A Stability-Indicating LC Assay Method for Bicalutamide

A stability-indicating HPLC method for the quantitative determination of Bicalutamide is described. Bicalutamide is a nonsteroidal antiandrogen and is an oral medication that is used for treating prostate cancer. Separation was achieved on a Waters Symmetry shield RP18 HPLC column using a mobile phase consists of a mixture of phosphate buffer (Solvent A) and organic modifier acetonitrile (Solvent B). Degradation studies were performed on bulk samples of bicalutamide using acid (0.5 N methanolic hydrochloric acid), base (0.5 N methanolic sodium hydroxide), oxidation (10% v/v methanolic hydrogen peroxide), heat (60 °C) and UV light (254 nm). Degradation was observed under base hydrolysis to give the starting material used during the synthesis of bicalutamide. The degraded samples were assayed and gave a mass balance greater than 99.5%, thus proving the stability-indicating power of the developed method. The method was validated with respect to linearity, accuracy, precision and robustness.     

A Stability-Indicating LC Assay Method for Pemetrexed Disodium

This present work describes the development of a stability-indicating high performance liquid chromatographic method for the quantitative determination of pemetrexed disodium. Pemetrexed disodium is an antifolate antineoplastic agent that exerts its action by disrupting folate-dependent metabolic processes essential for cell replication. The chromatographic separation was achieved on an ACE 3 C18 HPLC column using a mobile phase consisting of a mixture of buffer (solvent A) and organic modifier acetonitrile (solvent B). Forced degradation studies were performed on bulk sample of pemetrexed disodium using acid (0.5 N hydrochloric acid), base (0.5 N sodium hydroxide), oxidation (10% v/v hydrogen peroxide), heat (60 °C) and UV light (254 nm). Degradation of the drug substance was observed in base hydrolysis. Degradation product formed under acid and base hydrolysis was found to be starting material. The stressed samples were assayed using the developed LC method and the mass balance found was close to 99.5%, thus proving its stability-indicating power. The developed method was validated with respect to linearity, accuracy, precision and robustness.     

A Stability-Indicating LC Method for Assay of Topotecan Hydrochloride

This paper describes the development of a stability-indicating high-performance liquid chromatographic (HPLC) method for quantitative determination of topotecan hydrochloride, a semi-synthetic derivative of camptothecin and anti-tumor drug with topoisomerase I-inhibitory activity. Chromatographic separation was achieved on a C₁₈ column with a mixture of phosphate buffer and acetonitrile as mobile phase. The method was validated for linearity, accuracy, precision, and robustness. Forced degradation studies were performed by treating bulk samples of topotecan hydrochloride with acid (0.5 M hydrochloric acid), base (0.5 M sodium hydroxide), oxidizing agent (10% v/v hydrogen peroxide), heat (60 °C), and UV light (254 nm).     

Determination of Atazanavir in the Presence of its Degradation Products by a Stability-Indicating LC Method

A forced degradation study was successfully applied for the development of a stability-indicating assay method for the determination of atazanavir in presence of its degradation products. The method was developed and optimized by analyzing the forcefully degraded samples. Degradation of the drug was done under acidic, alkaline, oxidative, photolytic and thermal stress conditions. The proposed method was able to resolve all of the possible degradation products formed during the stress studies. The major impurities are generated in acidic and alkaline conditions. The product was found stable under thermal, photolytic and oxidative conditions. The developed method was validated for determination of atazanavir, and the method was found to be equality applicable to study the impurities formed during routine and forced degradation of atazanavir.     

Stability-Indicating HPTLC Method for Simultaneous Determination of Ezetimibe and Simvastatin

Simvastatin and ezetimibe are used to treat hyperlipidemia. A simple, selective and stability-indicating HPTLC method has been established for analysis of simvastatin and ezetimibe. The method has been validated so that both drugs can routinely be analyzed simultaneously. The method uses aluminum-backed silica gel 60F₂₅₄ TLC plates as stationary phase with n-hexane–acetone 6:4 (v/v) as mobile phase. Densitometric analysis of both drugs was carried out in absorbance mode at 234 nm. This system was found to give compact bands for simvastatin and ezetimibe (R _F 0.39 ± 0.05 and 0.50 ± 0.05, respectively). Linear relationships were obtained between response and amount of drug in the range 200–1,600 ng per band with high correlation coefficients (r ² = 0.9917 ± 0.0018 for simvastatin and r ² = 0.9927 ± 0.0021 for ezetimibe). The method was validated for precision, robustness, and recovery. The limits of detection and quantitation were 25 and 150 ng per band, respectively. Simvastatin and ezetimibe were subjected degradation by acid, pH 6.8 phosphate buffer, oxidation, dry heat, and wet heat. The degradation products were well resolved from the pure drug with significantly different R _F values. Because the method could effectively separate the drug from its degradation products, it can be used for stability-indicating analysis.     

Development and Validation of a Stability-Indicating LC Method to Quantify Hydrochlorothiazide in Oral Suspension for Pediatric Use

A stability-indicating reversed-phase high-performance liquid chromatography (LC) method was developed and validated for the determination of hydrochlorothiazide in an oral suspension. Isocratic chromatography was performed on a C₁₈ column with 0.1 M sodium phosphate buffer pH 3.0/acetonitrile (70:30 v/v) as mobile phase, at a flow rate of 1.3 mL min⁻¹, and UV detection at 254 nm. The method was linear (r ² = 0.9998), accurate (mean recovery = 100.3%), and precise (RSD <2%). It was also validated for specificity and robustness. The method was successfully applied for the quality control analysis of a new pharmaceutical formulation of HCTZ for pediatric use.     

Development of a Validated Stability-Indicating LC Method for Nitazoxanide in Pharmaceutical Formulations

A reversed-phase liquid chromatographic (LC) method was developed for the assay of nitazoxanide (NTZ) in solid dosage formulations. An isocratic LC separation was performed on a Phenomenex Synergi Fusion C₁₈ column (250 mm × 4.6 mm, i.d., 4 μm particle size) using a mobile phase of 0.1% o-phosphoric acid solution, pH 6.0: acetonitrile (45:55, v/v) at a flow rate of 1.0 mL min⁻¹. Detection was achieved with a photodiode array detector at 240 nm. The detector response for NTZ was linear over the concentration range from 2 to 100 μg mL⁻¹ (r = 0.9999). The specificity and stability-indicating capability of the method were proved using stress conditions. The RSD values for intra-day precision were less than 1.0% for tablets and powder for oral suspension. The RSD values for inter-day precision were 0.6 and 0.7% for tablets and powder for oral suspension. The accuracy was 100.4% (RSD = 1.8%) for tablets and 100.9% (RSD = 0.3%) for powder for oral suspension. The limits of quantitation and detection were 0.4 and 0.1 μg mL⁻¹. There was no interference of the excipients on the determination of the active pharmaceutical ingredient. The proposed method was precise, accurate, specific, and sensitive. It can be applied to the quantitative determination of drug in tablets and powder for oral suspension.     

A Stability-Indicating LC Method for Candesartan Cilexetil

An isocratic reversed-phase liquid chromatographic method has been developed for quantitative determination of candesartan cilexetil, used to treat hypertension, in the bulk drug and in pharmaceutical dosage forms. The method is also applicable to analysis of related substances. Chromatographic separation was achieved on a 250 mm × 4.6 mm, 5 μm particle, CN column with a 50:50 (v/v) mixture of phosphate buffer, pH 3.0, and acetonitrile as mobile phase. The flow rate was 1.0 mL min⁻¹ and the detection wavelength was 210 nm. Resolution of candesartan cilexetil and six potential impurities was greater than 2.0 for all pairs of compounds. The drug was subjected to hydrolytic, oxidative, photolytic, and thermal stress and substantial degradation occurred in alkaline and acidic media and under oxidative and hydrolytic stress conditions. The major product obtained as a result of basic hydrolysis was different from that produced by acid hydrolysis and aqueous hydrolysis. The stress samples were assayed against a reference standard and the mass balance was found to be close to 99.6%. The method was validated for linearity, accuracy, precision, and robustness.     

A Stability-Indicating LC Method for the Simultaneous Determination of Telmisartan and Ramipril in Dosage Form

A simple, rapid, and precise method is developed for the quantitative simultaneous estimation of telmisartan and ramipril in combined pharmaceutical dosage form. A chromatographic separation of the two drugs was achieved with an ACE 5 C₁₈, 25-cm analytical column using buffer–acetonitrile (55:45 v/v). The buffer used in mobile phase contains 0.1 M sodium perchlorate monohydrate in double distilled water pH adjusted 3.0 with trifluoroacetic acid. The instrumental settings are flow rate of 1.5 mL min⁻¹, column temperature at 30 °C, and detector wavelength of 215 nm using a photodiode array detector. The resolution between ramipril and telmisartan were found to be more than 5. Theoretical plates for ramipril and telmisartan were 13,022 and 6,629. Tailing factor for ramipril and telmisartan was 0.94 and 0.98. Telmisartan, ramipril and their combination drug product were exposed to thermal, photolytic, hydrolytic and oxidative stress conditions, and the stressed samples were analysed by the proposed method. Peak homogeneity data of telmisartan and ramipril is obtained using photodiode array detector, in the stressed sample chromatograms, demonstrated the specificity of the method for their estimation in presence of degradants. The described method shows excellent linearity over a range of 20–400 μg mL⁻¹ for telmisartan and 2.5–50 μg mL⁻¹ for ramipril. The correlation coefficient for telmisartan and ramipril are 1. The relative standard deviation for six measurements in two sets of each drug in tablets was always less than 2%. The proposed method was found to be suitable and accurate for quantitative determination and the stability study of telmisartan and ramipril in pharmaceutical preparations.     

A Rapid Stability-Indicating LC Method for Ziprasidone Hydrochloride

A simple and rapid reversed-phase liquid chromatographic method was developed for the related substances determination and quantitative evaluation of ziprasidone hydrochloride, which is used as an antipsychotic agent. Forced degradation studies were performed on bulk sample of ziprasidone hydrochloride using acid, base, oxidative hydrolysis, thermal stress and photolytic degradation. Mild degradation of the drug substance was observed during thermal stress and considerable degradation observed during base hydrolysis. The chromatographic method was fine tuned using the samples generated from forced degradation studies. Good resolution between the peaks corresponds to synthetic impurities and degradation products from the analyte were achieved on YMC Pack Pro C18 column using the mobile phase consists of a mixture of 0.05% v/v of phosphoric acid in water and acetonitrile. The stressed test solutions were assayed against the qualified working standard of ziprasidone hydrochloride and the mass balance in each case was close to 99.7% indicating that the developed method was stability-indicating. Validation of the developed method was carried out as per ICH requirements.     

Stress Degradation Studies on Tadalafil and Development of a Validated Stability-Indicating LC Assay for Bulk Drug and Pharmaceutical Dosage Form

A stability-indicating HPLC assay method was developed for the quantitative determination of tadalafil in bulk samples and in pharmaceutical dosage forms in the presence of the degradation products. It involved a 250 mm × 4.6 mm, 5 μm C-18 column. The gradient LC method employs solution A and B as mobile phase. Solution A contains a mixture of buffer (phosphate buffer and tetra-n-butyl ammonium hydrogen sulfate) pH 2.5: acetonitrile (80:20, v/v) and solution B contains a mixture of water: acetonitrile (20:80, v/v). The flow rate was 1.0 mL min⁻¹ and the detection wavelength was 220 nm. The retention time of tadalafil is about 17 min. Tadalafil was subjected to different ICH prescribed stress conditions. Degradation was found to occur in hydrolytic and to some extent in oxidative stress conditions, while the drug was stable to photolytic and thermal stress. The drug was particularly labile under alkaline hydrolytic conditions. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. The assay of stress samples was calculated against a qualified reference standard and the mass balance was close to 99.5%. The developed RP-LC method was validated with respect to linearity, accuracy, precision and ruggedness.     

Development and Validation of a High Performance Liquid Chromatographic Method for the Determination of Voriconazole Content in Tablets

This paper describes the validation of an isocratic HPLC method for the assay of voriconazole in tablets. The method employs a Merck LiChrospher^? 100 RP-8 (125 × 4.6 mm I.D., 5 μm particle size) column, with a mobile phase of methanol : triethylamine solutions 0.6 %, pH 6.0 (50:50, v/v) and UV detection at 255 nm. A linear response (r > 0.9999) was observed in the range of 20.0–100.0 μg mL⁻¹. The method showed good recoveries (average 100.4%) and the relative standard deviation intra and inter-day were ≤ 1.0 %. Validation parameters as specificity and robustness were also determined. The method can be used for both quality control assay of voriconazole in tablets and for stability studies as the method separates voriconazole from its degradation products and tablet excipients.     

Determination and Validation of an LC Method for Fluvoxamine in Tablets

A new, simple, rapid and specific reversed-phase high-performance liquid chromatography (HPLC) method was developed and validated for the determination of fluvoxamine in pharmaceutical dosage forms. The HPLC separation was achieved on a C₁₈ μ-Bondapack column (250 mm × 4.6 mm) using a mobile phase of acetonitrile–water (80:20, v/v) at a flow rate of 1 mL min⁻¹. Proposed method is based on the derivatization of fluvoxamine with 1,2-naphthoquinone-4-sulphonic acid sodium salt (NQS) in borate buffer of pH 8.5 to yield a orange product. The HPLC method is based on measurement of the derivatized product using UV-visible absorbance detection at 450 nm. The method was validated for specificity, linearity, precision, accuracy, robustness. The degree of linearity of the calibration curves, the percent recoveries of fluvoxamine, the limit of detection and quantification, for the HPLC method were determined. The assay was linear over the concentration range of 45–145 ng mL⁻¹ (r = 0.9999). Limit of detection and quantification for fluvoxamine were 15 and 50 ng mL⁻¹, respectively. The results of the developed procedure (proposed method) for fluvoxamine content in tablets were compared with those by the official method. The method was found to be simple, specific, precise, accurate, reproducible and robust.     

A Validated, Stability-Indicating, LC Method for Rabeprazole Sodium

We present a simple and reliable method for simultaneous determination of voriconazole and its main metabolite resulting from N-oxidation (UK-121,265), in human plasma. The work-up procedure used acetonitrile and potassium salts to precipitate plasma proteins. No internal standard was used. The chromatographic system used a LiChroCART^® 250-4 cartridge packed with LiChrospher^® 100 RP-8 (diameter particules, 5 μm). The UV monochromatic detector was set on 260 nm. The mobile phase consisted of a 60/40 (v/v) mixture of acetonitrile and water. The flow rate was 1 mL min^?1. The retention times for voriconazole and its metabolite were 8.98 and 4.02 min respectively, and total run time was 12 min. The linearity of the method was investigated from 0.31 to 10.0 mg L^?1; the lowest limit of quantification was 0.30 mg L^?1. Precision ranged from 2.41% to 6.32% for voriconazole and 0.80% to 11.6% for the N-oxide voriconazole metabolite. Accuracy was between 93.0% and 101% for voriconazole and 90.0% and 101% for the N-oxide voriconazole metabolite. This rapid and accurate method could be interesting to investigate metabolite/voriconazole ratio with respect to CYP2C19 genetic status and CYP3A4 activity changes.     

A Stability-Indicating LC Assay and Degradation Behavior of Temozolomide Drug Substances

This study deals with a stability indicating HPLC reverse phase method for quantitative determination of temozolomide. A chromatographic separation was achieved on an Inertsil ODS 3V, 250 × 4.6 mm ID, 5 μm column using mobile phase A (buffer 5 mL glacial acetic acid in 1,000 mL of Milli Q water ) and mobile phase B (methanol). Forced degradation studies were performed on bulk sample of temozolomide using acid (0.5 N hydrochloric acid), base (0.5 N sodium hydroxide), oxidation (10% v/v hydrogen peroxide), heat (60 °C) and UV light (254 nm). Degradation of the drug substance was observed in base hydrolysis and oxidation. Degradation product formed under these conditions was found to be Imp-A. When the stress samples were assayed, the mass balance was close to 99.5%. The sample solution was stable up to 48 h at 5 °C and mobile phase was found to be stable up to 48 h at 25 °C. The developed method was validated with respect to linearity, accuracy, precision, robustness and forced degradation studies prove the stability indicating power of the method.     

Method Development and Validation of a Rapid Determination of Ropinirole in Tablets by LC-UV

A reversed-phase high-performance liquid chromatographic (HPLC) method with UV detection was developed and validated for the determination of ropinirole (ROP) in tablets. The assay utilized UV detection at 250 nm and a Luna CN column (250 × 4.6 mm I.D, 5 μm). The mobile phases were comprised of acetonitrile: 10 mM nitric acid (pH 3.0) (75:25, v/v). Validation experiments were performed to demonstrate linearity, accuracy, precision, limit of quantitation (LOQ), limit of detection (LOD), and robustness. The method was linear over the concentration range of 0.5–10.0 μg mL⁻¹. The method showed good recoveries (99.75–100.20%) and the relative standard deviations of intra and inter-day assays were 0.38–1.69 and 0.45–1.95%, respectively. The method can be used for quality control assay of ropinirole.     

Development and Validation of a Liquid Chromatography-Tandem Mass Spectrometry Method for the Determination of Ezetimibe in Human Plasma and Pharmaceutical Formulations

An analytical method based on liquid chromatography-tandem mass spectrometry (LC-MS-MS) was developed and validated for the determination of ezetimibe in human plasma. Ezetimibe and etoricoxib (internal standard) were extracted from the plasma by liquid-liquid extraction and separated on a C₁₈ analytical column (50 × 3.0 mm I.D.) with acetonitrile:water (85:15, v/v) as mobile phase. Detection was carried out by positive electrospray ionization (ESI+) in multiple reaction monitoring (MRM) mode. The chromatographic separation was obtained within 2.0 min and was linear in the concentration range of 0.25–20ng mL⁻¹ for free ezetimibe and of 1–300ng mL⁻¹ for total ezetimibe. The mean extraction recoveries for free and total ezetimibe from plasma were 96.14 and 64.11%, respectively. Method validation investigated parameters such as linearity, precision, accuracy, specificity and stability, giving results within the acceptable range. The proposed method was successfully applied to the quantitation of ezetimibe and its glucuronide in human plasma to support clinical and pharmacokinetic studies. Moreover, the method was used for the quality control analysis of pharmaceutical dosage forms. Revised: 4 January and 3 February 2006     

Stability-Indicating LC Method for the Determination of Lacidipine in Tablets. Application to Degradation Kinetics and Content Uniformity Testing

A simple, stability-indicating, reversed-phase liquid chromatographic method was developed for the determination of lacidipine in the presence of its degradation products. The analysis was carried out using a 150 mm × 4.6 mm i.d., 5 μm particle size Nucleodur MN-C18 column. Mobile phase containing a mixture of acetonitrile and 0.02 M phosphate buffer (70:30) at pH = 5.0 was pumped at a flow rate of 1 mL min^?1 with UV-detection at 254 nm. The method showed good linearity in the range of 0.06–15 μg mL^?1 with a limit of detection (S/N = 3) of 0.016 μg mL^?1 (3.5 × 10^?8 M). The suggested method was successfully applied for the analysis of lacidipine in bulk and in commercial tablets with average recoveries of 100.19 ± 0.81% and 100.05 ± 0.69%, respectively. The results were favorably compared to those obtained by a reference method. The suggested method was utilized to investigate the kinetics of alkaline, acidic, peroxide and photo-induced degradation of the drug. The apparent first-order rate constant, half-life times and activation energies of the degradation process were calculated. The pH profile curve was derived. The proposed method was successfully applied to the content uniformity testing of tablets.     

Stability-Indicating LC Method for the Determination of Lacidipine in Tablets. Application to Degradation Kinetics and Content Uniformity Testing

A simple, stability-indicating, reversed-phase liquid chromatographic method was developed for the determination of lacidipine in the presence of its degradation products. The analysis was carried out using a 150 mm × 4.6 mm i.d., 5 μm particle size Nucleodur MN-C18 column. Mobile phase containing a mixture of acetonitrile and 0.02 M phosphate buffer (70:30) at pH = 5.0 was pumped at a flow rate of 1 mL min⁻¹ with UV-detection at 254 nm. The method showed good linearity in the range of 0.06–15 μg mL⁻¹ with a limit of detection (S/N = 3) of 0.016 μg mL⁻¹ (3.5 × 10⁻⁸ M). The suggested method was successfully applied for the analysis of lacidipine in bulk and in commercial tablets with average recoveries of 100.19 ± 0.81% and 100.05 ± 0.69%, respectively. The results were favorably compared to those obtained by a reference method. The suggested method was utilized to investigate the kinetics of alkaline, acidic, peroxide and photo-induced degradation of the drug. The apparent first-order rate constant, half-life times and activation energies of the degradation process were calculated. The pH profile curve was derived. The proposed method was successfully applied to the content uniformity testing of tablets.