Search

This work describes how to utilize the electrochemical technique to determine labetalol hydrochloride (Lab) in pure form and combined pharmaceutical formulation for quality control purposes. Four membrane sensors were developed using two plasticizers, dioctyl phthalate with 2-hydroxypropyl-?-cyclodextrin and ammonium reineckate (RNC) for sensors 1a and 2a, and tributyl phthalate with 2-hydroxypropyl-?-cyclodextrin and ammonium reineckate for sensors 1b and 2b as ionophores in polyvinyl chloride (PVC) matrix. Fast response and stable Nernstian slopes of 59.60, 57.58, 53.00 and 55.00 mV/decade for sensors 1a, 2a, 1b, and 2b, respectively, were obtained by developed sensors within a concentration range 10-4 M-10-2 M over pH range 2.00-5.10. Developed sensors showed good selectivity for Lab in pure form, in the presence of co-administered drugs, many of interfering ions, and excipients present in pharmaceutical formulation. No remarkable difference was detected upon the statistical comparison between the results of proposed sensors and the official method.

Alfuzosin and tamsulosin are recently co-administrated with vardenafil to treat symptoms of benign prostatic hyperplasia and erectile dysfunction. A highly sensitive and simple liquid chromatographic method was developed and validated for the simultaneous determination of the three drugs using moxifloxacin as an internal standard. Isocratic separation was achieved within 7.0 min using phenyl-hexyl column (250 × 4.6 mm i.d.) and a mobile phase composed of acetonitrile/0.25% phosphoric acid (30:70, v/v) at pH 3.0. The analysis was performed at a flow rate of 1.2 mL/min with fluorescence detection at 246/450 nm for Alfuzosin and vardenafil, and 226/322nm for tamsulosin using time programming technique. The proposed method was linear over the concentration ranges of 5.0-50.0ng/mL, 10.0-200.0ng/mL and 20.0-400.0ng/mL for alfuzosin, vardenafil and tamsulosin, with limits of detection of 0.56ng/mL, 0.98ng/mL and 2.81 ng/mL in a respective order. The developed method was successfully applied to determine the studied drugs in dosage forms and human plasma samples and the results were satisfactory as revealed by statistical analysis of the data., (Copyright © 2018 Académie Nationale de Pharmacie. Published by Elsevier Masson SAS. All rights reserved.)

Background: Poor drug adherence in hypertensive patients can lead to treatment failure and increased cardiovascular morbidity, as well as increased costs to society. An analytical method based on ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MSMS) was developed and validated for use in routine therapeutic drug monitoring (TDM). The method includes 21 antihypertensive drugs or active metabolites from the groups beta blockers (n=5), calcium antagonists (n=5), angiotensin II receptor antagonists (n=4), angiotensin converting enzyme (ACE) inhibitors (n=3) and diuretics (n = 3), in addition to one α1-selective alpha blocker., Method: A 200 µL serum sample was handled automatically using a pipetting robot. Protein precipitation was performed with 600 µL of 1% formic acid in acetonitrile (v:v) and phospholipid removal was carried out using a Waters OSTRO™ 96-well plate. After evaporation and reconstitution the eluent was injected thrice with different inlet and mass spectrometric methods to cover the different physico-chemical properties of the drugs and the variations in therapeutic concentration ranges between drugs. Acquity UPLC BEH C18 (2.1x50mm, 1.7 µm) column equipped with a corresponding pre-column was used for chromatographic separation. For every analyte an isotopically labelled analogue served as internal standard, except for lisinopril where enalaprilat-d5 was used., Results: Accuracies were in the range of -13.7 to 13.2% and intra-day and inter-day precisions in the range of 1.1 to 10.5%. The linearity within the calibration ranges expressed as coefficient of determination was higher than 0.995 for all compounds. Matrix effects and recovery efficiencies were within acceptable limits. The limits of quantitation varied from 0.02 to 10.7 µg/L. The stability of the drugs in serum at different conditions was tested. Diltiazem was not stable at 4-8 °C with up to 23.5 % loss after six days. Degradation of atenolol, irbesartan, bendroflumethiazide, hydrochlorothiazide and diltiazem was observed when stored at 30 °C. The suitability of the method was demonstrated in a routine TDM setting, analysing samples from 127 patients undergoing antihypertensive drug treatment., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective: Our study aimed to investigate the effect of mirtazapine on bone metabolism in the orchidectomized rat model., Methods: Rats were divided into three groups. A sham-operated control group (SHAM group) and a control group after orchidectomy (ORX group) received the standard laboratory diet (SLD). An experimental group after orchidectomy (ORX MIRTA group) received SLD enriched with mirtazapine for 12 weeks. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry. Bone marker concentrations of osteoprotegerin (OPG), amino-terminal propeptide of procollagen type I, bone alkaline phosphatase (BALP), sclerostin and bone morphogenetic protein 2 were examined in bone homogenate. The femurs were used for biomechanical testing., Results: Compared with the control ORX group, we found a lower BMD in the ORX MIRTA group. The differences were statistically significant, although not in the lumbar vertebrae. BMD was lower in the MIRTA group, suggesting a preferential effect on cortical bone. However, although the thickness of the diaphyseal cortical bone was not different, the fragility in the femoral neck area was statistically significantly different between the groups in biomechanical testing. Regarding the bone metabolism markers, there was a significant decrease in OPG and BALP levels, suggesting a reduction in osteoid synthesis., Conclusions: The results suggest that prolonged use of mirtazapine may have a negative effect on the synthesis of bone and on its mechanical strength, especially in the femoral neck. Further studies are warranted to establish whether mirtazapine may have a clinically significant adverse effect on bone exclusively in the model of gonadectomized rats, or whether the effect occurs also in the model of gonadally intact animals and in respective human models., (© 2015 S. Karger AG, Basel)

Introduction: Mirtazapine is an antidepressant that acts by enhancing serotonergic and noradrenergic neurotransmission. This study aimed to evaluate mirtazapine pharmacokinetic data from Korean psychiatric patients and to identify the potential factors affecting its steady-state concentration., Methods: A total of 337 samples of steady-state mirtazapine concentrations from 188 adult psychiatric outpatients were retrospectively evaluated. Serum mirtazapine concentrations were measured by high-performance liquid chromatography-tandem mass spectrometry., Results: Median mirtazapine concentration was 43.6 μg/L (164.37 nmol) at a daily dosage range of 7.5-60 mg. At the steady state, mirtazapine dose had a positive correlation with the drug concentration. Mean concentration-to-dose (C/D) ratio was 1.48 μg/L/mg/day (5.58 nmol/mg/day), which was higher than that in a previous study in Caucasian subjects. Age and paroxetine co-medication were positively associated with C/D ratio. Initial mirtazapine concentration and C/D ratio did not show an association with responsiveness in depressive patients., Discussion: This study presented the therapeutic drug monitoring data for mirtazapine and pharmacokinetic variations of mirtazapine in an Asian population. A further study could be helpful for clinical decision making based on the characteristics of patients., (© Georg Thieme Verlag KG Stuttgart · New York.)

Objective: To describe the effects of alpha2 -adrenergic receptor antagonists on the pharmacodynamics of sublingual (SL) detomidine in the horse., Study Design: Randomized crossover design., Animals: Nine healthy adult horses with an average age of 7.6 ± 6.5 years., Methods: Four treatment groups were studied: 1) 0.04 mg kg(-1) detomidine SL; 2) 0.04 mg kg(-1) detomidine SL followed 1 hour later by 0.075 mg kg(-1) yohimbine intravenously (IV); 3) 0.04 mg kg(-1) detomidine SL followed 1 hour later by 4 mg kg(-1) tolazoline IV; and 4) 0.04 mg kg(-1) detomidine SL followed 1 hour later by 0.12 mg kg(-1) atipamezole IV. Each horse received all treatments with a minimum of 1 week between treatments. Blood samples were obtained and plasma analyzed for yohimbine, atipamezole and tolazoline concentrations by liquid chromatography-mass spectrometry. Behavioral effects, heart rate and rhythm, glucose, packed cell volume (PCV) and plasma proteins were monitored., Results: Chin-to-ground distance increased following administration of the antagonists, however, this effect was transient, with a return to pre-reversal values as early as 1 hour. Detomidine induced bradycardia and increased incidence of atrioventricular blocks were either transiently or incompletely antagonized by all antagonists. PCV and glucose concentrations increased with tolazoline administration, and atipamezole subjectively increased urination frequency but not volume., Conclusions and Clinical Relevance: At the doses administered in this study, the alpha2 -adrenergic antagonistic effects of tolazoline, yohimbine and atipamezole on cardiac and behavioral effects elicited by SL administration of detomidine are transient and incomplete., (© 2013 Association of Veterinary Anaesthetists and the American College of Veterinary Anesthesia and Analgesia.)

Tolazoline is an α2-adrenergic receptor antagonist, used in veterinary medicine to antagonize the central nervous system depressant and cardiovascular effects of α2 receptor agonists. The pharmacokinetics and pharmacodynamic effects of tolazoline when administered subsequent to detomidine in the horse were recently reported, although the reversal of the sedative and cardiovascular effects following detomidine may not be complete. The current study therefore investigated the pharmacokinetics and pharmacodynamic effects of tolazoline when administered as a sole agent. Nine healthy adult horses were administered tolazoline (4mg/kgIV) and blood samples were collected at time 0 (prior to drug administration) and at various times up to 72h post drug administration. Plasma samples were analyzed using liquid chromatography-mass spectrometry and resulting data analyzed using compartmental analysis. Systemic clearance, steady state volume of distribution and terminal elimination half-life were 0.820±0.182L/h/kg, 1.68±0.379L/kg and 2.69±0.212h, respectively. Tolazoline administration had no effect on chin to ground distance, but the heart rate decreased (relative to baseline) and the percentage of atrial-ventricular block increased in all horses within 2min of administration. Packed cell volume and glucose concentrations were also increased throughout the sampling period. While not commonly used as a sole agent, caution is indicated whenever tolazoline is administered since the effects may be unpredictable., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Mirtazapine pharmacokinetics was studied in 10 healthy cats. Blood was collected before, and at intervals up to 72 h after, oral dose of 3.75 mg (high dose: HD) or 1.88 mg (low dose: LD) of mirtazapine. Liquid chromatography coupled to tandem mass spectrometry was used to measure mirtazapine, 8-hydroxymirtazapine and glucuronide metabolite concentrations. Noncompartmental pharmacokinetic modeling was performed. Median half-life was 15.9 h (HD) and 9.2 h (LD). Using Mann-Whitney analysis, a statistically significant difference between the elimination half-life, clearance, area under the curve (AUC) per dose, and AUC(∞) /dose of the groups was found. Mirtazapine does not appear to display linear pharmacokinetics in cats. There was no significant difference in glucuronidated metabolite concentration between groups. Pharmacodynamics was studied in 14 healthy cats administered placebo, LD and HD mirtazapine orally once in a crossover, blinded trial. In comparison with placebo, cats ingested significantly more food when mirtazapine was administered. No difference in food ingestion was seen between HD and LD, but significantly more behavior changes were seen with the HD. Limited serum sampling during the pharmacodynamic study revealed drug exposure comparable with the pharmacokinetic study, but no correlation between exposure and food consumed. Mirtazapine (LD) was administered daily for 6 days with no drug accumulation detected., (© 2010 Blackwell Publishing Ltd.)

A rapid and sensitive reversed phase liquid chromatography- tandem mass spectrometry (LC-MS-MS) method is developed for the determination of doxazosin in canine plasma. The samples are prepared by precipitation of proteins using a mixture of methanol and acetonitrile, followed by freezing and evaporation of the organic solvent. The remaining dry residue is redissolved in mobile phase and analyzed by LC-MS-MS with positive electrospray ionization using the selected reactions monitoring mode. An XTerra MS C(18) column, a mobile phase composed of acetonitrile and 2mM ammonium acetate with gradient elution, and a flow rate of 400 microL/min are employed. The elution times for prazosin (internal standard) and doxazosin are approximately 8 and 10 min, respectively. Calibration curves are linear in the 1-20 ng/mL concentration range. Limits of detection and quantification are 0.4 ng/mL and 1.2 ng/mL, respectively. Recovery is higher than 94%. Intra- and inter-day relative standard deviations are below 7% and 8%, respectively. The method is applied for the determination of doxazosin plasma levels following a single administration of doxazosin base and doxazosin mesylate tablets (2 mg dose) to dogs in the fed state. The results indicate possible superiority of the mesylate salt on the plasma input rates of doxazosin.

Two second-order calibration methods based on the parallel factor analysis (PARAFAC) and the alternating penalty trilinear decomposition (APTLD) method, have been utilized for the direct determination of terazosin hydrochloride (THD) in human plasma samples, coupled with the excitation-emission matrix fluorescence spectroscopy. Meanwhile, the two algorithms combing with the standard addition procedures have been applied for the determination of terazosin hydrochloride in tablets and the results were validated by the high-performance liquid chromatography with fluorescence detection. These second-order calibrations all adequately exploited the second-order advantages. For human plasma samples, the average recoveries by the PARAFAC and APTLD algorithms with the factor number of 2 (N=2) were 100.4+/-2.7% and 99.2+/-2.4%, respectively. The accuracy of two algorithms was also evaluated through elliptical joint confidence region (EJCR) tests and t-test. It was found that both algorithms could give accurate results, and only the performance of APTLD was slightly better than that of PARAFAC. Figures of merit, such as sensitivity (SEN), selectivity (SEL) and limit of detection (LOD) were also calculated to compare the performances of the two strategies. For tablets, the average concentrations of THD in tablet were 63.5 and 63.2 ng mL(-1) by using the PARAFAC and APTLD algorithms, respectively. The accuracy was evaluated by t-test and both algorithms could give accurate results, too.

Yohimbine is an alkaloid obtained from the Corynanthe yohimbe tree and other biological sources. Yohimbine is currently approved in the United States for erectile dysfunction and has undergone resurgence in street use as an aphrodisiac and mild hallucinogen. In recent years yohimbine use has become common in body-building communities for its presumed lipolytic and sympathomimetic effects. We describe a 37-year-old bodybuilder in which severe acute neurotoxic effects occurred in 2 h after yohimbine ingestion. The patient presented with malaise, vomiting, loss of consciousness, and repeated seizures after ingestion of 5 g of yohimbine during a body-building competition in a gymnasium. His Glasgow Coma Score was 3, requiring orotracheal intubation. Two hours after admission, vital signs were blood pressure 259/107 mmHg and heart rate 140 beats/min. Treatment with furosemide, labetalol, clonidine, and urapidil and gastrointestinal decontamination were performed. Twelve hours later the patient was extubated with normal hemodynamic parameters and neurological examination. The yohimbine blood levels at 3, 6, 14, and 22 h after ingestion were 5,240; 2,250; 1,530; and 865 ng/mL, respectively, with a mean half-life of 2 h. Few data are available about yohimbine toxicity and the related blood levels. This is a case of a large ingestion of yohimbine in which severe hemodynamic and neurological manifestations occurred and elevated blood levels of yohimbine were detected.

Aims: To evaluate the influence of sex and CYP2D6 genotype on mirtazapine disposition within two bioequivalence studies in healthy volunteers., Methods: Seventy-two healthy volunteers were included in two standard 2 x 2 crossover bioequivalence trials. Subjects received a single 30-mg oral dose of each mirtazapine formulation in each study period. Plasma concentrations were measured from 0 to 96 or 120 h by a HPLC with coupled mass spectrometry validated method. CYP2D6 genotyping was available for 68 subjects that were classified into three phenotypic groups depending on the number of active gene copies: extensive/ultrarapid metabolizers (UM-EM), intermediate (IM) and poor metabolizers (PM). To evaluate the influence of sex and genotype on mirtazapine disposition we performed a linear mixed model for repeated measures. Pharmacokinetic data were log-transformed and AUC and C(max) adjusted to the administered dose/weight. Factors included in the model were centre, formulation, period, sequence, sex and genotype as fixed effects, and subject nested sequence x sex x genotype as random one. A second model was also performed adding the interaction sex x genotype to the previous model., Results: Mirtazapine disposition evaluated as AUC(0-infinity) is influenced by sex (p=0.007) and CYP2D6 phenotype group (p=0.01). Attending to the theoretical figures provided by the model, mean (95% CI) dose/weight adjusted AUC(0-infinity) (ng h/ml)/(mg/kg) is 1516.62 (1411.27-1628.22) in EM/UM, 1613.63 (1482.14-1758.55) in IM and 2049.28 (1779.78-2357.24) in PM. In the case of C(max) these figures also show a trend to higher values in PM, but it did not reach statistical significance. Females show a lower dose/weight adjusted AUC(0-infinity): 1594.39 (1477.70-1720.28) vs. 1837.65 (1694.67-1992.70). On the contrary dose/weight adjusted C(max) is higher in females than in males: 38.33 (34.79-42.28) vs. 32.66 (29.44-36.21)., Conclusions: Both CYP2D6 genotype group and sex influence the disposition of mirtazapine in healthy volunteers and confirm reported data in the literature obtained by different methods. No sex-by-genotype interaction could be detected.

An accurate, selective and sensitive bioanalytical method has been developed and validated for the simultaneous quantification of alfuzosin and solifenacin in human plasma using propranolol as internal standard (IS). The analytes and IS were extracted in methyl tert-butyl ether, separated on Hypurity C8 column and detected by tandem mass spectrometry with a turbo ion spray interface. The method had a chromatographic run time of 3.0 min and linear calibration curves over the concentration range of 0.25-25 ng/mL for alfuzosin and 0.6-60 ng/mL for solifenacin. The intra- and inter-day accuracy and precision (%CV) evaluated at four quality control levels were within 88.2-106.4% and 0.9-7.7% respectively. The absolute recovery from spiked plasma samples was 71.8% for alfuzosin and 93.1% for solifenacin. Stability of alfuzosin and solifenacin was assessed under different storage conditions. The validated method was successfully employed for bioavailability study after oral administration of 10 mg of alfuzosin hydrochloride and 5mg of solifenacin succinate tablet formulations in eight healthy volunteers under fed condition.

Phentolamine mesylate accelerates recovery from oral soft tissue anesthesia in patients who have received local anesthetic injections containing a vasoconstrictor. The proposed mechanism is that phentolamine, an alpha-adrenergic antagonist, blocks the vasoconstriction associated with the epinephrine used in dental anesthetic formulations, thus enhancing the systemic absorption of the local anesthetic from the injection site. Assessments of the pharmacokinetics of lidocaine and phentolamine, and the impact of phentolamine on the pharmacokinetics of lidocaine with epinephrine were performed to characterize this potentially valuable strategy. The blood levels of phentolamine were determined following its administration intraorally and intravenously. Additionally, the effects of phentolamine mesylate on the pharmacokinetics of intraoral injections of lidocaine with epinephrine were evaluated. Sixteen subjects were enrolled in this phase 1 trial, each receiving 4 drug treatments: 1 cartridge lidocaine/epinephrine followed after 30 minutes by 1 cartridge phentolamine (1L1P), 1 cartridge phentolamine administered intravenously (1Piv), 4 cartridges lidocaine/epinephrine followed after 30 minutes by 2 cartridges phentolamine (4L2P), and 4 cartridges lidocaine/epinephrine followed by no phentolamine (4L). Pharmacokinetic parameters estimated for phentolamine, lidocaine, and epinephrine included peak plasma concentration (Cmax), time to peak plasma concentration (Tmax), area under the plasma concentration-time curve from 0 to the last time point (AUClast) or from time 0 to infinity (AUCinf), elimination half-life (t1/2), clearance (CL), and volume of distribution (Vd). The phentolamine Tmax occurred earlier following the intravenous administration of 1Piv (7 minutes than following its submucosal administration in treatment 1L1P (15 minutes) or 4L2P (11 minutes). The phentolamine t1/2, CL, and Vd values were similar for 1L1P, 1Piv, and 4L2P. The Tmax for lidocaine occurred later and the Cmax for lidocaine was slightly higher when comparing the 4L2P treatment and the 4L treatment. The phentolamine-induced delay of the lidocaine Tmax likely represents phentolamine's ability to accelerate the systemic absorption of lidocaine from oral tissues into the systemic circulation.

Modified release (MR) formulations are used to enhance the safety and compliance of existing drugs by improving their pharmacokinetics. Predicting the likely success of MR formulations is often difficult before clinical studies. A systematic in vitro approach using mouse and human tissues was adopted to rationalize the in vivo pharmacokinetics of 9- and 15-h MR formulations of an alpha-adrenoreceptor antagonist, 4-amino-6,7-dimethoxy-2-(5-methanesulfonamido-1,2,3,4 tetrahydroisoquinol-2-yl)-5-(2-pyridyl)quinazoline (UK-338,003). Immediate release UK-338,003 was well absorbed in humans consistent with moderate Caco-2 cell monolayer permeability. In contrast, 9- and 15-h modified release formulations showed marked reductions in C(max) (47.1 and 68.9%) and AUC(0-72) (32.6 and 54.0%). Colonic intubation resulted in 81.3 and 73.8% reductions in C(max) and AUC(0-72). Mechanistic studies in isolated mouse tissues showed that colonic UK-338,003 permeability (P(app) < 0.5 x 10(-6) cm/s) was at least 40 times lower than that for ileum with marked asymmetry. UK-338,003 was found to be a substrate for P-glycoprotein (PGP) with a weaker interaction for multidrug resistance-associated protein-type transporters in mouse intestine. PGP inhibition dramatically increased colonic UK-338,003 permeability to the levels observed in ileum. Low UK-338,003 apical to basolateral permeability was also observed in ex vivo human distal intestine, but both the asymmetry and increase in permeability after PGP inhibition were significantly lower. In conclusion, the poor absorption of MR UK-338,003 in humans can be explained by a combination of PGP-dependent efflux and low intrinsic permeability in the lower bowel. Regional permeability studies in ex vivo tissues used during drug development can highlight absorption problems in the distal bowel and assess the feasibility of developing successful MR formulations.

An on-line SPE-HPLC method, using a monolithic poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) (poly(GMA-EDMA)) based weak cation-exchange (WCX) column, was developed for simultaneous determination of alpha1-adrenergic receptor antagonists in human plasma. The monolithic WCX column was prepared by an in-situ polymerization protocol and modified stepwise with ethylenediamine and chloroacetic acid. On connecting this column to an injection valve, an on-line SPE protocol could be established for removal of matrices (mainly proteins and lipids) and preconcentration of four alpha1-adrenergic receptor antagonists in human plasma. This method was validated and then used for determination of terazosin, alfuzosin, prazosin, and doxazosin in clinical plasma samples. For all analytes, each calibration curve was found to be linear over a range of 0.005-5 microg/mL (R>0.997), and the limit of detection for each analyte was 0.5 ng/mL. Recovery (>80%) and precision (RSD<15%) for inter- and intra-day assay were tested at three concentration levels of each analyte and showed acceptable results for quantitative assay. Real samples from hypertensive patients were monitored and results were in agreement with those of the conventional liquid-liquid extraction-HPLC method. Furthermore, due to its good permeability and biocompatibility, the monolithic WCX sorbent could be reused more than 300 times. The proposed method was especially appropriate for multi-analyte monitoring in plasma samples.

Validated stability-indicating spectrophotometric and spectrofluorimetric assays (SIAMs) were developed for the determination of alfuzosin hydrochloride (ALF) in the presence of its oxidative, acid, and alkaline degradation products. Three spectrophotometric methods were suggested for the determination of ALF in the presence of its oxidative degradation product; these included the use of zero order (0D), first order (1D), and third order (3D) spectra. The absorbance was measured at 330.8 nm for (0D) method, while the amplitude of first derivative (1D) method and that of third derivative (3D) method were measured at 354.0 and 241.2 nm, respectively. The linearity ranges were 1.0-40.0 microg/ml for (0D) and (1D) methods, and 1.0-10.0 microg/ml for (3D) method. Two spectrofluorimetric methods were developed, one for determination of ALF in the presence of its oxidative degradation product and the other for its determination in the presence of its acid or alkaline degradation products. The first method was based on measuring the native fluorescence of ALF in deionized water using lamda(excitation) 325.0 nm and lamda(emission) 390.0 nm. The linearity range was 50.0-750.0 ng/ml. This method was also used to determine ALF in human plasma with the aid of a suggested solid phase extraction method. The second method was used for determination of ALF via its acid degradation product. The method was based on the reaction of fluorescamine with the primary aliphatic amine group produced on the degradation product moiety. The reaction product was determined spectrofluorimetrically using lamda(excitation) 380.0 nm and lamda(emission) 465.0 nm. The linearity range was 100.0-900.0 ng/ml. All methods were validated according to the International Conference on Harmonization (ICH) guidelines, and applied to bulk powder and pharmaceutical formulations.

Doxazosin mesylate is a selective alpha-adrenoreceptor antagonist for the treatment of hypertension and benign prostatic hyperplasia. A simple high performance liquid chromatographic method has been developed and validated for the quantitative determination of doxazosin in plasma. A reversed phase C18 column was used for the separation of doxazosin and prazosin (internal standard) with a mobile phase composed of water, acetonitrile, triethylamine (68:32:0.2 v/v, pH 5.0) at a flow rate of 1.2 mL/min. The fluorescence detector was operated at 246 (excitation) and 389 nm (emission). Intra- and inter-day precision and accuracy were acceptable for all quality control samples including the lower limit of quantification of 1 ng/mL. Recovery of doxazosin from human plasma was greater than 93.4%. Doxazosin was stable in human plasma under various storage conditions. This method was used successfully for a pharmacokinetic study in plasma after oral administration of multiple 4-mg dose of doxazosin gastrointestinal therapeutic system formulation to 16 healthy volunteers. At steady state the mean area under the curve for a dosing interval and elimination half-life were calculated to be 367.0 +/- 63.5 ng x hr/mL and 29.2 +/- 4.5 hr, respectively. There was no difference in pharmacokinetic parameters between male and female.

Alpha1-adrenoceptor antagonists are clinically useful for the improvement of urinary obstruction due to benign prostatic hyperplasia (BPH), and their therapeutic effects are mediated through the blockade of prostatic alpha(1)-adrenoceptors. The present study was undertaken to predict the magnitude and duration of alpha(1)-adrenoceptor occupancy in the human prostate after oral alpha(1)-adrenoceptor antagonists. Prostatic alpha(1)-adrenoceptor-binding parameters of silodosin were estimated by measuring specific [(3)H]prazosin binding in rat prostate after oral administration of this drug. The plasma concentration of silodosin after oral administration in rats and healthy volunteers was measured using a high-performance liquid chromatographic method. The alpha(1)-adrenoceptor-binding affinities (K(i)) of silodosin, tamsulosin, and terazosin in the human prostate and plasma concentrations of tamsulosin and terazosin were obtained from the literature. Using the alpha(1)-adrenoceptor binding parameters of silodosin in rat prostate, alpha(1)-adrenoceptor occupancy in the human prostate was estimated to be around 60-70% at 1-6 h after oral administration of silodosin at doses of 3.0, 8.1, and 16.1 micromol. Thereafter, the receptor occupancy was periodically decreased, to 24% (8.1 micromol) and 54% (16.1 micromol) 24 h later. A similar magnitude and time course of alpha(1)-adrenoceptor occupancy by silodosin in the human prostate were estimated using alpha(1)-adrenoceptor-binding affinities (K(i)) in the human prostate. Despite about two orders of differences in the plasma unbound concentrations after clinically effective oral dosages of silodosin, tamsulosin, and terazosin, there was a comparable magnitude of prostatic alpha(1)-adrenoceptor occupancy by these drugs. In conclusion, the prediction of alpha(1)-adrenoceptor occupancy in the human prostate by alpha(1)-adrenoceptor antagonists may provide the rationale for the optimum dosage regimen of these drugs in the therapy of BPH.

A high performance liquid chromatography method for the determination of mianserin in human serum was developed and validated. Doxepin was used as an internal standard. Mianserin was extracted from human serum using a liquid-liquid extraction with hexane:isoamyl alcohol (99:1, v/v). The sample was then dissolved in 0.05 M phosphoric acid (pH=3.0), and after separation on a Hichrom RPB (250 x 4.6 mm, 5 mm) column, the analytes were measured by ultraviolet detection at 214 nm. The recovery ranged from 86.1 to 94.5% for mianserin. The method was specific and linear over the concentration range of 2.0-128.0 ng/mL. The limit of quantification (LOQ) was established at 2.0 ng/mL (CV=13.8%). The accuracy range was from 92.5 to 107.5%. The method was used to measure mianserin in human serum samples obtained from healthy volunteers who had received a single oral dose of 30 mg mianserin. Pharmacokinetic parameters obtained for the mianserin were in agreement with the existing data.

We investigated the relationship between the pharmacological effect of tamsulosin and its concentrations in plasma and several lower urinary tract (LUT) and arterial tissues in conscious male dogs. Oral administration of tamsulosin (30 and 100 microg/kg) inhibited phenylephrine-induced intraurethral pressure (IUP) elevation. Inhibition peaked at 1, 2 h after dosing and lasted up to 6--8 h. Basal mean blood pressure did not significantly change throughout the observation period. Plasma concentration reached maximum within 0.5 h after dosing, whereas that in LUT tissues (prostate, urethra and bladder) reached maximum at 1, 2 h, and prostatic and urethral concentrations remained higher than those in plasma and arterial tissues at almost all observation points. Prostatic concentrations of tamsulosin at individual time points were 2.2- to 18.0-fold higher than plasma and 3.7- to 12.3-fold higher than mesenteric artery concentrations. Urethral concentrations of tamsulosin were also higher than those in both plasma and mesenteric artery. The prostatic and urethral concentrations of tamsulosin correlated well with its effect on IUP response [r2 = 0.98 (p<0.01) and r2 = 0.99 (p<0.01), respectively]. Our data demonstrate that tamsulosin is selectively retained in LUT tissues compared with plasma and arterial tissues and that its sustained effect on IUP response appears to be related to the prostatic and urethral retention of tamsulosin.

A simple, sensitive and selective LC-MS/MS method was developed for the determination of tamsulosin in human aqueous humor and serum to study the recently reported eye-related adverse effects of this alpha(1)-blocker drug. Aqueous humor samples were analyzed by direct injection, after addition of the internal standard, labetalol. Liquid-liquid extraction with ethyl acetate was used for serum sample preparation. The chromatographic separation was performed on a reversed phase column by gradient elution with acetonitrile -0.1% formic acid at a flow-rate of 0.2 ml/min. Detection and quantification of the analytes were carried out with a linear ion trap mass spectrometer, using positive electrospray ionization (ESI) and multiple reaction monitoring (MRM). The limit of quantification was 0.1 ng/ml for both aqueous humor and serum samples and linearity was obtained over the concentration ranges of 0.1-4.7 ng/ml and 0.1-19.3 ng/ml for aqueous humor and serum samples, respectively. Acceptable accuracy and precision were obtained for concentrations within the standard curve ranges. The method has been used for the determination of tamsulosin in aqueous humor and serum samples from patients that were on tamsulosin medication and underwent cataract surgery.

A molecularly imprinted polymer (MIP) was synthesized and evaluated to selectively extract an alpha-blocker, i.e. alfuzosin, from human plasma. The synthesis of the MIP was performed in dichloromethane with methacrylic acid as monomer and the target drug as template. A first series of experiments was carried out in dichloromethane to estimate the potential of the MIP in its specific recognition medium, i.e. dichloromethane, by developing a selective procedure and by measuring the capacity of the sorbent. An optimized procedure was developed for the selective extraction of alfuzosin with a recovery close to 100% in this medium and a specific capacity of 1.3 micromol g(-1) of MIP was measured. A study in aqueous media was also carried out by a comprehensive approach of the retention mechanism in order to build a selective procedure of extraction. The effects of the amount and of the charge of cations were studied and an optimal pH value was defined to limit matrix effects. Then, the alfuzosin MIP was then directly used to selectively extract the target drug from human plasma with an extraction recovery of 60%. Lastly, a soil was extracted by a pressurized solvent and the resulting extract was cleaned up on the MIP, showing the possibility to use this selective sorbent for the sample treatment of various complex matrices.

A highly sensitive HPLC method for enantioselective determination of carvedilol in human whole blood and plasma was developed. Carvedilol and S-carazolol as an internal standard extracted from whole blood or plasma were separated using an enantioselective separation column (Chiralpak AD column; 2.0 diameter x 250 mm) without any chiral derivatizations. The mobile phase was hexane:isopropanol:diethylamine (78:22:1, v/v). The excitation and emission wavelengths were set at 284 and 343 nm, respectively. The limits of quantification for the S(-)- and R(+)-carvedilol enantiomers in plasma and blood were both 0.5 ng/ml. Intra- and inter-day variations were less than 5.9%. As an application of the assay, concentrations of carvedilol enantiomer in plasma and blood samples from 15 patients treated with carvedilol for congestive heart failure were determined.

A simple, rapid, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of tamsulosin (I), a highly selective alpha1-adrenoceptor antagonist used for the treatment of patients with symptomatic benign prostatic hyperplasia. The analyte and internal standard, mosapride (II) were extracted by liquid-liquid extraction with diethyl ether-dichloromethane (70:30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on a reverse phase Waters symmetry C18 column with a mobile phase of 0.03% formic acid-acetonitrile (30:70, v/v). The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 409.1 solidus in circle 228.1 and m/z 422.3 solidus in circle 198.3 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.1-50.0 ng/mL for tamsulosin in human plasma. The lower limit of quantitation was 100 pg/mL with a relative standard deviation of less than 10%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 2.0 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies., ((c) 2005 John Wiley & Sons, Ltd.)

A new self-emulsifying drug delivery system (SEDDS) and self-microemulsifying drug delivery system (SMEDDS) have been developed to increase the solubility, dissolution rate, and, ultimately, oral bioavailability of a poorly water soluble drug, carvedilol. Ternary phase diagrams were used to evaluate the self-emulsification and self-microemulsfication domains. The self-emulsification time following introduction into an aqueous medium under gentle agitation was evaluated. The minimum self-emulsification time was found at a Tween 80 content of 40%. The particle size distribution and zeta-potential were determined. Benzoic acid had a dual function, it improved the self-emulsification performance of SEDDS and SMEDDS in 0.1 N HCl and lead to a positively charged emulsion. The in vitro dissolution rate of carvedilol from SEDDS and SMEDDS was more than two-fold faster compared with that from tablets. The developed SEDDS formulations significantly improved the oral bioavailability of carvedilol significantly, and the relative oral bioavailability of SEDDS compared with commercially available tablets was 413%.

A high-performance liquid chromatographic method with fluorescence detection for the determination of tamsulosin in human plasma is reported. The sample preparation involved liquid-liquid extraction of tamsulosin from alkalised plasma with butyl acetate and back-extraction of the drug to the phosphate buffer (pH 2). Butyl acetate is preferable to more commonly used ethyl acetate because of its much lower solubility in water. Liquid chromatography was performed on an octadecylsilica column (55 mm x 4 mm, 3 microm particles), the mobile phase consisted of acetonitrile-30 mM dihydrogenpotassium phosphate (25:75 v/v). The run time was 3.5 min. The fluorimetric detector was operated at 228/326 nm (excitation/emission wavelength). An analogue of tamsulosin, (R)-5-[2-[(3-(2-ethoxyphenoxy)propyl)amino]-2-methylethyl]-2-methoxybenzensulfonamide was used as the internal standard. The limit of quantitation was 0.4 ng/ml using 1 ml of plasma. Within-day and between-day precision expressed by relative standard deviation was less than 10% and inaccuracy did not exceed 5%. The assay was applied to the analysis of samples from several pharmacokinetic studies., (Copyright 2004 Elsevier B.V.)

The deterioration of the long-term fractal component of heart rate variability (beta) is reported in patients with organic heart disease. To examine the day-to-day variability of beta and to study the effects of nipradilol (NIP), an antianginal agent, on beta in patients with coronary artery disease (CAD), we performed 24-hour ambulatory ECG (AECG) in 24 patients with silent myocardial ischemia (SMI) and obtained the conventional frequency domain heart rate variability (HRV) indices and beta using the MemCalc system. Patients were divided into two groups: group I consisted of 10 patients (64 +/- 5 years of age) for the assessment of the day-to-day variability of beta; group II comprised 14 patients (60 +/- 6 years of age) for evaluation of the effect of NIP (3 mg b.i.d). In group I, AECG was repeated twice a week, while AECG was recorded before NIP treatment (placebo) and on the last day of the 4-week NIP treatment in group II. In group I, there was significant correlation between the values of beta on day 1 and day 2 (r = 0.79, p < 0.05), the difference being 0.09 +/- 0.15 (coefficient of variance: 6.8%). Bland and Altman's analysis indicated that the results were easily reproducible, making this approach feasible for clinical use. In group II, NIP significantly decreased the values of beta by 10.6% (from -1.23 +/- 0.09 to -1.18 +/- 0.07; p < 0.05). NIP also significantly increased the conventional HRV indices. The degree of decrement of beta by NIP was larger than that of the day-to-day variability. In conclusion, a day-to-day variability exists in beta in patients with SMI, but its value is relatively small. A daily dose of 6 mg NIP ameliorates beta and overcomes this variability.

Phentolamine is a reversible competitive alpha-adrenergic antagonist with similar affinities for alphal and alpha2 receptors. It has a long history of safe clinical use, and was developed as a potential therapy for male erectile dysfunction because of its capacity to increase the arteriolar blood flow to the corpora cavernosa. Phentolamine mesylate was administered to rats by oral gavage at daily doses of 10, 50, and 150 mg/kg for 24 months. A dose-related increase in mortality, ascribed to an exaggerated pharmacologic effect, was seen at high doses. Systemic exposure as measured by plasma drug concentration increased with dose and duration of dosing and slight drug accumulation occurred, particularly in high-dose males. In the treated groups, 10 males and 1 female were diagnosed with hibernomas, neoplasms of brown adipose tissue, which appeared in the thoracic cavity or retroperitoneal area as circumscribed, tan to reddish-brown lobulated masses. Histologically, the masses were well circumscribed with variably sized lobules defined by a rich capillary network and consisted of closely apposed oval to polygonal cells with large amounts of cytoplasm and a centrally located nucleus. The cytoplasm's appearance varied from multivacuolated to univacuolated to granular eosinophilic. In a few cases, neoplastic emboli were observed in capsular vessels. Ultrastructurally, the neoplastic cells contained numerous mitochondria with transverse parallel cristae that occupied over 60% of the cytoplasm and lipid droplets. This study documents the previously unreported development of hibernomas in rats treated with phentolamine mesylate.

1 This double-blind, cross-over, placebo-controlled study on six healthy male volunteers was designed to evaluate the effects of alpha2-adrenoceptor antagonism on cardiac parasympathetic regulation. 2 The subjects received atipamezole intravenously as a three-step infusion, which aimed at steady-state serum concentrations of 10, 30 and 90 ng ml(-1) at 50-min intervals. 3 Drug effects were assessed with repeated recordings of blood pressure and electrocardiogram, in which the high-frequency (0.15-0.40 Hz) R-R interval variation is supposed to reflect cardiac parasympathetic efferent neuronal activity. 4 At the end of the three steps of the infusion, the mean (+/-SD) concentrations of atipamezole were 10.5 (3.9), 26.8 (5.6) and 81.3 (21.1) ng ml(-1). 5 Within this concentration range, atipamezole appeared to reduce slightly the high-frequency R-R interval fluctuations, indicating a minor vagolytic effect in the heart. 6 Atipamezole increased systolic and diastolic arterial pressure, on average by 20 and 14 mmHg (maxima at the second step of the infusion), which evidently reflects an overall sympathetic augmentation.

Background: Carvedilol but not metoprolol exhibits persistent binding to beta-adrenergic receptors (beta-ARs) even after washout in cell culture experiments. Here, we determined the significance of this phenomenon on human beta-ARs in vitro and in vivo., Methods and Results: Experiments were conducted on human atrial trabeculae (n=8 to 10 per group). In the presence of metoprolol, isoproterenol potency was reduced compared with controls (P<0.001). In the presence of carvedilol, isoproterenol identified 2 distinct binding sites of high (36+/-6%; -8.8+/-0.4 log mol/L) and low affinity (-6.5+/-0.2 log mol/L). After beta-blocker washout, isoproterenol potency returned to control values in metoprolol-treated muscles, whereas in carvedilol-treated preparations, isoproterenol potency remained decreased (P<0.001 versus control). In vivo studies were performed in 9 individuals receiving metoprolol succinate (190 mg/d) or carvedilol (50 mg/d) for 11 days in a randomized crossover design. Dobutamine stress echocardiography (5 to 40 microg x kg(-1) x min(-1)) was performed before, during, and 44 hours after application of study medication. Beta-blocker medication reduced heart rate, heart rate-corrected velocity of circumferential fiber shortening, and cardiac output compared with baseline (P<0.02 to 0.0001). After withdrawal of metoprolol, all parameters returned to baseline values, whereas after carvedilol, all parameters remained reduced (P<0.05 to 0.001) despite complete plasma elimination of carvedilol., Conclusions: Carvedilol but not metoprolol inhibits the catecholamine response of the human heart beyond its plasma elimination. The persistent beta-blockade by carvedilol may be explained by binding of carvedilol to an allosteric site of beta-ARs.

A rapid, sensitive and accurate liquid chromatographic-tandem mass spectrometric method is described for the determination of tamsulosin in dog plasma. Tamsulosin was extracted from plasma using a mixture of hexane-ethyl acetate (2:1, v/v) and separated on a C18 column interfaced with a triple quadrupole tandem mass spectrometer. The mobile phase consisting of a mixture of methanol, water and formic acid (80:20:1, v/v/v) was delivered at a flow rate of 0.5 ml/min. Atmospheric pressure chemical ionization (APCI) source was operated in positive ion mode. Selected reaction monitoring (SRM) mode using the transitions of m/z 409-->m/z 228 and m/z 256-->m/z 166.9 were used to quantify tamsulosin and the internal standard, respectively. The linearity was obtained over the concentration range of 0.1-50.0 ng/ml for tamsulosin and the lower limit of quantitation was 0.1 ng/ml. For each level of QC samples, inter- and intra-run precision was less than 5.0 and 4.0% (relative standard deviation (R.S.D.)), respectively, and accuracy was within +/-0.3% (relative error (R.E.)). This method was successfully applied to pharmacokinetic study of a tamsulosin formulation product after oral administration to beagle dogs.

A selective and sensitive HPLC method is described for therapeutic drug monitoring of the antidepressant drug mirtazapine and its active metabolite desmethylmirtazapine. Liquid/solid extraction with C18 cartridges was used for cleanup of plasma samples. The chromatographic separation was carried out on a phenylhexyl column. No interference from other coadministered antidepressants has been observed in 234 samples from 184 patients. The calibration range used was from 1 ng/mL to 100 ng/mL. The analytic method has proven robust and well suited for therapeutic drug monitoring. In addition to qualitative and quantitative validation data for the assay method, concentration measurements in samples from patients on mirtazapine therapy and the relevant dosing information are presented. Median drug levels after a 15-mg dose were 37 ng/mL mirtazapine and 20 ng/mL desmethylmirtazapine. When a 60-mg dose was administered, median concentrations of 83 ng/mL mirtazapine and 65 ng/mL desmethylmirtazapine were found.

4-amino-5-(4-fluorophenyl)-6,7-dimethoxy-2-[4-(morpholinocarbonyl)-perhydro-1,4-diazepin-1-yl]quinoline (UK-294,315) is an antagonist of the human alpha1-adrenoceptor and exhibits nonlinear oral pharmacokinetics in humans. Superproportional increases in Cmax occur (220-fold, over a 1- to 50-mg dose range), area under the curve increases linearly, but time to maximum concentration decreases with dose, suggesting variation in rate but not extent of absorption. Oral absorption in humans is extensive, with only 14% of an orally administered (20 mg) radiolabeled dose excreted unchanged in the feces. In rats and dogs, UK-294,315 is partially eliminated as unchanged drug in feces (29 and 14% of an intravenous dose, respectively). Oral bioavailability is low in rats (11%) and high in dogs (71%), in keeping with systemic clearance. Fecal elimination of unchanged drug was 60% after oral administration to rats, indicating incomplete absorption in this species, whereas absorption in dogs is complete. UK-294,315 is a P-glycoprotein (P-gp) substrate (Km, 15 microM) exhibiting polarized flux in Caco-2 cell monolayers, saturable across a concentration range of 5 to 200 microM. Furthermore, the observations in vitro occurred at similar concentrations to those estimated in the gut lumen in clinical trials (dose range, 1-100 mg). It is considered that P-gp acts as a saturable absorption barrier to UK-294,315, slowing the rate of absorption at low doses, and is responsible for the observed nonlinearity in oral disposition in humans. Rat and dog pharmacokinetic studies offered limited insight into the process(es) driving nonlinear pharmacokinetics in humans. Our current understanding of the functional effects of P-gp in the human intestine, in combination with in vitro studies at clinically relevant concentrations, has helped rationalize the clinical data for UK-294,315.

Xylazine hydrochloride was administered i.m. at 0.35 mg/kg to 13 steers and 10 lactating dairy cows at Time 0. Ten minutes later, tolazoline hydrochloride was given i.v. at 4 mg/kg. Tissue and milk samples were analyzed using gas chromatography with nitrogen and phosphorous detection to determine concentrations of xylazine, 2,6-dimethylaniline (a toxic metabolite of xylazine), and tolazoline (at various intervals). Concentrations of xylazine and 2,6- dimethylaniline were below the limit of quantitation (10 microg/kg) by 72 hours in tissues and 12 hours in milk. The concentration of tolazoline was below 10 microg/kg by 96 hours in tissues and 48 hours in milk. Based on the results of these residue studies submitted by the sponsoring agency to the Ministry of Agriculture and Forestry in New Zealand, withholding periods for both xylazine hydrochloride and tolazoline hydrochloride injection were established.

A selective, sensitive and rapid liquid chromatography-tandem mass spectrometry method for the determination of alfuzosin in plasma was developed. A PE Sciex API 2,000 triple quadrupole mass spectrometer in multiple reaction monitoring (MRM) mode, using TurboIonSpray with positive ionisation was used. Using prazosin as an internal standard, liquid-liquid extraction was followed by C(18) reversed-phase liquid chromatography and tandem mass spectrometry. The mean recovery for alfuzosin was 82.9% with a lower limit of quantification set at 0.298 ng/ml, the calibration range being between 0.298 and 38.1 ng/ml. This assay method makes use of the increased sensitivity and selectivity of tandem mass spectrometric (MS-MS) detection to allow for a more rapid (extraction and chromatography) and selective method for the determination of alfuzosin in human plasma than has previously been described. The assay method was used to quantify alfuzosin in human plasma samples generated in a multiple-dose (5 mg bd.) study at steady state.

The effect of renal impairment on the safety and pharmacokinetics of a once-daily formulation of alfuzosin, 10 mg, was evaluated. In an open, single-dose study, 26 volunteers, ages 18 to 65 years, were classified as having normal renal function (n = 8) or mild (n = 6), moderate (n = 6), or severe (n = 6) renal impairment. Mean Cmax values increased by a factor of 1.20, 1.52, and 1.20 in subjects with mild, moderate, or severe renal impairment, respectively, compared with controls. Values for AUC(0-infinity) were 1.46, 1.47, and 1.44, respectively. The t(1/2z) was increased only in the group with severe renal impairment. Emergent vasodilatory adverse events were reported by 4 of 26 subjects. No discontinuations due to adverse events occurred. Laboratory parameters were satisfactory in all groups. In conclusion, once-daily alfuzosin, 10 mg, could be safely administered to patients with impaired renal function, and dosage adjustment does not seem necessary.

Solid-phase extraction, utilizing a 96-well plate format, was used to isolate an alpha-1a receptor antagonist and internal standard from human plasma. Following the isolation procedure, the analyte and internal standard were separated and detected using reversed-phase HPLC coupled with atmospheric pressure chemical ionization (APCI) mass spectrometry operated in the positive ion multiple reaction monitoring (MRM) mode. Based upon the peak area ratio (analyte: internal standard) the analyte was quantified over a concentration range of 0.02-2 ng/ml. Assay validation results including parameters such as precision and accuracy are presented. The validated method was subsequently used to support human pharmacokinetic studies., (Copyright 2002 Elsevier Science BV.)

Objective: A new patented prolonged release formulation of the alpha1-adrenoceptor antagonist alfuzosin has been developed for once-daily (OD) administration in benign prostatic hyperplasia (BPH). This study was designed to compare 2 dose regimens: 10 mg OD alfuzosin and 2.5 mg TID alfuzosin at steady state., Methods: In an open, randomized crossover study with a 9-day washout between treatments, 18 healthy male subjects (50 - 65 years) received OD or TID alfuzosin tablets orally over 5 days. Both formulations were administered according to the schedule recommended for therapeutic use: OD was administered 5 min after the evening meal, TID was administered in the evening, then in the morning and at noon (30 min before meals). On the fifth day, plasma concentrations were quantitated by HPLC with spectrofluorometric detection., Results: The following pharmacokinetic parameters refer to the geometric mean values for both formulations. Mean Cmax value of 10 mg OD alfuzosin was 15.8 ng/ml at a median t(max) of 9.0 h; Cmax was higher and reached earlier from 2.5 mg alfuzosin TID: 19.3 ng/ml, 19.7 ng/ml and 20.3 at 1.0 hour after each dosing, respectively. Mean AUC(0-24) values after OD and TID were 228.3 and 226.0 ng x h/ml, respectively. Based on AUC(0-24) values corrected by the administered daily dose, the relative bioavailability of alfuzosin OD was 75.7% with a 90% confidence interval of 68.0 - 84.3%. Non-corrected AUC(0-24) values were bioequivalent with a ratio estimate of 101.0% and a 90% confidence interval of 90.7 - 112.5%. The higher daily dose compensated for the loss of bioavailability observed with the OD formulation. Mean t1/2z value was longer for the OD (8.9 h) than the TID formulation (6.9 h). Variability between individuals was similar for the 2 formulations. Both dose regimens were well tolerated., Conclusions: Alfuzosin 10 mg once-daily provides a suitable pharmacokinetic profile for a once-daily administration, equivalent bioavailability between the 2 dosage regimens and a good safety profile justify the use of alfuzosin 10 mg in patients with BPH.

To determine the acute effects of carvedilol (beta-blocker) on cardiovascular and renal function and its pharmacokinetics in dogs. Fifteen mature mongrel dogs (7-15 kg) of both sexes were used in these experiments. Eight dogs served as controls, and seven dogs served as iatrogenic mitral regurgitation (MR) experimental animals. Carvedilol (0.2, 0.4, and 0.8 mg/kg, P.O.) was administered, and the blood carvedilol concentration was analyzed by reverse-phase high-performance liquid chromatography. The response to isoproterenol or phenylephrine was also evaluated. Isoproterenol (0.025 microg/kg/min) was infused via the saphenous vein for 5 min, and phenylephrine (5 microg/kg) was injected with carvedilol (0.2, 0.4 mg/kg) or placebo for 4 days. The heart rate and arterial blood pressure were measured, and LV fractional shortening was measured by echocardiography. Glomerular filtration rate (GFR) and renal plasma flow (RPF) were measured by intravenous infusion of sodium thiosulfate and sodium para-aminohippurate. Carvedilol (0.2 mg/kg) decreased the heart rate, whereas renal function, arterial blood pressure, and left ventricular contractile function were not affected. Carvedilol (0.4 mg/kg) decreased heart rate, blood pressure, and renal function. The tachycardic response to isoproterenol was significantly diminished for 36 hr by 0.4 mg/kg carvedilol. Carvedilol 0.2 mg/kg inhibited this effect for 24 hr. Thus, it is necessary to titrate the dosage of carvedilol, it should be initiated at less than 0.2 mg/kg and titrated up to 0.4 mg/kg for heart failure dogs.

A highly sensitive method for quantitation of tamsulosin in human plasma using 1-(2,6-dimethyl-3-hydroxylphenoxy)-2-(3,4-methoxyphenylethylamino)-propane hydrochloride as the internal standard (I.S.) was established using liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS). After alkalization with saturated sodium bicarbonate, plasma were extracted by ethyl acetate and separated by HPLC on a C18 reversed-phase column using a mobile phase of methanol-water-acetic acid-triethylamine (620:380:1.5:1.5, v/v). Analytes were quantitated using positive electrospray ionization in a quadrupole spectrometer. LC-ESI-MS was performed in the selected ion monitoring (SIM) mode using target ions at m/z 228 for tamsulosin and m/z 222 for the I.S. Calibration curves, which were linear over the range 0.2-30 ng/ml, were analyzed contemporaneously with each batch of samples, along with low (0.5 ng/ml), medium (3 ng/ml) and high (30 ng/ml) quality control samples. The intra- and inter-assay variability ranged from 2.14 to 8.87% for the low, medium and high quality control samples. The extraction recovery of tamsulosin from plasma was in the range of 84.2-94.5%. The method has been used successfully to study tamsulosin pharmacokinetics in adult humans.

Fiduxosin is a new alpha(1)-adrenoceptor antagonist targeted for the treatment of symptomatic benign prostatic hyperplasia. The purpose of this study was to determine and compare the potencies of the alpha(1)-adrenoceptor antagonists terazosin, doxazosin, tamsulosin, and fiduxosin, based on relationships between plasma drug concentrations and blockade of phenylephrine (PE)-induced intraurethral (IUP) and mean arterial pressure (MAP) responses after single oral dosing in conscious male beagle dogs. Magnitude of blockade and plasma concentrations were evaluated at selected time points over 24 h. All drugs produced dose-dependent antagonism of PE-induced IUP and MAP responses. When IUP and MAP blockade effects were plotted against drug plasma concentrations, direct relationships were observed that were well described by the sigmoidal maximal effect model. IUP IC(50) values for terazosin, doxazosin, tamsulosin, and fiduxosin were 48.6, 48.7, 0.42, and 261 ng/ml, respectively. MAP IC(50) values were 12.2, 13.8, 1.07, and 1904 ng/ml, respectively. Uroselectivity index values, defined as MAP IC(50)/IUP IC(50), were 0.25, 0.28, 2.6, and 7.3, respectively. These results extend previous observations with terazosin in this model, showing that doxazosin exhibits a uroselectivity index comparable to terazosin, consistent with the lack of alpha(1)-adrenoceptor subtype selectivity or uroselectivity of these drugs. Tamsulosin, an alpha(1a)-/alpha(1d)-subtype selective agent, had an index value approximately 10-fold greater than the nonselective drugs. Based on its pharmacokinetic profile and a relative uroselectivity 29-fold greater than the nonselective drugs, fiduxosin is expected to exhibit greater selectivity for urethral compared with vascular alpha(1)-adrenoceptors in human and should be a novel, long-acting, uroselective alpha(1)-adrenoceptor antagonist.

The effect of food on the pharmacokinetics of fiduxosin, a novel selective alpha1a-receptor antagonist, was determined in healthy male subjects. This was a Phase I, open-label, single-center, randomized, two-period, crossover, single oral dose study of fiduxosin. Healthy male subjects (N= 14) were administered single oral doses of 30 mg of fiduxosin under fasting or nonfasting (1026 Kcal, 54 g fat, 46% calories from fat) conditions in each period. Fiduxosin plasma concentration profiles were used to assess fiduxosin pharmacokinetics. The mean Cmax, Tmax, AUC(infinity), CL/F and Vbeta/F values under fasting and nonfasting conditions were 34.3 and 150 ng/mL, 5.4 and 5.4 h, 822 and 1940 ng x h/mL, 42.5 and 17.2 L/h, and 924 and 235 L, respectively. The harmonic mean t 1/2 under fasting and nonfasting conditions were 13.9 and 9.28 h, respectively. Food significantly increased the bioavailability of fiduxosin. Under the nonfasting regimen, the Cmax central value was more than 4-fold and the AUC(infinity) central value more than 2-fold the central value of the fasting regimen. Tmax was not significantly different between fasting and nonfasting regimens. Food also decreased fiduxosin oral clearance (CL/F) by 60% and volume of distribution (Vbeta/F) by 75%.

Objective: To establish a high-performance liquid chromatography (HPLC) for the determination of naftopidil in plasma, and make a study of pharmacokinetics of high-dose naftopidil capsules in dogs., Methods: Five healthy dogs were treated with naftopidil capsules in an oral single dose of 200 mg. Naftopidil in dog plasma was determined by HPLC. The mobile phase consisted of acetonitrile and 0.05 mol.L-1 phosphate buffer (pH 6.5) (60:40), verapamil was selected as internal standard, and the eluate was monitored at 230 nm. Pharmacokinetics calculations were carried out by practice pharmacokinetics programs (Version 97, 3P97)., Results: The calibration curve of naftopidil was linear within the range of 10-1,200 ng.ml-1, and the correlation coefficient was 0.9992, the recovery rate of HPLC was (100.23 +/- 3.00)%, within day RSD and between day RSD were no more than 5.56% and 3.30% respectively. The plasma drug concentration-time course in dogs conformed to the one-compartment model. T1/2Ke value was (1.91-4.99) h, Tmax was (1.87-3.21) h, Cmax was (338.79-414.04) ng.ml-1., Conclusion: HPLC is accurate and reliable, and can be used for studying the pharmacokinetics of high-dose naftopidil capsules in dogs.

A sensitive capillary electrophoretic method for the determination of carvedilol enantiomers in 100 microl of human plasma has been developed and validated. Carvedilol and the internal standard carazolol are isolated from plasma samples by liquid-liquid extraction using diethylether. A sensitive and selective detection is provided by helium-cadmium laser-induced fluorescence. The total analysis time is 17.5 min, about 30 min are needed for the sample preparation. The linearity of the assay ranges from 1.56 to 50 ng/ml per carvedilol enantiomer. The limits of quantification (LOQ) for the carvedilol enantiomers in 100 microl of human plasma are 1.56 ng/ml. The inter-day accuracy for R-carvedilol is between 95.8 and 103% (104% at LOQ) and for S-carvedilol between 97.1 and 103% (107% at LOQ); the inter-day precision values are between 3.81 and 8.64% (10.9% at LOQ) and between 5.47 and 7.86% (7.91% at LOQ) for R- and S-carvedilol, respectively. The small sample volume needed is especially advantageous for the application in clinical studies in pediatric patients. As an application of the assay concentration/time profiles of the carvedilol enantiomers in a 5-year-old patient receiving a test dose of 0.09 mg/kg carvedilol are reported.

In order to determine nicergoline pharmacokinetics after oral administration to humans, we have developed two radioimmunoassays, one directed against nicergoline and the other directed against known nicergoline metabolites. The assays were validated according to the recommendations of international regulatory agencies and their limits of quantification were 40 and 10 pg/ml, respectively. In order to further validate the methods, a chromatographic separation of immunoreactive entities was performed with samples from healthy volunteers who were given 15 mg of Sermion (nicergoline orally administered). Chromatographic determination of assay specificity showed that the metabolite radioimmunoassay recognised known nicergoline metabolites but also a new metabolite. Using the antibodies directed against nicergoline, we were unable to detect nicergoline in the human plasma. This suggests that nicergoline is absent in the circulation because of complete metabolism through its first-pass effect.

In the present study the pharmacokinetics and pharmacodynamics of tamsulosin were investigated in anesthetized male dogs. Hypogastric nerve stimulation elevated the intraurethral pressure (IUP), which was inhibited dose dependently by intraduodenal administration of tamsulosin (3-30 microg/kg). The inhibition peaked about 90 min after dosing and lasted up to 240 min. The basal mean blood pressure did not change significantly during the observation period. The plasma, prostatic, and urethral concentrations of tamsulosin were determined by the liquid chromatography-mass spectrometry/mass spectrometry method. The plasma concentration reached the maximal level within 30 min after dosing and gradually declined thereafter. The maximal total plasma concentration of tamsulosin (C(max, t)) and its unbound concentration (C(max, u)) correlated with the maximal effect on IUP response [r(2) = 0.81 (p<0.01, n = 15) and r(2) = 0.84 (p<0.01, n = 15), respectively]. Each individual unbound plasma concentration did not correlate, however, with its associated inhibition of IUP response (r(2) = 0.04, n = 126). Although the plasma concentration of tamsulosin decreased nearly to the lower limit of quantitation 240 min after dosing, the prostatic and urethral concentrations remained high, i.e., 13 to 44 times greater than the plasma concentration. Our data demonstrate that the maximal inhibition by tamsulosin of IUP response is well correlated with the maximal plasma concentration in the early phase. The sustained effect of tamsulosin on IUP response that follows may be related to prostatic and urethral retention of tamsulosin.

A high-performance liquid chromatographic (HPLC) analysis of terazosin in 1 ml of human plasma was developed using prazosin as an internal standard. The plasma sample was extracted with dichloromethane and ethylether and a 100-microl aliquot was injected onto the reversed-phase column. The mobile phase, 0.02 M sodium phosphate buffer:acetonitrile:tetrahydrofuran = 720:220:60 (v/v/v), was run at a flow rate of 0.8 ml/min and the column effluent was monitored using a florescence detector set at 370 and 250 nm for the emission and excitation wave numbers, respectively. The retention times for terazosin and prazosin were approximately 6.4 and 9.8 min, respectively, and the coefficients of variation of terazosin were generally low, below 6.4%. The present HPLC method was successful for the pharmacokinetic study of terazosin in healthy volunteers. Following oral administration of terazosin, 2 mg, to 20 healthy male volunteers, the area under the plasma concentration-time curve from time zero to time infinity was 421 +/- 71.8 ng h/ml and terminal half-life was 9.83 +/- 1.29 h.

The application of liquid chromatography/ion-trap mass spectrometry for simultaneous quantification of multiple drugs and detection of their metabolites for in vitro experiments was reported recently. In the current study, the use of these techniques was extended to in vivo pharmacokinetic (PK) studies of alpha-1a antagonists. In combination with limited time-point PK, greatly increased throughput was demonstrated for the in vivo screening and investigation of in vivo-in vitro correlation. In addition to quantitative analyses, the technique allowed simultaneous detection of major in vivo metabolites without having to reanalyze the plasma samples. The drugs were individually dosed in mice intravenously via tail vein injection and the blood samples were collected 5 min and 2 h after dosing. After the plasma samples for the different drugs had been prepared separately, they were pooled for cassette analysis. The concentrations of five test compounds in the plasma samples at 2 h ranged from 36-1062 ng/mL, whereas their 5-min plasma levels were similar. From the same cassette analysis, major metabolites in the samples were also detected simultaneously through the interpretation of full-scan mass spectra. The metabolite identification confirmed the results from a previous report that the major sites of metabolism are hydroxylation of the phenyl ring not bearing the alkylsulfonamide substitutent, piperidine N-dealkylation, and N-demethylation of the alkylsulfonamide group., (Copyright 2001 John Wiley & Sons, Ltd.)