hemodynamics were examined in conscious dogs. Sixteen dogs were chronically instrumented for measurements of coronary blood flow (CBF), mean arterial pressure (MAP), and heart rate (HR). Regadenoson (5 microg/kg, IV) increased CBF from 34 +/- 2 to 191 +/- 7 mL/min. The duration of the 2-fold increase in CBF was 515 +/- 71 seconds. Regadenoson decreased MAP by 15 +/- 2% and increased HR by 114 +/- 14%. Regadenoson-induced maximum increases in CBF were not significantly lower in the presence of caffeine at 1, 2, 4, and 10 mg/kg (2 +/- 3, 0.7 +/- 3, 16 +/- 5, and 13 +/- 8%, respectively; all P > 0.05). Caffeine at 1, 2, 4, and 10 mg/kg significantly decreased the duration of the 2-fold increase in CBF induced by regadenoson by 17% +/- 4%, 48% +/- 8%, 62% +/- 5%, and 82% +/- 5%, respectively (all P < 0.05). Caffeine at 4 and 10 mg/kg significantly attenuated the effects of regadenoson on MAP and HR. The results indicate that 1 to 10 mg/kg caffeine dose-dependently reduced the duration, but not the peak increase of CBF caused by 5 microg/kg regadenoson.

To determine whether an increase in the plasma concentration of uric acid by sucrose intake is ascribable to enhanced purine degradation and/or decreased urinary excretion of uric acid, we measured the plasma concentrations of purine bases (uric acid, hypoxanthine, and xanthine) and uridine, as well as the urinary excretion of purine bases in 7 healthy subjects before and after administering sucrose at 1.5 g/kg of body weight in 2 related experiments, with and without an administration of 300 mg of allopurinol. In addition, in the control experiment without an administration of sugar and with an administration of 300 mg of allopurinol, we measured the same parameters in those 7 subjects. Without added allopurinol, sucrose increased the plasma concentration of uric acid by 11% (P<.01) as well as that of uridine, although it did not significantly increase the plasma concentrations of hypoxanthine and xanthine or the urinary excretion of uric acid. On the other hand, the plasma concentration and urinary excretion of hypoxanthine were increased by 2.4-fold (P<.05) and 3.42-fold (P<.05), respectively, and the plasma concentration of xanthine was increased by 1.2-fold (P<.05) together with an increase in the plasma concentration of uridine in the experiment with allopurinol administration. In contrast, the plasma concentration and urinary excretion of uric acid and the urinary excretion of xanthine were not increased. In addition, in the control experiment, all parameters did not change significantly. These results indicate that purine degradation enhanced by sucrose plays a major role in the increased plasma concentration of uric acid.

A method which employed high-performance liquid chromatography coupled with electrochemical detection was developed for the simultaneous determination of sildenafil and its metabolite, N-desmethyl sildenafil, in human plasma has. The method was developed and validated for purposes of its application to a pharmacokinetic study in healthy volunteers after an oral dose of 50mg/tablet under fasting conditions. High precision and accuracy were demonstrated. A one-step liquid-liquid extraction further provides a simple and practical way to process plasma samples containing sildenafil with good quantitative recovery. Sampling lasted for 24h after dosing; consequently a limit of quantitation (LOQ) of 7.858 ng/mL was achieved for sildenafil whereas a LOQ of 8.675 ng/mL was obtained for N-desmethyl sildenafil. The mobile phase consisted of acetonitrile, methanol and phosphate buffer (0.05 M) (18.5:34.5:47.0, v/v/v) pH 7.68. The stationary phase was a C(8) (150 mm x 4.6 mm), 5 microm particle size operated at 27 degrees C. All analytes were stable at the pH of the supernatant, and during the analytical time window. At the applied potential of +1.20 V versus Ag/AgCl, no interferences from endogenous plasma compounds were recorded at the retention times of sildenafil, N-desmethyl sildenafil. High resolution was obtained between the analytes and the employed internal standards.

This review summarizes currently available information about a crucial part of erythrocyte metabolism, that is, purine nucleotide conversions and their relationships with other conversion pathways. We describe the cellular resynthesis, interconversion, and degradation of purine compounds, and also the regulatory mechanisms in the conversion pathways. We also mention purine metabolism disorders and their clinical consequences. The literature is fragmentary because studies have concentrated only on selected aspects of purine metabolism; hence the need for a synthetic approach.

HPLC method for the determination of sildenafil and its metabolite (N-demethylsildenafil) in rat serum has been developed. The technique included a solid phase extraction of the serum samples on a [poly(divinylbenzene-co-N-vinylpyrrolidone)] solid phase extraction sorbent. After conditioning, the cartridge was loaded with 0.5 mL of buffered serum containing internal standard. Elution was made with 1 mL of acetonitrile. After evaporation of the eluates to dryness and reconstitution with methanol, the samples were analyzed on Kromasil C18 column phase with phosphate buffer 0.05 M/acetonitrile: 54/46, pH 8. Detection was carried out using a photodiode array detector. For sildenafil and demethylsildenafil, full validation of the proposed method was provided (linearity range, calibration curves, average extraction efficiency; average intra-day and interday variabilities, limit of detection, limit of quantification, specificity). The proposed method was successfully utilised to quantify sildenafil and N-demethylsidenafil in rat serum for a pharmacokinetic study.

To determine the effects of allopurinol on beer-induced increases in plasma and urinary excretion of purine bases (hypoxanthine, xanthine, and uric acid), we performed three experiments on five healthy study participants. In the first experiment (combination study), the participants ingested beer (10 ml/kg body weight) eleven hours after taking allopurinol (300 mg). In the second experiment (beer-only study), the same participants ingested beer (10 ml/kg body weight) alone, while in the third experiment (allopurinol-only study), they took allopurinol (300 mg) alone. There was a two-week interval between each of the studies. Beer-induced increases in plasma concentration and urinary excretion of hypoxanthine in the combination study were markedly higher than those in the beer-only study. On the other hand, the sum of increases in plasma concentrations of purine bases in the beer-only study was greater than in the combination study, whereas the increase in plasma uridine concentration in the combination study did not differ from the beer-only study. In addition, allopurinol administration inhibited the beer-induced increase in plasma concentration of uric acid. These results suggest that abrupt adenine nucleotide degradation may increase plasma concentration and urinary excretion of hypoxanthine under conditions of low xanthine dehydrogenase activity, which is mostly ascribable to allopurinol. Further, the difference in the sum of increases in plasma concentrations of purine bases between the combination study and beer-only study was largely ascribable to a greater increase in urinary excretion of hypoxanthine in the combination study. In addition, allopurinol intake seems to be effective in controlling the rapid increase in plasma uric acid caused by ingestion of alcoholic beverages.

To investigate the effect of long-term beer ingestion on the plasma concentrations and urinary excretion of purine bases, 5 healthy males participated in the present study, during which they ingested beer every evening for 30 days. Blood and 24-hour urine samples were collected in the morning one day before and 14 and 30 days after the initiation of the beer ingestion. During the beer ingestion period, the plasma concentration and the urinary excretion of uric acid were increased significantly, while uric acid clearance was not decreased. Further, purine ingestion was not significantly different throughout the study. These results suggest that production of uric acid by ethanol ingestion was the main contributor to the increased plasma uric acid. Therefore, patients with gout should be encouraged to avoid drinking large amounts of beer on a daily basis.

To investigate the effects of exercise on the plasma concentrations and urinary excretion of purine bases and oxypurinol, we performed 3 experiments with 6 healthy male subjects. The first was a combination of allopurinol intake (300 mg) and exercise (VO2max, 70%) (combination experiment), the second was exercise alone (exercise-alone experiment), and the third was allopurinol intake alone (allopurinol-alone experiment). In the combination experiment, exercise increased the concentrations of purine bases and noradrenaline in plasma, as well as lactic acid in blood and the urinary excretion of oxypurines, whereas it decreased the urinary excretion of uric acid and oxypurinol as well as the fractional excretion of hypoxanthine, xanthine, uric acid, and oxypurinol. In the exercise-alone experiment, exercise increased the concentrations of purine bases and noradrenaline in plasma, lactic acid in blood, and the urinary excretion of oxypurines, whereas it decreased the urinary excretion of uric acid and fractional excretion of purine bases. In contrast, in the allopurinol-alone experiment, the plasma concentration, urinary excretion, and fractional excretion of purine bases and oxypurinol remained unchanged. These results suggest that increases in adenine nucleotide degradation and lactic acid production, as well as a release of noradrenaline caused by exercise, contribute to increases in plasma concentration and urinary excretion of oxypurines and plasma concentration of urate, as well as decreases in urinary excretion of uric acid and oxypurinol, along with fractional excretion of uric acid, oxypurinol, and xanthine. In addition, they suggest that oxypurinol does not significantly inhibit the exercise-induced increase in plasma concentration of urate.