Effect of oral organic nitrates on expression and activity of vascular soluble guanylyl cyclase.

BACKGROUND AND PURPOSE: The regulation of vascular soluble guanylyl cyclase (sGC) expression by nitric oxide (NO) is still under discussion. In vitro, NO has been shown to downregulate the expression of sGC but it is unclear if this mechanism is operative in vivo and occurs during nitrate treatment. EXPERIMENTAL APPROACH: We investigated whether high dose isosorbide mononitrate (ISMN) or pentaerythrityl tetranitrate (PETN) treatment changes vascular sGC expression and activity in vivo. New Zealand White rabbits received a standard diet, 2 or 200 mg ISMN kg(-1) d(-1) for 16 weeks, and C57BL/6 mice received a standard diet, 6, 60 or 300 mg PETN kg(-1) d(-1) for four weeks. Absorption was checked by measuring the plasma levels of the drug/metabolite. KEY RESULTS: Western blots of rabbit aortic rings showed similar protein levels of sGC alpha1- (P=0.2790) and beta1-subunits (P=0.6900) in all groups. Likewise, ANOVA showed that there was no difference in the expression of sGC in lungs of PETN-treated mice (P=0.0961 for alpha1 and P=0.3709 for beta1). The activities of isolated sGC in response to SNAP (1 microM-1 mM) were identical in aortae of ISMN-treated rabbits (P=0.0775) and lungs of PETN-treated mice (P=0.6348). The aortic relaxation response to SNAP slightly decreased at high ISMN but not at high PETN. CONCLUSIONS AND IMPLICATIONS: These data refute the hypothesis that therapeutic treatment with long acting NO donors has a significant impact on the regulation of vascular sGC expression and activity in vivo.

Potency and in vitro tolerance of organic nitrates: partially denitrated metabolites contribute to the tolerance-devoid activity of pentaerythrityl tetranitrate.

Neither therapeutic dosage of nitrovasodilators nor the development of tolerance correlates with nitrate groups in these molecules. Clinically, low dosages of glyceryl trinitrate (GTN) develop tolerance, but 100-fold higher dosages of pentaerythrityl tetranitrate (PETN) do not. Vasorelaxation was studied on prostaglandian F2alpha (PGF2alpha)-precontracted porcine pulmonary arteries in organ bath procedure. In vitro tolerance was induced by incubating the arteries with different nitrate concentrations and thereafter concentration-response curves were repeated. Furthermore, 14 mg/kg PETN were daily administered to rats by gavage; PETN and metabolites were measured in feces and blood. In vitro, the vasodilator potencies increased from mononitrates to tetranitrates (pD2: 4.14 to 8.18); PETN was the most potent vasodilator. In vitro tolerance was found with PETN and trinitrates but not with dinitrates and mononitrates. Thus, in vitro tolerance correlated with the in vitro potency of nitrates but not with the vasodilator potency of NO donors in general, because S-nitroso-N-aectyl-D-penicillamine and N-phenylpiperazin-NONOate were more potent than GTN but did not induce tolerance. After feeding of rats with PETN, pentaerythrityl dinitrate (PEdiN) and mononitrate (PEmonoN) but neither PETN nor PEtriN (both detected in feces) were found in the blood. The missing systemic bioavailability of PETN and PEtriN may explain the discrepancy between in vitro and in vivo findings. We conclude that the partially denitrated metabolites PEdiN and PEmonoN contribute to the moderate and tolerance-devoid clinical activity of PETN.

[Quantitative gas chromatography-mass spectrometry determination of pentaerythrityl tetranitrate metabolites pentaerythrityl trinitrate, pentaerythrityl dinitrate and pentaerythrityl in human plasma]. / Quantitative gaschromatographische/massenspektrometrische Bestimmung der Pentaerithrityltetranitrat-Metaboliten Pentaerithrityltrinitrat, Pentaerithrityldinitrat und Pentaerithritylmononitrat in Humanplasma.

Assay methods based on gas chromatography/mass spectroscopy (GC-MS) may be used to measure PE1N (pentaerithrityl mononitrate, CAS 1607-00-7), PE2N (pentaerithrityl dinitrate, CAS 1607-01-8) and intermediate pentaerithrityltrinitrate (PE3N, CAS 1607-17-6) in human plasma. Based on this method a simplified method to quantify the metabolites of PETN (pentaerithrityl tetranitrate, CAS 78-11-5, Pentalong) is described. In the present study a consistent method to extract the metabolites of human plasma and following derivatisation is described. Since PE1N can be quantified up to 150 ng/ml, PE2N and PE3N up to 30 ng/ml in human plasma, a dilution of the plasma samples can be avoided. The mean recovery rate is not so high as in other described methods, and inaccuracy is about 10%. Therefore a calibration range between 0.2-30 ng/ml of PE2N and 1-150 ng/ml of PE1N has to be considered. The described method offers an alternative and applicable option to quantify the PETN-metabolites and elucidate their part as NO-donors.

[Actions of pentaerithritol tetranitrate, isosorbide mononitrate and placebo on headache and ability to work of healthy subjects]. / Wirkungen von Pentaerithrityltetranitrat, Isosorbidmononitrat und Plazebo auf den Kopfschmerz und auf die Beeinträchtigung der Arbeitsfähigkeit gesunder Probanden.

In a randomised, double-blind, four-way crossover study, 24 healthy volunteers received 240 mg/d pentaerithritol tetranitrate (PETN, CAS 78-11-5), 150 mg/d PETN, 60 mg/d isosorbide mononitrate slow release (ISMN, CAS 16051-77-7) or placebo in each study period for two days. Headache and disability to work were self-rated six times per day; individual measurements were combined to total scores. ISMN caused headaches more frequently (in approx. 90% of volunteers) and more severe (average total score 15.2) and a greater disability (average total score 6.0) than the high or low PETN-dosage (both in approx. 50%, headache score 4.9 or 6.4, disability score 1.1 or 2.1, resp.) and placebo (in approx. 10%, headache 0.8, disability 0), all these differences were statistically significant (p < 0.01, Wilcoxon). The high PETN-dosage showed a non-significant trend to produce fewer systemic side effects than the low PETN-dosage (not vice versa). With ISMN six volunteers prematurely terminated the study period and one volunteer who was replaced withdrew from the entire study due to side effects; all volunteers completed the study periods with the other medications.

Clinical pharmacokinetics of vasodilators. Part II.

Stimulating cardiac beta 1-adrenoceptors with oxyfedrine causes dilatation of coronary vessels and positive inotropic effects on the myocardium. beta 1-adrenergic agonists increase coronary blood flow in nonstenotic and stenotic vessels. The main indication for the use of the phosphodiesterase inhibitors pamrinone, mirinone, enoximone and piroximone is acute treatment of severe congestive heart failure. Theophylline is indicated for the treatment of asthma, chronic obstructive pulmonary disease, apnea in preterm infants ans sleep apnea syndrome. Severe arterial occlusive disease associated with atherosclerosis can be beneficially affected by elcosanoids. These drugs must be administered parenterally and have a half-life of only a few minutes. Sublingual or buccal preparations of nitrates are the only prompt method (within 1 or 2 min) of terminating anginal pain, except for biting nifedipine capsules. The short half-life (about 2.5 min) of nitroglycerin (glyceryl trinitrate) makes long term therapy impossible. Tolerance is a problem encountered with longer-acting nitric oxide donors. Knowledge of the pharmacokinetic properties of vasodilating drugs can prevent a too sudden and severe blood pressure decrease in patients with chronic hypertension. In considering the administration of a second dose, or another drug, the time necessary for the initially administered drug to reach maximal efficacy should be taken into account. In hypertensive emergencies urapidil, sodium nitroprusside, nitroglycerin, hydralazine and phentolamine are the drugs of choice, with the addition of beta-blockers during catecholamine crisis or dissecting aortic aneurysm. Childhood hypertension is most often treated with angiotensin-converting enzyme (ACE) inhibitors or calcium antagonists, primarily nifedipine. Because of the teratogenic risk involved with ACE inhibitors, extreme caution must be exercised when prescribing for adolescent females. The propagation of health benefits to breast-fed infants, combined with more women delaying pregnancy until their fourth decade, has entailed an increase in the need for hypertension management during lactation. Low dose hydrochlorothiazide, propranolol, nifedipine and enalapril or captopril do not pose enough of a risk of preclude breastfeeding in this group. The most frequently used antihypertensive agents during pregnancy are methyldopa, labetalol and calcium channel antagonists. Methyldopa and beta-blockers are the drugs of choice for treating mild to moderate hypertension. Prazosin and hydralazine are used to treat moderate to severe hypertension and hydralazine, urapidil or labetalol are used to treat hypertensive emergencies. The use of overly aggressive antihypertensive therapy during pregnancy should be avoided so that adequate uteroplacental blood flow is maintained. Methyldopa is the only drug accepted for use during the first trimester of pregnancy.

Quantitative determination of pentaerythrityl tetranitrate and its metabolites in human plasma by gas chromatography/mass spectrometry.

Up to now, there has been no data available on the pharmacokinetics of pentaerythrityl tetranitrate (PETN, CAS 78-11-5) and its metabolites, pentaerythrityl-trinitrate (PE-tri-N), pentaerythrityl-dinitrate (PE-di-N), pentaerythrityl-mononitrate (PE-mono-N) in human plasma. Therefore, in order to determine PETN and its metabolites in plasma sensitive and highly selective GC/MS methods had to be developed and validated. PETN and its metabolite PE-tri-N were validated in the concentration range 50 pg/ml to 10 ng/ml. Isosorbide dinitrate (ISDN) was used as the internal standard and the analytes were extracted with dichloromethane from the plasma. The mass spectrometric tests were carried out using chemical ionization in the negative mode (NlCl) with the application of ammonia as a reagent gas. The nitrate ion m/z 62 was determined in the analytes and internal standard. The accuracy of the mean of the quality control samples during the three days (between days) was between 100 and 110% (PETN), as well as 90 and 106% (PE-tri-N). After an oral application of 100 mg PETN in a pilot study, unchanged PETN and PE-tri-N was measured in plasma. Both metabolites PE-di-N and PE-mono-N were validated at the concentration range of 0.25 ng/ml to 25 ng/ml plasma. After extraction, these analytes were derivatized with BSTFA (N,O-bis[trimethylsilyl]trifluoro-acetamide). The applied internal standard was isosorbide-5-mononitrate (IS-5-MN). The mass spectrometric tests were carried out in the same manner as for PETN and PE-tri-N with chemical ionization in the NlCl mode. The detected masses were m/z 324 for PE-di-N, m/z 351 for PE-mono-N and m/z 217 for IS-5-MN. The accuracy of the mean of the quality control samples during 5 days were between 104 and 107% (PE-di-N) and 102 and 106% (PE-mono-N). The maximum concentration of these analytes in the subject samples were on the average all over 5 ng/ml plasma after the oral administration of 100 mg PETN.

Absorption and bioavailability of pentaerithrityl-tetranitrate (PETN, Dilcoran 80).

The effects of 80 mg pentaerithrityl-tetranitrate (PETN) as suspension or formulated as tablets were compared to placebo in a single blind, randomized, crossover study in 18 healthy subjects (study A), and the bioequivalence of two tablet formulations (marketed Dilcoran 80 vs a new formulation) was studied in 24 healthy subjects after administration of single oral doses of 80 mg PETN according to a placebo controlled, randomized, double blind, two-way crossover study design (study B). The perfusion of the right middle finger was measured by rheography (altitude A of the changes of resistance and of the incisure D) before and 24 h post-dose, and blood pressure and heart rate were measured in supine position at the same time. The values of area under curve (AUC) of the ratio A/D were calculated by the trapezoidal rule. In study A the mean A/D-values were reduced from about 2.0 to about 1.3 after intake of PETN (solution or tablet) with a minimum 60 to 90 min postdose (solution) and 2 h postdose (tablet). A significant reduction in this ratio was seen up to 8 (solution) or 12 h (tablet) post dose. Changes in blood pressure were not observed while the heart rate decreased in the subjects of all three groups 1 to 2 h postdose followed by an increase by 6 to 10 beats per min. After subtraction of the AUC values of placebo from the PETN-derived AUC values, mean values of 6.61 (SD 1.52, solution) and 7.25 (SD 1.48, A/D*h, tablet) were calculated (p > 0.1, study A).(ABSTRACT TRUNCATED AT 250 WORDS)