Pharmacokinetics and pharmacodynamics of nitroglycerin and its dinitrate metabolites in conscious dogs: Intravenous infusion studies

Intravenous infusions of nitroglycerin (GTN), 1,2-glyceryl dinitrate (1,2-GDN), and 1,3-glyceryl dinitrate (1,3-GDN) were given to four conscious dogs at 10 g/min, 30 g/min, 50 g/min, and 70 g/min of GTN and 20 g/min and 100 g/min of GDNs. The steady state plasma concentrations (Css)of GTN were reached after about 60 min whereas for 1,2-GDN and 1,3-GDN the Csswere reached at about 150 min after the infusion began. Except for one dog, the Cssof GTN were not proportional to infusion rate, however, all dogs together showed a good linear relationship between Cssof GTN and infusion rates with an average correlation coefficient of 0.917±0.102. Large variability in GTN clearance after various infusion rates was observed in all dogs. The Cssratios of 1,2-GDN/GTN and 1,3-GDN/GTN yield overall averages of 31.5 ±17.2 and 5.47 ±3.19,respectively. Average Cssratios of metabolites 1,2-GDN/1,3-GDN were 5.78±1.23. This ratio is different from those obtained after iv bolus and oral dosing indicating that the biotransformation of GTN to 1,2-GDN and 1,3-GDN differs for each dosing route. The clearances for 1,2-GDN and 1,3-GDN were not changed over the dose range of 20 g/min to 100 g/min. Terminal half-lives of 1,2-GDN and 1,3-GDN postinfusion were similar to those values obtained after a single bolus dose (45 min). It appears that all the GTN dose at steady state can be accounted for by the formation of measurable 1,2-GDN and 1,3-GDN. Large intra- and interdog variations in systolic blood pressure decrease (SPD) following infusions of GTN were observed, however, all dogs showed a clear systolic blood pressure decrease when the highest infusion rate (70 g/min) was given. No significant systolic blood pressure drop was detected following 20 g/min infusions of 1,2-GDN or 1,3-GDN. It was clear that systolic blood pressure in all dogs decreased following 100 g/min infusions of 1,2-GDN or 1,3-GDN. When SPD values were plotted vs. log GTN concentrations following the infusion of 70 g/min of GTN in all four dogs, a counterclockwise hysteresis was observed indicating the significant contribution of the active dinitrate metabolites to GTN pharmacodynamics.This work was supported in part by NIH grant HL32243.     

Variable Glyceryl Dinitrate Formation Following Infusions of Glyceryl Trinitrate at Different Vascular Sites in the Rat

The availability of glyceryl trinitrate (GTN) and the differential formation of dinitrate metabolites (GDNs) in various organs as a function of routes of administration were investigated in the rat. GTN was infused at 2.0 µg/min via the left femoral vein (LFV), left external jugular vein (LJV), left femoral artery (LFA), and hepatic portal vein (HPV). Blood concentrations of GTN and GDNs were measured in femoral arterial samples. Different infusions yielded GTN steady-state concentrations in the following rank order: LJV LFV > LFA HPV. Furthermore, the GDN formation ratios (1,2-GDN/1,3-GDN) are different: LFV LJV > LFA > HPV. The availabilities of GTN through the leg, vein, and liver were derived. GTN is significantly extracted and metabolized in these organs, and the leg and the vein prefer 1,2-GDN formation, while the liver forms 1,3-GDN predominantly.     

Nitroglycerin dinitrate metabolites do not affect the pharmacokinetics and pharmacodynamics of nitroglycerin in the dog: A preliminary report

Studies were carried out in conscious dogs to determine the effects of 1,2-glyceryl dinitrate (1,2-GDN) and 1,3-glyceryl dinitrate (1,3-GDN) on nitrogtycerin (GTN) pharmacokinetics and pharmacodynamics. In the first set of experiments, steady state plasma levels (Css) of either 1,2-GDN or 1,3-GDN in three dogs were rapidly achieved by giving an iv bolus (77 g/kg), followed immediately by an infusion (50 g/min) of the same GDN. A single iv bolus dose of GTN (0.025 g/kg) was given 50 min after beginning the GDN infusion and compared with plasma concentrations following a similar GTN dose in the absence of dosed GDNs. No significant differences in GTN AUC (p0.9) and CL_app (p 0.7) were found. In a second set of experiments, an infusion of nitroglycerin was begun in each of 4 dogs and continued for 160 min at an infusion rate of 100 gm/min. Steady state concentrations of GTN were achieved within 100 min, at which time the dog received, simultaneously, an iv bolus dose (5.14 mg) of one of the GDNs and an infusion dose (100 gmg/min) of the same GDN. For both dinitrate metabolites no significant differences (p 0.5) were found between control and interaction arterial and venous clearances, although venous GTN clearances tended to decrease in the presence of dosed GDNs. Steady state systolic blood pressure during GDN infusions could be further reduced when GTN doses were administered; however, the steady state systolic blood pressure decrease caused by GTN could not be further reduced by the GDN infusions. Results suggest that the GDNs do not inhibit nitroglycerin metabolism or hemodynamics at the dose levels studied here.Supported in part by National Institutes of Health Grant HL32243.     

Simultaneous pharmacodynamic modeling of the non-steady-state effects of three oral doses of 1,3-glyceryl dinitrate upon blood pressure in healthy volunteers

The organic nitrate 1,3-glyceryl dinitrate (1,3-GDN) is one of the primary dinitrate metabolites of the antianginal agent nitroglycerin (GTN). Investigational New Drug Approval was sought to administer oral solution doses of 1,3-GDN to a small number (n=3) of healthy volunteers; each subject receiving three doses at 1.2, 2.4, and 3.6 mg. With volunteers confined to a semirecumbent posture for the duration of each treatment (4-hr period postdose), diastolic blood pressure (DBP) was recorded and plasma samples collected for 1,3-GDN concentration analysis. Appreciable concentration-related decreases in DBPwere observed, with maximal decreases from predose baseline values approximating 11 to 25 mm Hg. For each subject parametric pharmacodynamic modeling was performed with simultaneous analysis utilizing the DBPvs. time data from all three doses; an inhibitory E_max pharmacodynamic model was adopted. The temporal relationship between plasma 1,3-GDN concentrations and DBPdisplayed rapid equilibration. For subjects 1, 2 and 3,respectively, E_max was predicted as 12.9, 23.4, and 29.7 mm Hg, representing 21.5, 31.6, and 39.5% decreases in DBPfrom predose baseline values;plasma concentrations at half E_max (C₅₀)were 2.75, 2.43, and 5.93 g/L. Utilizing pharmacokinetic-pharmacodynamic modeling, 1,3-GDN plasma concentrations appear to relate to a systemic effect measure that is mechanistically representative of the therapeutic actions of organic nitrates as peripheral vasodilators. The establishment of a GDN plasma concentration-effect relationship together with the relatively high plasma levels of GDN achieved following GTN dosing supports the hypothesis that the GDNs contribute significantly to the hemodynamic effect observed with GTN.Supported in part by National Institute of Health Grant HL 32243.     

Methaqualone pharmacokinetics after single- and multiple-dose administration in man

Serum levels of methaqualone (MTQ) were determined in eight unfasted subjects following single- and multiple-dose administration of 1×300 mgtablet over a 28-day period. Data were analyzed by a two-compartment open model. Following a fairly rapid absorptive phase (K _a =0.82±0.32 hr^–1),the serum elimination curve was biexponential, consisting of a phase predominantly due to distribution (=0.97±0.55 hr^–1)and a phase predominantly due to elimination (=0.036±0.004 hr^–1).A steady-state MTQ serum concentration profile was observed within the first week. There were no significant changes in the kinetics of absorption, distribution, or elimination over the 28-day period of drug administration. Urinary D-glucaric acid excretion, which increased two-to threefold after the first week of MTQ dosing, returned to normal levels when the drug was discontinued. The significance of the pharmacokinetic parameters in relation to bioavailability and biological disposition of single and multiple dose MTQ administration is discussed.     

Cutaneous Metabolism of Nitroglycerin in Vitro. II. Effects of Skin Condition and Penetration Enhancement

The effects of skin storage, skin preparation, skin pretreatment with a penetration enhancer, and skin barrier removal by adhesive tape-stripping on the concurrent cutaneous transport and metabolism of nitroglycerin (GTN) have been studied in vitro using hairless mouse skin. Storing the skin for 10 days at 4°C did not alter barrier function to total nitrate flux [GTN + 1,2-glyceryl dinitrate (1,2-GDN) + 1,3-glyceryl dinitrate (1,3-GDN)]. However, metabolic function was significantly impaired and suggested at least fivefold loss of enzyme activity. Heating skin to 100°C for 5 min appreciably damaged hairless mouse skin barrier function. The ability to hydrolyze GTN was still present, however, and remained constant over the 10-hr experimental period, in contrast to the control, which showed progressively decreasing enzymatic function with time. Pretreatment of hairless mouse skin in vivo (prior to animal sacrifice, tissue excision, and in vitro transport/metabolism studies) with 1-dodecylazacyclo-heptan-2-one (Azone), a putative penetration enhancer, significantly lowered the skin barrier to nitrate flux (relative to the appropriate control). Again, barrier perturbation resulted in essentially constant metabolic activity over the observation period. The ratio of metabolites formed (1,2-GDN/1,3-GDN) was increased from less than unity to slightly above 1 by the Azone treatment. Adhesive tape-stripping gradually destroyed skin barrier function by removal of the stratum corneum. The effects of 15 tape-strips were identical to those of Azone pretreatment: a greatly enhanced flux, a constant percentage formation of metabolites over 10 hr (once again), and an increase in the 1,2-GDN/1,3 GDN ratio. Overall, the experiments caution that, for transdermal drug delivery candidates susceptible to skin metabolism, the status of barrier function (enhancer pretreated, skin damage or disease, etc.) may significantly affect systemic availability.     

Glutathione S-Transferase-Mediated Metabolism of Glyceryl Trinitrate in Subcellular Fractions of Bovine Coronary Arteries

The possible role of glutathione S-transferases (GTSs) in vascular glyceryl trinitrate (GTN) metabolism was investigated. GTN degradation to form its dinitrate metabolites (GDNs) in the 9000g (9k) supernatant fraction of bovine coronary arteries (BCA) was examined. BCAs were homogenized with a 3x volume of phosphate buffer, and the 9k fraction was obtained by centrifugation. GTN (40 ng/ml; 1.76 x 10^–7 M) was incubated for 2 hr in the 9k fraction of BCA in the presence of reduced glutathione (2 x 10^–3 M). Samples were taken at 10, 20, 40, 60, and 120 min. GTN was observed to degrade readily, exhibiting a half-life of 26 min in the incubate. While both 1,2- and 1,3-GDNs were generated from GTN, formation of 1,3-GDN was predominant (GDN ratio, as 1,2/1,3-GDN, = 0.7–0.8). Coincubation with 2 x 10^–5 Mconcentrations of two GST inhibitors, sulfobromophthalein (SBP) and ethacrynic acid (ECA), decreased the rate of GTN loss. The GTN half-lives in SBP- and ECA-treated incubations were 66 and 84 min, respectively. In addition, the pattern of GDN formation was also altered. The resultant GDN ratios exceeded unity in the presence of these inhibitors, indicating that 1,3-GDN formation was attenuated to a greater extent than that of 1,2-GDN. These data suggest that vascular GTN metabolism in BCA is carried out by cytosolic GST isozymes which possess a preference for C-2 denitration of GTN.     

Species differences in butadiene metabolism between mice and rats evaluated by inhalation pharmacokinetics

Metabolism of 1,3-butadiene to 1,2-epoxybutene-3 in rats follows saturation kinetics. Comparative investigation of inhalation pharmacokinetics in mice also revealed a saturation pattern. For both species linear pharmacokinetics apply at exposure concentrations below 1000 ppm 1,3-butadiene; saturation of butadiene metabolism is observed at atmospheric concentrations of about 2000 ppm.For mice metabolic clearance per kg body weight in the lower concentration range where first order metabolism applies was 7300ml×h^–1 (rat: 4500 ml×h^–1). Maximal metabolic elimination rate (V_max) was 400 mol×h^–1 ×kg^–1 (rat: 220 mol ×h^–1×kg^–1). This shows that 1,3-butadiene is metabolized by mice at higher rates compared to rats.Based on these investigations, the metabolic elimination rates of butadiene in both species were calculated for the exposure concentrations applied in two inhalation bioassays with rats and with mice. The results show that the higher rate of butadiene metabolism in mice when compared to rats may only in part be responsible for the considerable difference in the susceptibility of both species to butadiene-induced carcinogenesis.     

Pharmacokinetics and safety of ILX23-7553, a non-calcemic-vitamin D3 analogue, in a phase I study of patients with advanced malignancies

Purpose: Differentiation therapy is an alternative to chemotherapy with potentially less toxicity, improved quality of life, and survival. We conducted a phase I trial of ILX23-7553, a formulation of 1,25-dihydroxy-16-ene-23-yne-vitamin D₃, a 1,25-dihydroxyvitamin D₃ analog with preclinically demonstrated antitumor and differentiating effects and diminished hypercalcemic effects. Patients and methods: The protocol consisted of five daily oral treatments during 14-day cycles at 15 dose levels from 1.3 to 45.0g/m²/day. We treated 42 heavily pretreated patients who had a variety of malignancies with 162 treatment cycles, and obtained pharmacokinetics from three patients at the two highest dose levels. Results: There were no grade 3 or 4 toxicities. Grade 1–2 toxicities included diarrhea, nausea, fatigue, constipation, and one grade 1 hypercalcemia. Average day 6 calcium was 9.26 ± 0.55mg/dl in cycle 1 and 9.30 ± 0.67mg/dl in cycle 2. Pharmacokinetics at dose levels 14 (40g/m²/day) (1 patient) and 15 (45g/m²/day) (2 patients) demonstrated an average C _max of 30.4 ± 7.8pg/ml (0.07nM) and 104 ± 38.2pg/ml (0.25nM), and AUCs of 222.5 ± 225.2pg·h/ml and 855 ± 536pgh/ml, respectively. Eight patients (19%) had stable disease. While in vitro effects have been reported at these concentrations, they were at least 10-fold lower than ED₅₀s, and the study was terminated before an MTD was reached. Conclusion: The drug is safe and has potential benefits at serum concentrations where effects begin to be noted in vitro. Further study is needed with a reformulated higher unit dose compound to determine the safety and efficacy of higher serum concentrations.     

The depolarizing action of 5-hydroxytryptamine on rabbit isolated preganglionic cervical sympathetic nerves

Summary This study describes a depolarizing action of 5-hydroxytryptamine (5-HT) on rabbit isolated preganglionic cervical sympathetic nerves using an extracellular recording technique. From cumulative concentration-response curves for 5-HT (1 mol/1-1 mmol/1), the mean maximal depolarization was shown to be 277 ± 32 V and EC₅₀ was 9.4 mol/l(6.5–13.6 mol/l, geometric mean, 95% confidence limits, n = 42). The responses to 5-HT displayed marked tachyphylaxis. When cumulative concentration-response curves to 5-HT and 2-methyl-5-HT were determined in the same preparations (n = 4), the mean maximal response to 5-HT was 519 ± 167 V, EC₅₀ 32.2 mol/l (8.8–118 mol/l) and the mean maximal response to 2-methyl-5-HT was 317 ± 63 V, EC₅₀ 35.1 mol/l (12.9–95.5 mol/l, geometric means, 95 % confidence limits). The action of selective 5-HT antagonists was tested on repeated cumulative concentration-response curves to 5-HT. Neither methiothepin (0.1–1 mol/l, _n = 3) nor ketanserin (0.1–1 mol/l, n = 3) had an action on 5-HT responses. The selective 5-HT₃ antagonists MDL 72222, ICS 205-930 and SDZ 206–830 were all potent antagonists of the 5-HT depolarizations. The action of these antagonists was quantified by determining the apparent pA₂ from the dose ratios and a Schild plot. For MDL 72222 (1 nmol/1-0.1 mol/l), the apparent pA₂ was 9.1 ± 0.1 (n = 12), Schild plot: 9.2; for ICS 205–930 (0.1 nmol/l–3 nmol/1), the apparent pA₂ was 10.4 ± 0.1 (n = 11), Schild plot 10.3, and for SDZ 206–830 (0.03 nmol/l-1 nmol/1), the apparent pA₂ was 11.2 ± 0.1 (n = 12), Schild plot 11.2. 5-HT depolarizations were unaffected by hexamethonium (0.5 mmol/1). 5-HT depolarizations were reduced by superfusion with both Na-free (42 ± 8% of controls, n = 4) and Na/Ca-free media (35 ± 7% of controls, n = 4). It is concluded that 5-HT depolarizations of rabbit preganglionic cervical sympathetic nerve are mediated by 5-HT₃ receptors. The data with selective 5-HT₃ receptor antagonists suggest that the receptor profile may be more like that for the 5-HT₃ receptor on the terminals of sympathetic nerves than that for the 5-HT₃ receptor on the soma of superior cervical ganglion cells or on vagal afferent neurones. Send offprint requests to D. I. Wallis at the above address     

The Potential Use of the South African River Crab,Potamonautes warreni,as a Bioindicator Species for Heavy Metal Contamination

In 1995, preliminary water and sediment analyses of the river bed and burrow sediments from 9 locations along the Mooi River, NW Province, South Africa had shown cadmium concentrations up to 0.009 mg l^–1±0.003 and up to 0.33 and 0.89 weight % with scanning electron microscopy (SEM) and x-ray microanalysis. Samples of the adult river crab (Potamonautes warreni) were collected from the Mooi River at Noordbrug (26°40S/27°05E), 1 km north of Potchefstroom Town, and exposed to 0.2 or 2.0 mg Cd²⁺ l^–1 in situ to determine tolerance, uptake and bioaccumulation of cadmium. Using flame atomic absorption spectroscopy (FAAS) the gills, haemolymph and digestive gland of naturally exposed P. warreni showed wet mass values of 0.74±0.27 g Cd²⁺ g^–1, 0.007±0.007 g ml^–1 and 0.12±0.09 g g^–1 respectively. The tolerance of crabs to aqueous Cd reached its limit (ET₅₀=42 hours) at 2.0 mg l^–1 aqueous Cd exposure. At an exposure to 0.2 mg Cd²⁺ l^–1 for 21 days, the greatest Cd (n=11; 9.99±5.09 g g^–1 wet mass) and Cu concentrations (n=11; 17.90±4.66 g g^–1 wet mass) were associated with the gills, and to a lesser extent the digestive gland (n=11; 0.38±0.20 g g^–1 wet mass), whereas concentrations of Zn were variable in both organs. In the haemolymph Cd levels were relatively small (n=11; 0.012–0.006 g ml^–1) with exposure and time and Cu, Zn concentrations varied. Changes in the uptake of Cd in P. warreni indicated that transport, storage and possibly regulatory mechanisms are likely to operate in adult crabs. The potential of P. warreni as a bioindicator species of pollution is also discussed.     

Isoprenaline-induced increase in mRNA levels of inhibitory G-protein α-subunits in rat heart

Summary Long-term -adrenergic stimulation has been shown to desensitize the -adrenoceptor/adenylyl cyclase signalling pathway at both the receptor and the G-protein level. To further elucidate the cellular mechanism of G-protein regulation we investigated the influence of prolonged infusion of isoprenaline (2.4 mg/kg·d) on myocardial mRNA levels of different G-protein -subunits in rats. For comparison rats were treated with triiodothyronine (T₃; 0.5 mg/kg·d) which induces cardiac hypertrophy like isoprenaline but has different effects on the adenylyl cyclase system. Isoprenaline- and T₃-treated animals developed an increase in heart/body weight ratio of 41±3% and 27±4%, respectively (P<0.05). Isoprenaline increased myocardial total RNA concentration by 39±6% (P<0.05). Hybridization with ³²P-labeled rat cDNAs demonstrated an expression rank order of G_s-mRNA>G_i-2-mRNA>G_i–3-mRNA and no detectable expression of G_i–1-mRNA in rat myocardium. mRNA levels of G_s G_i–2 and G_i–3 were 36.9±1.28, 10.7±1.07 and 3.7±0.19 pg/g total RNA, respectively. Isoprenaline increased G_i–2 ^– and G_i–3-mRNA concentrations per g total RNA by 49±18% and 27±710, respectively (P<0.05). This effect was abolished by simultaneously administered propranolol (9.9 mg/kg·d), indicating a,-adrenoceptor-mediated mechanism. In contrast, T₃-induced cardiac hypertrophy was not accompanied by changes in G_i-mRNA expression. G_sa-mRNA levels were unaffected by either treatment.In conclusion, long-term stimulation with isoprenaline in vivo induces a -adrenoceptor-mediated increase in myocardial G_i–2 ^– and G_i–3-mRNA without affecting G_s-mRNA. These results suggest that similar increases in myocardial G_i–2-mRNA in end-stage human heart failure may be at least partly explained by increased -adrenergic stimulation due to increased sympathetic activity.Parts of this work were presented at the wintermeeting of the Deutsche Gesellschaft fur Pharmakologie und Toxikologie in Hannover, 1990 (Eschenhagen et al.), Naunyn-Schmiedebergs Arch Pharmacol 342 (Suppl):R8. The work was supported by the Deutsche Forschungsgemcinschaft Send offprint requests to: T. Eschenhagen at the above address     

Binding of 3H-clonidine to an α-adrenoceptor in membranes of guinea-pig ileum

Summary The binding of ³H-clonidine to membrane particles from guinea-pig ileum was investigated. The specific binding, i.e. the binding that could be inhibited by high concentrations of unlabeled clonidine or noradrenaline, was of high affinity, K _D3 nM. The number of sites was approximately 25 fmol/mg protein. Rate constants of association and dissociation were 5.3×10⁷ M^–1 min^–1 and 0.18 min^–1, respectively. Affinites of various drugs to the binding site were determined by measuring their effect on the binding of ³H-clonidine. The affinity of adrenergic agonists decreased in the order clonidine = tramazoline > (–)-erythro--methylnoradrenaline > (–)-noradrenaline (–)-phenylephrine. (–)-Noradrenaline had about 20 times more affinity than the (+)-isomer. The affinity of -adrenoceptor antagonists decreased in the order phentolamine > rauwolscine = yohimbine > WB 4101 > pseudoyohimbine > prazosin = corynanthine. Yohimbine and rauwolscine had about 100 times more affinity than their stereoisomer corynanthine. Serotonin 10 M and metiamide 10 M did not affect the binding, and propranolol inhibited it only at high concentrations. — The results indicate that ³H-clonidine labels an ₂-adrenoceptor in guinea-pig ileum. The orders of affinity of -adrenoceptor agonists and antagonists agree well with their orders of potency in functional tests, namely as modulators of cholinergic transmission in the guinea-pig ileum and as modulators of noradrenaline release in the rabbit pulmonary artery. An -adrenoceptor should be classified as ₂ when the affinities of clonidine, tramazoline and -methylnoradrenaline greatly exceed the affinity of phenylephrine, and when the affinities of rauwolscine and yohimbine exceed those of prazosin and corynanthine.     

Plasma and muscle tocopherol contents during vitamin E therapy in arterial disease

Summary Plasma tocopherol levels have been determined after single oral doses of two different tocopheryl acetate formulations. It was shown that a tablet preparation of D--tocopheryl acetate in a water dispersible form gave significantly higher plasma tocopherol than an equal dose of a conventional preparation of DL--tocopheryl acetate. — The normal plasma tocopherol of 38 healthy persons was found to be 11.3 ± 4.28 g/ml. Atherosclerotic patients under oral treatment with 300–600 mg tocopheryl acetate daily for 0.5–6 years had a mean plasma concentration of 30.7 ± 14.4 g tocopherol/ml (n = 65). — In 23 atherosclerotic patients suffering from intermittent claudication the muscle contents of tocopherol after treatment with vitamin E increased to averagely three times that of non-treated atherosclerotics. In a limited series studied there was a striking correlation between muscle tocopherol and the improvement of walking distance.

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Tokopherolkonzentration in Plasma und Muskel unter Vitamin-E-Therapie bei Patienten mit Arteriosklerose

Zusammenfassung Es wurde der Tocopherolgehalt m Plasma nach peroralen einmaligen Dosen von zwei verschiedenen Tocopherylacetat-Zubereitungen bestimmt. Es zeigte sich, daß eine Tablettenzubereitung von D--Tocopherylacetat in einer in Wasser dispergierbaren Form einen wesentlich höheren Tocopherolgehalt im Plasma ergab, als die entsprechende Dosis eines konventionellen DL--Tocopherylacetat-Präparates. — Der normale Tocopherolgehalt im Plasma bei 38 gesunden Personen betrug 11.3 ± 4.28 g/ml. Arteriosklerotische Patienten wiesen unter peroraler Behandlung mit 300–600 mg Tocopherylacetat täglich während 0.5–6 Jahren eine durchschnittliche Plasmakonzentration von 30.7 ± 14.4 g Tocopherol/ml (n = 65) auf. — Bei 23 arteriosklerotischen Patienten mit intermittierendem Hinken war der Tocopherolgehalt in den Muskeln nach Behandlung mit Vitamin E im Durchschnitt dreimal größer als bei unbehandelten Arteriosklerotikern. In einer begrenzten Untersuchungsreihe konnte eine auffallende Korrelation zwischen dem Tocopherolgehalt in den Muskeln und der Verlängerung der Gehstrecke nachgewiesen werden.

     

Further studies on (±)-YM-12617, a potent and selective α1-adrenoceptor antagonist and its individual optical enantiomers

Summary YM-12617, 5-[2-[[2-(o-ethoxyphenoxy)ethyl]amino]propyl]-2-methoxybenzenesulfonamide HCl is structurally novel, an extremely potent and highly selective ₁-adrenoceptor antagonist. An asymmetric center exists at the -carbon atom in the phenethylamine portion of YM-12617, therefore two optical enantiomers exist. -Adrenoceptor blocking properties and hypotensive activities of YM-12617 and its enantiomers have been compared in vitro and in vivo. 1. In the isolated rabbit aorta, R(–)- and S(+)-YM-12617 competitively antagonized phenylephrine-induced contraction with pA₂ values of 9.95 and 7.69, respectively. Although R(–)- and S(+)-YM-12617 were also competitive antagonists toward UK-14,304 at prejunctional ₂-adrenoceptors in the isolated guinea-pig ileum, the affinities of R(–)-YM-12617 (pA₂ = 6.18) and S(+)-YM-12617 (pA₂ = 5.64) for these receptors were 5,900 and 110 times lower than those displayed for postjunctional ₁-adrenoceptors in the isolated rabbit aorta. 2. R(–)- and S(+)-YM-12617 displaced both ³H-prazosin and ³H-idazoxan binding to rat brain membranes; however, the affinities of the R(–)- and S(+)-enantiomers for ₁-adrenoceptors (pK_i = 9.95 and 7.83, respectively) were 21,000 and 72 times higher than those for ₂-adrenoceptors (pK _i = 5.62 and 5.97), respectively. 3. Based on pA₂ values obtained in the isolated tissues and pK _i values in the binding assays, R(–)-YM-12617 was 132–182 times more potent than S(+)-YM-12617 as an antagonist at ₁-adrenoceptors. In contrast, the R(–)- and S(+)-enantiomers were similar in potency at blocking ₂-adrenoceptors. 4. In normotensive pithed rats, R(–)- and S(+)-YM-12617 preferentially antagonized the ₁-adrenoceptor mediated pressor effect of phenylephrine with DR₁₀ values of 1.38 and 705 g/kg i. v., respectively, although a high dose (3,000 g/kg i.v.) also inhibited the effect of UK-14,304 at postjunctional ₂-adrenoceptors. R(–)-YM-12617 exhibited an over 2,000-fold selectivity for postjunctional ₁-adrenoceptors, and R(–)-YM-12617 was over 500 times more potent than S(+)-YM-12617 in antagonizing postjunctional ₁-adrenoceptors based on DR₁₀ values. 5. In anesthetized rats, R(–)- and S(+)-YM-12617 dose-dependently produced hypotension with ED₂₀ values, doses required decreased mean blood pressure by 20%, of 0.64 and 61 g/kg i. v., respectively. R(–)-YM-12617 exerted a 95 times more potent hypotensive activity than S(+)-YM-12617, and its isomeric activity ratio was consistent with that for ₁-adrenoceptors but not ₂-adrenoceptors. 6. The present results suggest that the high stereoselectivity of the optical enantiomers of YM-12617 is in the ₁-adrenoceptor, but not in the ₂-adrenoceptor, and their antagonist potency for ₁-adrenoceptors is likely to contribute to the hypotensive effect. Send offprint requests to K. Honda     

In vivo studies on alpha-adrenergic receptor subtypes in human veins

Summary We studied in vivo responsiveness of venous ₁ and ₂-adrenoceptors, measuring the diameter changes in superficial veins in response to -adrenergic agonists and antagonists in healthy human volunteers. The dorsal hand vein technique was used because it permits complete dose-response studies of venous constriction without confounding reflex alterations.Local infusions of all agonists studied induced dose-dependent contraction of the hand vein; the maximal effects (Emax) were: norepinephrine (88% ± 10%), methox amine (97% ± 5%), phenylephrine (95% ± 6%), clonidine (54% ± 12%), and azepexole (68% ± 26%). Clonidine reduced the norepinephrine-induced venoconstriction by 11% ± 10%. Oral doses of 1 mg prazosin antagonized the venoconstriction induced by norepinephrine, methoxamine, and clonidine, but not by azepexole. Yohimbineantagonism was observed against all agonists studied. Inhibition by yohimbine of clonidine-induced venoconstriction was irreversible over 60–180 min.Results show that the in vivo effects on veins of -adrenergic agonists are in good agreement with results from in vitro experiments. Agonists with ₁- and ₂-adrenoceptor subtype selectivity cause venoconstriction in vivo, but ₂-receptor mediated constriction is intrinsically weaker. Clonidine acts as a partial antagonist against norepinephrine, presumably on postsynaptic ₂-receptors. At high doses, ₂-adrenoceptor subtype selectivity of clonidine and yohimbine appear to be partially lost in vivo. Send offprint requests to H. G. Eichler at the above address     

Reflex epicardial coronary vasoconstriction elicited by nicotine in anaesthetized dogs

Summary The effects of arterial chemoreceptor activation by nicotine on coronary artery diameter was studied in anaesthetized, artificially ventilated dogs. Left circumflex coronary artery diameter, coronary blood flow, calculated mean coronary resistance, systemic arterial blood pressure and heart rate were measured. In control dogs (n = 10) the injection of nicotine (100 g) into the carotid artery evoked an increase of arterial pressure (+22 ±9 mm Hg) and a decrease in heart rate (– 36 + 13 beats/min), and tended to increase coronary blood flow (+7 ±4ml/min). Intracarotid nicotine had no effect on large coronary artery diameter (+ 0.02 ±0.03 mm) or total coronary resistance (+ 0.04 ±0.09 mm Hg min/ml) under these conditions. When heart rate was controlled by (1)-adrenoceptor blockade (propranolol, 1 mg/kg i. v.) plus pacing of the right ventricle (n = 4) or (2)-adrenoceptor blockade plus bilateral vagotomy (n = 7), the chemoreflex-induced constriction of the large coronary artery (–0.07 ±0.02 mm and –0.12 ± 0.03 mm, respectively;p < 0.05). In contrast, there was no chemoreflex-induced change in total coronary resistance after -adrenoceptor blockade plus pacing (+0.01 ±0.09 mm Hg min/ml, but after -adrenoceptor blockade plus vagotomy coronary resistance was increased (+0.75 ±0.31 mm Hg min/ml;p < 0.05). The constriction of both large and small coronary arteries was abolished by phentolamine (0.5 mg/kg i. v.). These results suggest that carotid body chemoreceptor stimulation by nicotine can produce reflex -adrenoceptor-mediated constriction of both large and small coronary arteries, and that the constriction of the small vessels is balanced by vagally-mediated dilatation.     

In Vitro Cutaneous Disposition of a Topical Diclofenac Lotion in Human Skin: Effect of a Multi-Dose Regimen

Purpose. This study determines comparative bioavailability of diclofenac sodium lotion compared to an aqueous solution after topical application to viable human skin in vitro. In addition, the difference between a single dose and multiple doses (8 times) was also determined. Methods. An in vitro flow-through diffusion cell system was employed, using radiolabelled diclofenac sodium. Results. Multiple doses of lotion (2 l/cm² and 5 l/cm²) delivered a total of 40.1 ± 17.6 g and 85.6 ± 41.4 g diclofenac, respectively, at 48 h, compared to only 9.4 ± 2.9 g and 35.7 ± 19.0 g absorbed after topical application of diclofenac as an aqueous solution (P < 0.05). A single dose study showed no statistical difference between diclofenac delivered in lotion or an aqueous solution. Over 48 h the total absorption from lotion was 10.2 ± 6.7 g and 26.2 ± 17.6 g (2 l/cm² and 5 l/cm², respectively), compared to 8.3 ± 1.5 g and 12.5 ± 5.7 g from an aqueous solution. Both single doses of lotion and aqueous diclofenac showed decreased diclofenac absorption into the receptor fluid between 12 and 24 h. However, when applied multiple times, absorption from lotion was continually increasing up to 48 h. The total dose accountability ranged from 76.8 ± 8.2% to 110.6 ± 15.1% of the applied dose. Conclusions. Diclofenac lotion exhibited enhanced diclofenac percutaneous absorption rate through human skin (mass, flux and partition coefficient) when applied a multiple number of times and this enhanced absorption was maintained over 48 h. This suggests that a constituent of the lotion (DMSO) will enhance human skin absorption of diclofenac when used in a multi-dose regimen, but not after a single dose.     

Environmental Impacts of Diesel Fuel on Bacteria and Phytoplankton in a Tropical Estuary Assessed Using <Emphasis Type="BoldItalic">In Situ</Emphasis> Mesocosms.

Dissolved or dispersed petroleum hydrocarbon concentrations (DDPH) were monitored in Ponggol estuary, Singapore, fortnightly from July 1999 to June 2000. DDPH concentrations ranged from 4.4 to 248.9gl^–1 and 0.4 to 1099.7gl^–1 for surface and subsurface waters, respectively and with mean concentrations of 41.01gl^–1 in the water column. Absorbed or adsorbed petroleum hydrocarbon (AAPH) concentrations measured in sediments ranged from 20.6 to 541.0 mg kg^–1, with mean concentrations of 148.23 mgkg^–1. In situ mesocosm studies of bacteria and phytoplankton were based on field monitoring ofenvironmentally measured concentrations of petroleum hydrocarbons, using diesel fuel as the source of contaminant. The mesocosm comprised of 25 L clear polycarbonate carboys incubated in situ for 6 days. Water and sediments from a clean site with undetectable levels of petroleum hydrocarbons were used in controls. The treatment mesocosms comprised of mean and highest concentrations of DDPH and AAPH. The study revealed signs of acute toxicity to autotrophs viz., phytoplankton and autotrophic bacteria in treatments simulating concentrations of diesel fuel found in the sediments. A stimulatory effect was seen at lower concentrations. Bacterial heterotrophs responded positively to all concentrations of diesel fuel because of the abundance of a carbon source, reduced grazing pressure and reduced competition for nutrients from phytoplankton.     

Distribution of Gacyclidine Enantiomers in Spinal Cord Extracellular Fluid

Purpose. Determination of the pharmacokinetics of gacyclidineenantiomers, a non-competitive NMDA antagonist, in plasma and spinal cordextracellular fluid (ECF) of rats. Methods. Implantation of microdialysis probes in spinal cord (T9).Serial collection of plasma samples and ECF dialysates over 5 hoursafter IV bolus administration of (±)-gacyclidine (2.5 mg/kg). Plasmaprotein binding determined in vivo by equilibrium dialysis. ChiralGC/MS assay. Results. Plasma concentrations of (+)-gacyclidine were 25% higherthan those of (–)-gacyclidine over the duration of the experiment inall animals. Plasma concentrations decayed in parallel in a biphasicmanner (t_1/2 9 min; t_1/2 90 min) with no significant differencebetween enantiomers. Clearance and volume of distribution of(–)-gacyclidine were approximately 20% higher than those of its opticalantipode (CL: 248 vs 197 ml.kg^–1.min^–1;Vd: 31.6 vs 23.5 l/kg).Protein binding (90%) was not stereoselective. Both gacyclidineenantiomers were quantifiable in spinal cord ECF 10 min after drugadministration and remained stable over the duration of the experimentin spite of changing blood concentrations. Penetration of(–)-gacyclidine was significantly higher (40%) than that of (+)-gacyclidine inall animals. Yet, exposure of spinal cord ECF was similar for bothenantiomers, and not correlated with plasma AUCs. Conclusions. The disposition of gacyclidine enantiomers isstereoselective. Both enantiomers exhibit a high affinity for spinal cord tissueand their distribution may involve a stereoselective and active transportsystem. This hypothesis could also explain the discrepancy betweendrug concentrations in plasma and spinal cord ECF.