Facile Access to Solifenacin Impurity K: One-Step Synthesis and an HPLC-MS Method for Its Determination.

Solifenacin (SFC) is a potent muscarinic antagonist that effectively reduces bladder muscle contraction, thereby alleviating symptoms such as frequency of micturition and urgency. Oxidation of SFC leads to the formation of impurities like Impurity K. Effective analysis and control of this impurity is crucial for ensuring compliance with regulatory standards and safeguarding patient health. To address these challenges, we propose a novel one-step synthesis of Impurity K from SFC. Impurity K was synthesized using cerium(IV) ammonium nitrate (CAN) in water/acetonitrile as the solvent. Additionally, we describe a new HPLC-MS method for the detection of Impurity K in solifenacin succinate tablets.

An ultrasound assisted extraction-solid-phase extraction-ultra-performance liquid chromatography combined strategy for atropine determination in Atropa belladonna leaves.

Atropine is an antimuscarinic alkaloid identified in Atropa belladonna. In pharmacopeias, percolation is standardized as an extraction method for A. belladonna leaves, along with liquid-liquid extraction as a cleanup procedure and titration as an analytical method for assaying the atropine in the leaves. In this study, a faster, solvent-saving, and more reliable method for quality control of A. belladonna samples was developed. Ultrasound-assisted extraction was proposed and optimized by fractional factorial design followed by Box-Behnken design. For modeling atropine content, the following optimal conditions were established: particle size, 180 µm; percentage methanol in water, 50%; volume of solvent, 15 ml; time of extraction, 60 min; and number of extractions, two. This led to a significant improvement in atropine extraction (P < 0.001). For cleanup, solid-phase extraction was used as an alternative to liquid-liquid extraction, giving similar results, with higher reproducibility. Finally, for the atropine assay, a UPLC method was validated as a substitute for the classic titration method. Taken together, the development of an ultrasound-assisted extraction-solid-phase extraction-UPLC approach allowed the determination of atropine content in A. belladonna leaves in a time- and solvent-saving manner, with high reliability.

Validated stability-indicating reversed-phase-HPLC method for simultaneous determination of orphenadrine citrate, caffeine and aspirin.

New, simple, rapid and precise reversed-phase high-performance liquid chromatographic method was developed for the simultaneous determination of orphenadrine citrate, caffeine and aspirin in presence of aspirin degradation products, orphenadrine citrate and caffeine process related impurities, and excipients. Good resolution and quantization were achieved on reversed-phase column [Phenomenex™ Luna ODS C(18) (25 cm×4.6 mm, 5 µm particles)]. Gradient elution based on; eluant [A]: 0.1% triethylamine in aqueous potassium dihydrogen phosphate buffer (50 mM; pH 3.0), while as, eluant [B]: acetonitrile, at a flow rate of 1.5 mL min(-1). UV quantitation was set at 215 nm. Linearity was exhibited for orphenadrine citrate, caffeine and aspirin within 0.5-150, 0.5-360 or 0.7-301 µg mL(-1) ranges, respectively. Satisfactory validation results were ascertained in terms of low limits of quantiation (6.33×10(-2)-7.94×10(-2)), mean percentage recovery (98.9-101.4%), precision (<2%) and robustness. The proposed method was proved to be specific, robust and accurate for the determination of cited drugs in pharmaceutical preparations in presence of their degradation products.

Reverse-phase, high performance liquid chromatographic method for the determination of tolterodine tartrate in routine quality control samples.

A selective, sensitive, and rugged reverse phase-high performance liquid chromatographic method has been developed for the determination of tolterodine tartrate in routine quality control samples. The mobile phase consisted of acetonitrile:phosphate buffer (pH 7.0) in 55:45 v/v ratio. The mobile phase was also used for the extraction of tolterodine tartrate from its formulations. The chromatography was carried out on a Luna 100A, C-18 (5-micro, 250 x 4.60 mm) column. The software used in the chromatographic analysis was Empower Photodiode Array (PDA) software (Waters, Milford, CT). The UV spectrophotometric determination was done at 210 nm. Retention time was found to be about 7.0 +/- 0.5 min. The standard curve was linear (r2 = 0.9997) over the concentration range of 0.1-0.3 mg/mL. The method was found to be accurate, precise, specific, and rugged. The limit of detection was 0.16 microg/mL and the limit of quantification was 0.489 microg/mL. With a short chromatographic run time, the proposed method can be used for the estimation of large number of quality control samples in a short period.

Novel Validated HPTLC Method for the Analysis of Two Binary Mixtures Containing Tamsulosin Hydrochloride with Antimuscarinic Agents.

A validated and selective high-performance thin-layer chromatography (HPTLC) method was developed for the analysis of mixures of tamsulosin hydrochloride (TAM) with either tolterodine tartrate (TOL) or solifenacin succinate (SOL) in bulk drug and in combined dosage forms. The proposed method is based on HPTLC separation of the three drugs followed by densitometric measurements of their spots at 224 nm. Separation was carried out on Merck HPTLC aluminum sheets of silica gel 60 F254 using ethyl acetate-methanol-ammonia (6:4:0.05, v/v) as mobile phase. The linear regression analysis data were used for the regression line in the range of 0.1-0.7, 0.4-4 and 1-6 µg band-1 for TAM, TOL and SOL, respectively. The proposed method was validated and successfully applied for the analysis of their pharmaceutical formulations and laboratory-prepared mixtures containing the two bicomponent combinations. The method was validated and showed good performances in terms of linearity, sensitivity, precision, accuracy and stability.

Development and validation of a stability-indicating high performance liquid chromatographic (HPLC) assay for biperiden in bulk form and pharmaceutical dosage forms.

Current compendial (USP) methods of assay for the analysis of biperiden in bulk form and pharmaceutical dosage forms involve the use of titrimetric and spectrophotometric procedures, respectively. These are non-selective and non-stability-indicating techniques. In this work, a stability-indicating high performance liquid chromatographic assay procedure has been developed and validated for biperiden. The liquid chromatographic separation was achieved isocratically on a symmetry C8 column (150 mm x 3.9 mm i.d., 5 microm particle size) using a mobile phase containing methanol-buffer (50:50, v/v, pH 2.50) at a flow rate of 1 ml/min and UV detection at 205 nm. The buffer was composed of sodium dihydrogen phosphate (50 mM) and 1-heptanesulfonic acid sodium salt (5 mM). The method was linear over the concentration range of 0.5-25 microg/ml (r=0.9998) with a limit of detection and quantitation 0.03 and 0.1 microg/ml, respectively. The method has the requisite accuracy, selectivity, sensitivity and precision to assay biperiden in bulk form and pharmaceutical dosage forms. Degradation products resulting from the stress studies did not interfere with the detection of biperiden and the assay is thus stability-indicating.

Impurity profile tracking for active pharmaceutical ingredients: case reports.

Tracking the impurity profile of an active pharmaceutical ingredient (API) is a very important task for all stages of drug development. A systematic approach for tracking impurity profile of API is described. Various real pharmaceutical applications are presented through successful examples of impurity profile tracking for three different novel APIs. These include MK-0969, an M3 antagonist; MK-0677, an oral-active growth hormone secretagogue and API-A, a cathepsin K inhibitor. A general strategy including selection of a reversed phase high performance liquid chromatographic (RP-HPLC) impurity profile method based on screening various stationary phases and changing the pH of the mobile phase and elucidation of impurity structures through the utilization of LC-MS, preparative-LC and NMR is demonstrated. A series of studies were conducted on the peak purity check by using the LC-UV diode-array and LC-MS detections. The advantages and disadvantages of each technique in the evaluation of peak purity are discussed.

Determination of (R,R)-glycopyrronium bromide and its related impurities by ion-pair HPLC.

A simple, rapid and specific ion-pair HPLC method for the determination of (R,R)-glycopyrronium bromide and its related impurities is presented, and parameters affecting the chromatographic properties of these compounds are discussed. Optimal analyte separation was achieved on base deactivated Nucleosil at 40 degrees C, using phosphate buffer pH 2.30 with sodium-1-decanesulfonate (0.01 M)/methanol (35/65; v/v) as eluent for isocratic elution at a flow rate 1 ml x min(-1). The analytical assay was validated according to international guidelines. The methodis suitable for in-process control and as stability indicating assay.

Analysis of hallucinogenic constituents in Amanita mushrooms circulated in Japan.

The constituents of seven mushrooms sold as Amanita muscaria or Amanita pantherina (five A. muscaria and two A. pantherina) and four "extracts purported to contain A. muscaria" products that are currently circulated in Japan were determined. All mushroom samples were identified as A. muscaria or A. pantherina by macroscopic and microscopic observation. The dissociative constituents, ibotenic acid (IBO) and muscimol (MUS), were extracted with 70% methanol twice and determined by gas chromatography/mass spectrometry. The IBO (as the hydrate)/MUS contents were in the range of <10-2845ppm/46-1052ppm in the cap of A. muscaria and 188-269ppm/1554-1880ppm in the cap of A. pantherina. In the caps, these compounds had a tendency to be more concentrated in the flesh than in the cuticle. On the other hand, the IBO/MUS contents in the stem were far lower than in the caps. In the "extracts purported to contain A. muscaria" products, IBO/MUS were detected below the lower limit of calibration curve (<10ppm/<25ppm) or not detected. However, these samples contained other psychoactive compounds, such as psychoactive tryptamines (5-methoxy-N,N-diisopropyltryptamine and 5-methoxy-N,N-dimethyltryptamine), reversible monoamine oxidase inhibitors (harmine and harmaline) and tropane alkaloids (atropine and scopolamine), which were not quantified. This is the first report of the chemical analysis of Amanita mushrooms that are circulated in the drug market.

Simultaneous spectrophotometric determination of indacaterol and glycopyrronium in a newly approved pharmaceutical formulation using different signal processing techniques of ratio spectra.

Three spectrophotometric methods have been developed and validated for determination of indacaterol (IND) and glycopyrronium (GLY) in their binary mixtures and novel pharmaceutical dosage form. The proposed methods are considered to be the first methods to determine the investigated drugs simultaneously. The developed methods are based on different signal processing techniques of ratio spectra namely; Numerical Differentiation (ND), Savitsky-Golay (SG) and Fourier Transform (FT). The developed methods showed linearity over concentration range 1-30 and 10-35 (µg/mL) for IND and GLY, respectively. The accuracy calculated as percentage recoveries were in the range of 99.00%-100.49% with low value of RSD% (<1.5%) demonstrating an excellent accuracy of the proposed methods. The developed methods were proved to be specific, sensitive and precise for quality control of the investigated drugs in their pharmaceutical dosage form without the need for any separation process.

Muscarinic Receptor Activation Affects Pulmonary Artery Contractility in Sheep: The Impact of Maturation and Chronic Hypoxia on Endothelium-Dependent and Endothelium-Independent Function.

Giang, Michael, Demosthenes G. Papamatheakis, Dan Nguyen, Ricardo Paez, Carla Blum Johnston, Joon Kim, Alexander Brunnell, Quintin Blood, Ravi Goyal, Lawrence D. Longo, and Sean M. Wilson. Muscarinic receptor activation affects pulmonary artery contractility in sheep: the impact of maturation and chronic hypoxia on endothelium-dependent and endothelium-independent function. High Alt Med Biol. 17:122-132, 2015.-Muscarinic receptor activation in the pulmonary vasculature can cause endothelium-dependent vasodilation and smooth muscle-dependent vasoconstriction. Chronic hypoxia (CH) can modify both of these responses. This study aimed to assess the combined influence of CH and maturation on endothelium-dependent and endothelium-independent muscarinic-induced vasoreactivity. This was accomplished by performing wire myography on endothelium-intact or endothelium-disrupted pulmonary arterial rings isolated from normoxic or CH fetal and adult sheep. In endothelium-intact arteries, vasodilation was evaluated using cumulative bradykinin doses in phenylephrine and carbachol precontracted pulmonary arterial segments; and vasoconstriction was examined using cumulative doses of carbachol following bradykinin predilation. Effects of nonselective (atropine) and selective M1 (pirenzepine), M2 (AFDX116), and M3 (4-DAMP and Dau5884) muscarinic receptor antagonists were assessed in disrupted arteries. In normoxic arteries, bradykinin relaxation was twofold greater in the adult compared to fetus, while carbachol contraction was fourfold greater. In adult arteries, CH increased bradykinin relaxation and carbachol contraction. In vessels with intact endothelium, maturation and CH augmented maximal response and efficacy for carbachol constriction and bradykinin relaxation. Approximately 50%-80% of adult normoxic and CH endothelium-disrupted arteries contracted to acetylcholine, while ∼50% of fetal normoxic and ∼10% of CH arteries responded. Atropine reduced carbachol-induced contraction in all vessels. Adult normoxic vessels were most responsive to M3 antagonism, fetal to M2 antagonism, while M1 inhibition had no effect. Overall, muscarinic-induced pulmonary arterial contraction is partially endothelium dependent and appears to develop after birth. Fetuses are more reliant on M3 receptors while M2 receptors predominate in adults, whereas CH augments muscarinic-dependent pulmonary vasoconstriction in both.

Determination of tropicamide in pharmaceutical formulations using high-performance liquid chromatography.

An isocratic, reversed-phase liquid chromatographic method was developed for determination of tropicamide using atropine as an internal standard in a pharmaceutical dosage form. Tropicamide and atropine sulfate were separated using a microBondapak ODS (C18) column by isocratic elution of mobile phase with flow rate of 2.0 ml/min. The mobile phase composition was methanol-50 mM phosphate buffer (pH 4; 30:70, v/v). The eluate was monitored at 257 nm with detector range setting fixed at 0.01 AUFS. Under these conditions, the retention times were 4.81 min for atropine and 11.89 min for tropicamide. The standard calibration curve was linear over a sample concentration range from 2 to 300 microg/ml, with limit of detection of 0.15 microg/ml. The assay linearity was good (typically r2 = 0.9992) and the standard curves were linear in the detection range. The precision of the method (expressed by relative standard deviation) and the accuracy (mean error in percent) were <5% for both intra- and inter-day assays. Recovery at 80-120% of labeled claim ranged from 98.4 to 100.7% for tropicamide. The proposed method was satisfactorily applied to the determination of tropicamide in pharmaceutical preparation and stability indicating studies.

Simultaneous determination of N-butylscopolamine and oxazepam in pharmaceutical formulations by first-order digital derivative spectrophotometry.

A simple method has been developed for the simultaneous determination of N-butylscopolamine bromide and oxazepam in pharmaceutical formulations using first-order digital derivative spectrophotometry. Acetonitrile was selected as the solvent in which both compounds showed well-defined bands. Both analytes showed good stability in this solvent when solutions of the analytes were exposed to light and temperatures between 20 degrees and 80 degrees C. The simultaneous determination of both drugs was performed by the zero-crossing method at 226.0 and 257.0 nm for N-butylscopolamine and oxazepam, respectively. The linear range of determination was found to be 2.5 x 10(-7) to 8.0 x 10(-5) mol/L for N-butylscopolamine and 7.1 x 10(-8) to 8.0 x 10(-5) mol/L for oxazepam. A very good level of repeatability (relative standard deviation) of 0.2% was observed for N-butylscopolamine and oxazepam. The ingredients commonly found in pharmaceutical formulations do not interfere. The proposed method was applied to the determination of these drugs in pharmaceutical formulations (capsules).

Novel strategy for online monitoring of the degradation kinetics of propantheline bromide via a calixarene-based ion-selective electrode.

Propantheline bromide (PB) is a hydrolysable anti-cholinergic drug. A novel strategy for the online monitoring of PB degradation kinetics catalysed by hydroxyl ions is presented. This is achieved by the incorporation of an on-site PB-selective electrode constructed using as an ionophore. This sensor was used to track the hydrolysis of PB by continuous measurement of the decrease in the produced emf over time. The use of this new technique provides real-time observation and yields a continuous profile of the hydrolysis behaviour of PB under various pH conditions as well as the temperature dependency of each reaction. Moreover, a great advantage of this proposed on-line system is its higher accuracy for rate constant estimation relative to other off-line methods. This kinetic data analysis permitted the determination of the hydrolysis activation energy and prediction of the drug shelf life. The estimated activation energy from Arrhenius plot was 20.77 kcal mol(-1).

Scopolamine poisoning among heroin users--New York City, Newark, Philadelphia, and Baltimore, 1995 and 1996.

Heroin is mixed ("cut") frequently with other substances primarily to increase its weight for retail sale (e.g., mannitol and starch) and to add pharmacologic effects (e.g., dextromethorphan and lidocaine). During 1995 and 1996, health departments and poison-control centers in New York City (NYC); Newark, New Jersey; Philadelphia; and Baltimore reported at least 325 cases of drug overdoses requiring medical treatment in persons who had used "street drugs" sold as heroin that probably also contained scopolamine, an anticholinergic drug. This report summarizes the clinical and epidemiologic features of these cases, which represent a new type of drug overdose.

Cellular dielectric spectroscopy: a powerful new approach to label-free cellular analysis.

The past decade has seen a number of significant changes in identifying higher quality lead compounds earlier in the drug discovery process. Cell-based assay technologies yielding high-content information have emerged to achieve this goal. Although most of these systems are based on fluorescence detection, this article describes the development and application of an innovative cellular assay technology based on radio frequency spectrometry and bioimpedance measurements. Using this technique, the authors have discovered a link between cellular bioimpedance changes and receptor-mediated signal transduction events. By performing dielectric spectroscopy of cells across as pectrum of frequencies (1 KHz to 110 MHz), a series of receptor-specific, frequency-dependent impedance patterns is collected. These raw data patterns are used to determine the identity of the cellular receptor-signaling pathway being tested and to quantify stimulation endpoints and kinetics. The authors describe the application of this technology to the analysis of ligand-induced cellular responses mediated by the 3 major classes of G-protein-coupled receptors (GPCRs) and protein tyrosine kinase receptors. This single assay platform can be used with ease to monitor G(s), G(i), and G(q) GPCRs without the need for chimeric or promiscuous G-proteins, fluorophors, or tagged proteins. In contrast to other methods of monitoring cellular signal transduction, this approach provides high information content in a simplified, noninvasive, and biologically relevant fashion.

Micellar electrokinetic chromatography of scopolamine-related anticholinergics.

A simple micellar electrokinetic chromatography (MEKC) method is described for the separation of scopolamine N-oxide hydrobromide (SO), scopolamine hydrobromide (SH), scopolamine N-methylbromide (SM) and scopolamine N-butylbromide (SB), and for the quantitation of SH, SM and SB (using SO as an internal standard). The analysis of these drugs was performed in a phosphate buffer (30 mM; pH 7.00) with sodium dodecyl sulfate (SDS) (30 mM) as an anionic surfactant. Several parameters affecting the separation of the drugs were studied, including the concentrations of the buffer and SDS. The stability of the drugs in the phosphate buffer (pH 7.00) was also examined. Partial application of the method to the determination of scopolamine N-butylbromide in tablets proved to be feasible.

Ultraviolet spectroscopic estimation of microenvironments and bitter tastes of oxyphenonium bromide in cyclodextrin solutions.

The UV absorbance and bitter taste of oxyphenonium bromide (OB), an antiacetylcholine drug, in cyclodextrin (CD) solutions are measured, and the local environment of the binding site and the reduction of the bitter taste intensity are quantitatively estimated from the UV data. The UV spectrum of OB is changed with the addition of alpha-, beta-, and gamma-CD, because the phenyl group of OB is included into the CD cavity. The maximum wavelength, lambda(max), senses environmental changes of OB best among several spectral characteristics. From comparison of lambda(max) between a CD solution and the reference ethanol-water and dioxane-water systems, the dielectric constant of the binding site is evaluated. This value leads us to estimate the microenvironment and structure of the binding site. The suppression of the bitter taste of 4 mM OB by CDs is in the increasing order alpha-CD < gamma-CD < beta-CD. The extent of this suppression can be quantitatively predicted from the UV absorbance by assuming that the free OB molecule alone exhibits the bitter taste, regardless of the kind and concentration of CD. Some implications and limitations of the present approach are discussed.

Nonaqueous versus aqueous capillary electrophoresis for the dosage of N-butylscopolamine in various pharmaceutical formulations.

A simple nonaqueous capillary electrophoresis method is described for the separation of several atropine and scopolamine related drugs. The analysis of these pharmaceutical compounds was performed in a methanol-acetonitrile (25/5, v/v) mixture containing 25 mM ammonium acetate and 1 M acetic acid. The robustness was proved using a full factorial design at two levels. The method was validated and successfully applied for the determination of N-butylscopolamine in different pharmaceutical preparations. Results were compared to those obtained by a capillary electrophoresis method based on aqueous media.