New formulation of in situ gelling Metolose-based liquid suppository.

CONTEXT: An in situ gelling liquid suppository is liquid at room temperature but forms a gel at body temperature. In our work, Metolose® SM-4000 (methylcellulose) is studied that basically shows thermal gelation at 68°C (2%, w/w). OBJECTIVE: The objective was to study the potency of different factors (concentration, pH, additives) to change the value of thermal gelation temperature (T (t)) for Metolose® to form an in situ gelling liquid suppository. MATERIALS AND METHODS: We studied the effect of Metolose® concentration, pH, and salts (sodium chloride, potassium chloride, sodium hydrogen carbonate, and sodium monohydrogen phosphate) on T (t) by viscosimetry. To choose the appropriate compound, in vitro drug release was examined. Rectal safety test was performed on rats in vivo after 12-hour application. RESULTS: Increasing the Metolose® concentrations (0.5-4%, w/w), T (t) can be decreased, but it also altered the consistency of gel. pH does not affect the T (t). The water-soluble salts allowed reducing the gelation temperature to 37°C. Sodium monohydrogen phosphate in 4.5% concentration was found to be the most appropriate. The impact of examined factors on in vitro drug release of piroxicam from the in situ-formed gel was characterized according to Fickian diffusion. Metolose® and the chosen salt did not cause any morphological damage on the rectal tissues. DISCUSSION: According to our study, Metolose® has the physical and chemical potential to be used as base for liquid suppositories.

Study on process parameters and optimization of microencapsulation based on phase separation.

As surfactants are capable of influencing the droplet formation, our study primarily aims the investigation of the effect of a nonionic surfactant e.g. Polysorbate 80 on the formation of microspheres on the course of vibrating nozzle method with coacervation. The experiments also concern the impact of the different process parameters (e.g. vibration frequency, feed rate and voltage) on the shape and size distribution of microspheres characterized by laser diffraction size determination completed with particle image analysis. The calcium-alginate microspheres were processed using freeze-drying to ensure solid state with better drug carrier capability. Addition of isomalt was advantageous in the formation of freeze-dried microspheres at low alginate concentration, which was explained by micro-CT analysis of the constructed particle structure. The internal three-dimensional network of calcium alginate demonstrated a more cancellous architecture ameliorating the roundness of microparticles.

Potential application of Metolose in a thermoresponsive transdermal therapeutic system.

The purpose of the present study was to formulate a novel thermoresponsive membrane controlled therapeutic system from Metolose for possible transdermal application. Metolose gel shows thermal gelation property, which can be characterized by two (T(1), T(2)) temperatures. A sharp decrease of viscosity can be measured at T(1), but gelation can be observed at T(2). Different types of Metolose polymers were compared considering their thermoresponsive behaviour. Only thermal gelation was observed in the case of Metolose SM, while Metolose SH showed a sudden decrease of viscosity at T(1). Since this temperature is above the body temperature, so it should be shifted to the skin temperature in case of possible transdermal application. Modulation of thermoresponsibility was followed by rheological method, and the thermoresponsive drug release from Metolose gel was studied by static liberation test. Our results demonstrated that the effect of different salts (NaCl, NaHCO(3), KCl) of various concentrations in Metolose SH gel reduced T(1) to the skin temperature, which enabled enhanced drug release.

Metabolism of moexipril to moexiprilat: determination of in vitro metabolism using HPLC-ES-MS.

Moexipril is a long-acting, non-sulfhydryl angiotensine-converting enzyme inhibitor. It is used for treatment of arterial hypertension. Moexipril is the prodrug, yielding moexiprilat by hydrolysis of an ethyl ester group. Moexiprilat is the metabolite responsible for the pharmacological effect after moexipril administration. Samples of rat and human microsomal preparations exposed to moexipril treatment were analyzed by HPLC using octyl silica stationary phase and isocratic elution. To detect moexipril and moexiprilat the separation was monitored by both ultraviolet and mass specific detection. The rat liver microsomal preparation was more effective to in producing moexiprilat than the similar one derived from human liver cell lines. While additional potential metabolites of moexipril were suggested by computer-modeling, moexiprilat was the sole metabolite detected after microsomal treatment.

Determination of alpha-methyldopa in human plasma by validated high-performance liquid chromatography with fluorescence detection.

A sensitive reversed-phase gradient elution high-performance liquid chromatographic method with fluorescence detection has been developed for the determination of alpha-methyldopa (AMD) in human plasma. Separation of the investigated compound and the 3,4-dihydroxyphenylalanine (DOPA) internal standard was achieved on a Nucleosil 7 C18 column with a 5 mM heptanesulphonic acid sodium salt containing 0.05 M potassium dihydrogenphosphate (pH 3.2)-acetonitrile mobile phase. The composition of the mobile phase was changed according to a linear gradient time program. Detection was performed at 270 nm fluorimetric excitation and 320 nm emission. The compounds were isolated from plasma by Bond-Elut C18 solid-phase extraction. The limit of quantitation was found to be 10 ng/ml plasma. The assay was validated with respect to accuracy, precision and system suitability. All validated parameters were found to be within the 20% required limits. On the basis of the sensitivity, linearity and validation parameters the developed analytical method was found to be suitable for application in a bioequivalency study.

Determination of panomifene in human plasma by high-performance liquid chromatography.

An ion-pair HPLC method was developed for the determination of the antiestrogenic drug panomifene, (E)-1,2-diphenyl-1-(4-[2-(2-hydroxyethylamino)ethoxy]phenyl)-3,3,3 - trifluoropropene, in human plasma. Tamoxifen, 20 ng in 1 ml of plasma, was used as an internal standard. The compounds were isolated from plasma by liquid-solid extraction. Fluorescence detection was achieved by on-line photochemical conversion of the compounds into highly fluorescent phenanthrene derivatives. The sensitivity of the method was 1 ng/ml. The within-day and between-day precision, linearity, extraction recovery and stability of panomifene in plasma and in deproteinized plasma were determined for validation of the method. The method is suitable for measuring plasma levels of panomifene and tamoxifen and for pharmacokinetic studies.

Determination of deramciclane and N-desmethylderamciclane in human plasma by liquid chromatography-tandem mass spectrometry using off-line robotic sample pretreatment.

A rapid and highly sensitive LC-MS-MS method using deuterium-labelled internal standards was developed and evaluated for the simultaneous determination of deramciclane and its pharmacologically active metabolite (N-desmethylderamciclane). The sample preparation based on liquid-liquid extraction was carried out with an off-line robotic system. Evaluation of this analytical method shows that samples can be assayed with acceptable accuracy and precision in the 0.1 to 50 ng/ml concentration range for both compounds. The method was applied for the quantitative determination of deramciclane and its metabolite in human plasma samples during a food interaction pharmacokinetic study.

Antipyrine, coumarin and glipizide affect n-acetylation measured by caffeine test.

The effect of various treatments on acetylation status measured by caffeine metabolites was investigated in 17 subjects with non-insulin dependent diabetes mellitus (NIDDM). The test drugs, caffeine (200 mg), antipyrine (20 mg/kg) and coumarin (5 mg), were given simultaneously, and urinary 5-acetylamino-6-formyl-amino-3-methyluracil/1-methylxanthine (AFMU/1X) molar ratio was measured before and after 8 weeks of therapy. The urinary AFMU/1X molar ratio decreased (p < 0.05) after 8 weeks of therapy with glipizide (2.5 mg), but remained unaltered in those treated with placebo or those who self-monitored blood glucose (SMBG) by glucometer. Antipyrine and coumarin decreased (p < 0.05) the AFMU/1X molar ratio both in diabetics and healthy volunteers. Our data demonstrate that glipizide, antipyrine and coumarin may interfere with the classification of acetylator status measured by caffeine metabolites.

Food interaction study of a new theophylline (Egifilin) 200 and 400 mg retard tablet in healthy volunteers.

The aim of the present food interaction study was to determine the blood plasma levels of theophylline upon the administration of new Egifilin 200 and 400 mg retard tablets, either on an empty stomach or after meal, and to make comparative pharmacokinetic evaluation in 26 healthy volunteers. For determination of the plasma levels of theophylline an improved isocratic HPLC-UV method was used in the concentration range of 0.1-18 micrograms/ml. The mean pharmacokinetic curves obtained with 200 and 400 mg tablets before and after meal were in good agreement also on the basis of statistical evaluation, although, as usual with theophylline, the evaluation of the individual pharmacokinetic curves indicated great variations. The pharmacokinetic parameters (AUCzero-t, AUCzero-infinity, HVD, MRT, Cmax, tmax) calculated for Egifilin 200 and 400 mg retard tablets were analyzed by ANOVA, ANOVAlog, Wilcoxon, and Schuirmann statistical tests as well as by the confidence interval calculation. As it was found, under the circumstances of the present food interaction study, food consumption did not have a biologically significant effect on the pharmacokinetic parameters of either the 200 or the 400 mg Egifilin retard preparations.

Food interaction pharmacokinetic study of cordaflex 20 mg retard filmtablet in healthy volunteers.

The aim of the present study was to investigate the effect of food consumption on the pharmacokinetics of Cordaflex 20 mg retard filmtablet in healthy volunteers through measuring nifedipine plasma levels by an HPLC-ED method both after fasting and food ingestion. The food interaction pharmacokinetic study of Cordaflex 20 mg retard filmtablet was carried out in 12 healthy male volunteers treated with a single dose of the preparation both after fasting and after food ingestion, in a crossover design allowing 1 week of wash-out period between the 2 treatments. Nifedipine concentration of plasma samples were determined by an isocratic HPLC-ED method [Horvai et al. 1994] with robotic sample processing [Horváth et al. 1995, 1996]. The pharmacokinetic parameters (AUC0-infinity, AUC0-t, Cmax, MRT) were analyzed by calculating 90% confidence interval for logarithmic transformed test/reference ratio values, and Schuirmann's statistical tests, the tmax and HVD values were analyzed by Wilcoxon's nonparametric statistical test. The above statistical tests of the present food interaction study indicated significant differences for each one of the respective pharmacokinetic parameter pairs calculated for treatments after fasting and after food ingestion. On the basis of the above findings and also by comparing the mean pharmacokinetic curves, it was evident, that, in agreement with the data of literature [Kleinbloesem et al. 1993, Schall et al. 1994], food ingestion increased the relative bioavailability and maximum plasma concentration (Cmax). Considering the average of the parameter values and also the respective statistical tests, it was also apparent that the time to maximum plasma concentration (tmax), the mean residence time (MRT), and the half-value duration (HVD) all decreased significantly upon the effect of food ingestion.

Pharmacokinetics and safety of deramciclane during multiple oral dosing.

UNLABELLED: Deramciclane is a new putative non-benzodiazepine-type anxiolytic compound. It is a selective serotonin 5-HT(2A) and 5-HT(2C) receptor antagonist and has also inverse agonist properties. The aim of this study was to reveal the pharmacokinetics and tolerability of deramciclane during repeated oral dosing in healthy male volunteers. SUBJECTS, MATERIAL AND METHODS: A randomized double-blind, placebo-controlled design was used. The study had three consecutive groups that received first a single oral dose of 10, 30 and 60 mg of deramciclane followed by twice a day administration for seven days. The total number of subjects was 28. The pharmacokinetic parameters were calculated for a single dose and after repeated administration. Tolerability was assessed by monitoring safety laboratory variables, electrocardiogram, heart rate, blood pressure and adverse events. RESULTS: The steady-state was reached during the seven-day administration. The pharmacokinetics of deramciclane was dose-proportional at steady-state at each dose level. Deramciclane accumulated about three-fold during repeated administration. The relative bioavailability of deramciclane increased about 1.4-fold compared to that of a single dose at each dose level. The mean elimination half-life of deramciclane for 10, 30 and 60 mg doses prolonged from 24.3, 20.9 and 22.9 h after a single dose to 30.5, 25.6 and 28.7 h at steady-state, respectively. Only few adverse events were reported, all mild and transient in nature. The most frequently reported adverse drug reactions were tiredness and headache. There were no deramciclane-induced changes in the clinical chemistry or hematology variables, blood pressure, heart rate or in electrocardiogram. CONCLUSIONS: In conclusion, the pharmacokinetics of deramciclane is linear over the dose range of 10 - 60 mg at steady-state. The slight non-linearity within the dose levels during repeated administration of seven days was regarded as clinically irrelevant. Deramciclane was safe and well tolerated up to doses of 60 mg b.i.d. for seven days.

HPLC method for rapid determination of acetylator phenotype by measuring urinary caffeine metabolites.

A validated reversed-phase high-performance liquid chromatographic (RP-HPLC) method is developed for the selective and rapid determination of two major metabolites of caffeine, namely 5-acetylamino-6-formylamino-3-methyluracil (AFMU) and 1-methylxanthine (MX) from human urine. HPLC separation is achieved by means of a Supersphere-60 RP-Select B (4 microns) analytical column using a non-linear gradient elution programme of 70-95% solvent B (2.5% acetic acid-methanol, 60:40, v/v) in solvent A (water-acetonitrile, 80:20, v/v). A selective UV detection method is used for determination of AFMU, MX and internal standard with readings at 284, 268 and 248 nm, respectively. Urine samples are prepared for measurement by a simple chloroform-diethyl ether (80:20, v/v) extraction. The assay is validated with respect to linearity, sensitivity, accuracy, precision and system suitability. All validation parameters are found to be within the required limits. The limit of detection of AFMU and MX is found to be 50 ng/200 microliters urine. Calibration curves show good linearity between 0.1 and 5 micrograms/200 microliters urine concentration range for both metabolites. The assay is sufficiently sensitive and rapid (4.5 min chromatographic run) to be applied for routine monitoring of change in AFMU/MX molar ratio, indicating acetylation phenotype and change of caffeine metabolism in clinical cocktail studies.

Whole-body autoradiography and quantitative organ-level distribution study of deramciclane in rats.

The distribution of 3H-labelled deramciclane (EGIS-3886), a new 5-HT2 antagonist with anxiolytic activity, has been investigated by whole-body autoradiography and quantitative organ-level determination after intravenous and oral administration to male and female rats at a dose of 3 mg kg(-1). Pregnant dams were also studied, but by autoradiography only. In the autoradiographic study 32 organs were investigated, while in the quantitative organ-level study the radioactivity in 15 organs were determined. There are no sex differences in the distribution of deramciclane, absorption is rapid, elimination is comparatively fast, no specific organ is targeted, and the accumulation of the compound is very unlikely. Penetration of the blood-brain barrier was complete and extremely fast, a very important feature of a potential anxiolytic drug. There is no penetration of the foetus in pregnant dams. The study demonstrated that deramciclane has advantageous pharmacokinetic properties in rats.

Different absorption profiles of deramciclane in man and in dog.

We have studied the dog model for predicting the oral absorption of deramciclane, a novel anxiolytic compound, as a model acid-labile drug. The absorption profile of deramciclane was studied in man and beagle dogs after administration of conventional capsules and enteric coated tablets. Absorption in dogs pretreated with pentagastrin or saline was also studied after administration of conventional capsules. The in-vitro stability of deramciclane was determined over the pH range 1.2-6.0. The rate of degradation of deramciclane increased ten-fold as the pH was reduced from 2.1 to 1.2 (t 1/2 beta (elimination half-life) 9 h and 39 min, respectively). Deramciclane was stable at pH > or = 3. The two formulations were bioequivalent in dogs and there were no significant differences between pharmacokinetic parameters measured for dogs pretreated with pentagastrin or saline. In man the mean relative bioavailability and Cmax (peak plasma concentration) for the conventional capsules were approximately 75% and 83% of those for the enteric coated tablets (P = 0.0004 and P = 0.0031, respectively). This was probably because of degradation of deramciclane at lower pH of man's stomach compared with that of the dog. Pentagastrin was probably unsuccessful in reducing gastric pH and thus no change in absorption was observed. It is concluded that the absorption of deramciclane, and possibly other acid-labile drugs, cannot be predicted by use of the dog model.

Oral, intraperitoneal and intravenous pharmacokinetics of deramciclane and its N-desmethyl metabolite in the rat.

The pharmacokinetic properties of deramciclane fumarate (EGIS-3886), a new potential anxiolitic agent, and its N-desmethyl metabolite have been investigated in Wistar rats after 10 mgkg(-1) deramciclane fumarate was administered orally, intraperitoneally or intravenously. A highly sensitive, validated and optimized gas chromatographic method with nitrogen selective detection (GC-NPD) using a solid-phase extraction technique was used to determine plasma levels of the parent compound and its N-desmethyl metabolite. After oral administration the absorption of the parent compound was very fast (t(max) 0.5h). The maximum plasma concentration (C(max)) was detected at 44.9, > or =177.8 and > or =2643.0 ngmL(-1) after oral, intraperitoneal and intravenous administration of deramciclane, respectively. For the metabolite the respective Cmax values were 32.0, > or =25.4 and 51.0 ngmL(-1). The pharmacokinetic curves of both the parent compound and its metabolite showed enterohepatic recirculation for all administration routes. The biological half-life (tbeta 1/2) for deramciclane ranged from 3.42 to 5.44 h and for the N-desmethyl metabolite the range was 2.90-5.44 h, after administration of the drug by the three different routes. After intravenous administration AUC0-infinity, of deramciclane was 29.2- and 5.4-times higher than that observed after oral and intraperitoneal treatment, respectively. These AUC0-infinity ratios were only 2.1- and 1.5-times higher for the metabolite. The absolute bioavailability of deramciclane in rats was 3.42% after oral and 18.49% after intraperitoneal administration. The comparative pharmacokinetic study of deramciclane in rat after the different administration routes showed fast absorption. Furthermore, plasma levels were found to be administration route-dependent, low bioavailability of the parent compound indicated an extremely fast and strong first-pass metabolism. The apparent volume of distribution suggested strong tissue binding after administration of the drug by any of the three routes studied.

Application of overpressured layer chromatography combined with digital autoradiography and mass spectrometry in the study of deramciclane metabolism.

Overpressured layer chromatography was combined with the highly sensitive and rapid digital autoradiography (DAR) and mass spectrometry to separate, detect, and identify 3H- and 14C-labeled deramciclane metabolites in different biological matrixes. Several minor and major metabolites were separated from plasma and urine samples. The radioactive metabolites localized by DAR were scraped from the thin-layer chromatographic plate and transferred to a mass spectrometer for structure identification. Several metabolites were isolated and characterized, including hydroxy-N-desmethyl deramciclane, which is described in detail. The combination of techniques is efficient and has good sensitivity: about 2 micrograms metabolite from a biological matrix was isolated and identified this way.

Metabolite analysis, isolation and purity assessment using various liquid chromatographic techniques combined with radioactivity detection.

During the discovery of metabolic routes of a drug candidate, radioactively labeled substances are administered. This study reports the multidimensional application of overpressured layer chromatography (OPLC) and high-performance liquid chromatography (HPLC) coupled with online or off-line nondestructive radioactivity detection methods in metabolism studies. Among these methods, digital autoradiography and flow-cell radioactivity detectors (RD) using solid scintillators are used. In this study, the hyphenation of OPLC with RD is reported. The application of the OPLC-RD technique is demonstrated on a metabolism study as well as the multidimensional chromatographic selectivity using normal-phase OPLC for the separation in the first dimension, followed by reversed-phase HPLC-RD, which provides additional selectivity to the separation. Information regarding the identity of radiolabeled metabolites and data obtained from spectroscopic methods could be advantageously used during structure elucidation.

Capillary electrophoretic separation of clomiphene isomers using various cyclodextrins as additives.

The cis (Z) and trans (E) isomers of clomiphene were separated using capillary electrophoresis. Various derivatives of cyclodextrins (CDs) were applied as buffer additives, achieving a resolution of greater than 18. The effect of various parameters, type and concentration of CDs, type and concentration of background electrolytes (BGEs), pH, and organic modifiers has been studied systematically. Migration reversal was observed in several cases. The load ability of the various buffer systems was also taken into account.

The effect of oil as a dietary component on in vitro dissolution of an acid-labile drug.

The non-benzodiazepine-like anxiolytic agent deramiclane fumarate (EGIA-3886) was used to demonstrate that the presence of high oil/fat content in dissolution media serves as a barrier against accelerated drug degradation in acidic media.

Pharmacokinetic studies of flumecinol in man and dog.

The pharmacokinetics of flumecinol (Zixoryn) a new hepatic enzyme inducer has been studied in four beagle dogs and six healthy volunteers. The beagle dogs and the volunteers received the drug orally in a dose of 40 mg/kg of body weight and of 100 mg single dose respectively. Flumecinol was extracted from plasma with diethyl ether and analysed by gas-liquid chromatography using a flame ionisation detector (FID). The pharmacokinetic parameters of flumecinol were determined by computer evaluation of the plasma concentration-time curves. The peak plasma concentrations were found to be 5.3 and 2.1 hours in dogs and humans, respectively. Flumecinol is eliminated from the plasma of dogs and humans with half-lives of 38.95 and 17.16 hours, corresponding to a clearance of 53.2 litres/hour and 94.0 litres/hour, respectively.