A Stability-Indicating LC Method for Rimonabant

A simple, inexpensive and rapid isocratic LC method has been developed for the quantative determination of Rimonabant, an anti-obesity drug. The method can also be employed for the determination of Rimonabant and its impurities in the bulk drug. Degradation studies were performed on the bulk drug by heating to 60 °C, exposure to UV light at 254 nm, acid (0.5 N hydrochloric acid), base (0.5 N sodium hydroxide) and aqueous hydrolysis and oxidation with 3.0% v/v hydrogen peroxide. Considerable degradation was observed under oxidation conditions. Good resolution between the peaks corresponding to impurities produced during synthesis, degradation products and the analyte was achieved on a Phenomenex Gemini C18 LC column using a mobile phase consisting of a mixture of aqueous potassium dihydrogen phosphate (pH 3.0) and acetonitrile. The degradation samples were assayed against the reference standard of Rimonabant and the mass balance in each case was close to 99.5%. Validation of the method was carried out as per ICH requirements.     

Stability-Indicating LC Method for the Determination of Olmesartan in Bulk Drug and in Pharmaceutical Dosage Form

A novel stability-indicating LC assay method was developed and validated for quantitative determination of olmesartan in bulk drugs and in pharmaceutical dosage form in the presence of degradation products generated from forced degradation studies. An isocratic, reversed phase LC method was developed to separate the drug from the degradation products, using an Ace5-C18 (250 mm × 4.6 mm, 5 μm) column, and 50 mM ammonium acetate (pH-5.5 by acetic acid) and acetonitrile (70:30 v/v) as a mobile phase. The detection was carried out at the wavelength of 235 nm. The olmesartan was subjected to stress conditions of hydrolysis (acid, base), oxidation, photolysis and thermal degradation. Degradation was observed for olmesartan in acid, base and in 30% H₂O₂ conditions. The drug was found to be stable in the other stress conditions attempted. The degradation products were well resolved from the main peak. The percentage recovery of olmesartan ranged from (99.89 to 100.95%) in pharmaceutical dosage form. The developed method was validated with respect to linearity, accuracy (recovery), precision, specificity and robustness. The forced degradation studies prove the stability-indicating power of the method.

     

Stability Indicating LC Assay Method for the Determination of Famciclovir in Bulk Drug and Pharmaceutical Dosage Forms

A simple isocratic stability indicating LC method was developed and validated for the determination of famciclovir in bulk drug and pharmaceutical dosage form. A mixture of 0.05 M potassium dihydrogen orthophosphate buffer and acetonitrile (80:20 v/v) was used as mobile phase at a flow rate of 1.0 mL min^?1. Hypersil BDS C18 (250 mm × 4.6 mm × 5 μm) column was used and the eluents were monitored at 220 nm. Forced degradation studies were performed for famciclovir active substance, reconstituted matrix and 500 mg tablets using the parameters like acid, base, peroxide, temperature, light, and relative humidity. Peak purity index was checked using PDA detection to demonstrate the specificity and stability indicating nature of the method. The developed method was validated for precision, ruggedness, linearity, LOD, LOQ, range, robustness and accuracy. The developed method can be used for regular quality control and stability study applications of famciclovir bulk drug and tablet dosage forms.     

A Stability Indicating LC Method for Deferasirox in Bulk Drugs and Pharmaceutical Dosage Forms

A novel, sensitive, stability indicating RP-LC method has been developed for the quantitative determination of deferasirox, its related impurities in both bulk drugs and pharmaceutical dosage forms. Efficient chromatographic separation was achieved on a C18 stationary phase with simple mobile phase combination delivered in an isocratic mode and quantitation was by ultraviolet detection at 245 nm. The mobile phase consisted of buffer, acetonitrile and methanol (50:45:5, v/v) delivered at a flow rate of 1.0 mL min^?1. Buffer consisted of 10 mM potassium dihydrogen orthophosphate monohydrate, pH adjusted to 3.0 by using orthophosphoric acid. In the developed LC method the resolution (R _s ) between deferasirox and its four potential impurities was found to be greater than 2.0. Regression analysis showed an r value (correlation coefficient) greater than 0.999 for deferasirox and its four impurities. This method was capable to detect all four impurities of deferasirox at a level of 0.002% with respect to test concentration of 0.5 mg mL^?1 for a 10 μL injection volume. The inter- and intra-day precision values for all four impurities and for deferasirox was found to be within 2.0% RSD. The method showed good and consistent recoveries for deferasirox in bulk drugs (98.3–101.1%), pharmaceutical dosage forms (100.2–103.1%) and for its all the four impurities (99.7–102.1%). The test solution was found to be stable in methanol for 48 h. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. Considerable degradation was found to occur in acid stress hydrolysis. The stress samples were assayed against a qualified reference standard and the mass balance was found close to 99.95%. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness.     

Stress Degradation Behavior of Entacapone and Development of LC Stability-Indicating Related Substances and Assay Method

A new, sensitive, stability indicating gradient RP-LC related substances and assay method has been developed for the quantitative determination of entacapone in bulk drugs. Efficient chromatographic separation was achieved on a C18 stationary phase with simple mobile phase combination of buffer and acetonitrile. Buffer consisted of 0.1% orthophosphoric acid, delivered in a gradient mode and quantitation was carried out using ultraviolet detection at 220 nm with a flow rate of 1.5 mL min^?1. In the developed LC method the resolution (R _s ) between entacapone and its three potential process impurities were found to be >2.0. Regression analysis showed an r ² value (correlation coefficient) >0.99 for entacapone and its three potential impurities. This method was capable to detect all three process impurities of entacapone at a level of 0.003% with respect to test concentration of 0.5 mg mL^?1 for a 20 μL injection volume. The inter- and intra-day precision values for all three impurities and for entacapone was found to be within 2.0% RSD. The method has shown good and consistent recoveries for entacapone in bulk drugs (99.2–101.5%) and its three impurities (99.5–102.2%). The test solution was found to be stable in diluent for 48 h. The drug substances were subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. Considerable degradation was found to occur in acid stress, base stress and oxidative conditions. The stressed test solutions were assayed against the qualified working standard of entacapone and the mass balance in each case was close to 99.7% indicating that the developed method was stability-indicating. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness.     

Stability-Indicating TLC Method for the Determination of Dutasteride in Pharmaceutical Dosage Forms

A simple, sensitive, selective, precise and stability-indicating thin-layer chromatographic method for determination of dutasteride both as a bulk drug and as pharmaceutical tablets was developed and validated as per the International Conference on Harmonization guidelines. The method employed thin-layer chromatography aluminium plates precoated with silica gel 60F₂₅₄ as the stationary phase and the mobile phase consisted of acetonitrile:methanol:dichloromethane in the ratio of 2.0:1.0:2.0, v/v/v. This solvent system was found to give compact spots for dutasteride (R _f value of 0.64 ± 0.02). Densitometric analysis of dutasteride was carried out in the absorbance mode at 244 nm. The linear regression analysis data for the calibration plots showed good linear relationship with r = 0.9943 with respect to peak area in the concentration range of 100–600 ng per band. The method was validated for precision, accuracy, ruggedness and recovery. The limits of detection and quantitation were 7.54 and 22.85 ng per band, respectively. Dutasteride was subjected to acid and alkali hydrolysis, oxidation, photo degradation, dry heat and wet heat treatment. The drug undergoes degradation under acidic, basic conditions, photolytic, oxidative and upon wet and dry heat treatment. The degraded products were well separated from the pure drug. The statistical analysis proves that the developed method for quantification of dutasteride as bulk drug and from pharmaceutical tablets is reproducible and selective. As the method could effectively separate the drug from its degradation products, it can be employed as stability-indicating.     

Development and Validation of a Stability Indicating LC Method for the Determination of Faropenem in Pharmaceutical Formulations

A simple, selective and sensitive stability indicating LC method has been developed and validated for the determination of faropenem in bulk drug and pharmaceutical formulations in the presence of degradation products. The separation was achieved by using an isocratic mobile phase mixture of acetate buffer of pH 3.5 and methanol (65:35, v/v) and 250 mm × 4.6 mm I.D., 5 μm particle size SGE make Wakosil C-18 AR column at flow rate of 1.0 mL min^?1 with detection at 305 nm. The retention time of faropenem is 6.63 min and was linear in the range of 5–75 μg mL^?1 (r = 0.9999). The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation and was found to be unstable in all the stress conditions. The proposed method was successfully employed for quantification of faropenem in bulk drug and its pharmaceutical formulations.     

A Validated Stability Indicating LC Method for Mitotane in Bulk Drugs and Pharmaceutical Dosage Forms

A novel, sensitive, stability indicating RP-LC method has been developed for the quantitative determination of mitotane, its impurity in both bulk drugs and pharmaceutical dosage forms. Efficient chromatographic separation was achieved using a C18 stationary phase with simple mobile phase combination delivered in an isocratic mode and quantitation was by ultraviolet detection at a wavelength of 230 nm. The mobile phase consisted of buffer and acetonitrile (25:75, v/v) delivered at a flow rate of 1.0 mL min^?1. Buffer consisted of 10 mM potassium dihydrogen orthophosphate monohydrate, pH adjusted to 2.5 by orthophosphoric acid. In the developed LC method the resolution (R _s ) between mitotane and its impurity namely Imp-1 was found to be greater than 2.5. Regression analysis shows an r value (correlation coefficient) greater than 0.999 for mitotane and its impurity. This method was capable to detect the impurity of mitotane at a level of 0.003% with respect to test a concentration of 0.2 mg mL^?1 for a 10 μL injection volume. The inter- and intra-day precision values for mitotane and its impurity was found to be within 2.0% RSD. The method has shown good and consistent recoveries for mitotane in bulk drugs (99.2–101.5%), pharmaceutical dosage forms (98.2–103.1%) and for its impurity (99.7–102.1%). The test solution was found to be stable in diluent for 48 h. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. Considerable degradation was found to occur in basic stress hydrolysis. The stress samples were assayed against a qualified reference standard and the mass balance was found close to 99.97%. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness.     

Stability-Indicating HPTLC Method for the Determination of Tamsulosin in Pharmaceutical Dosage Forms

A simple, sensitive, selective, precise and stability indicating high-performance thin-layer chromatographic method was developed for the determination of tamsulosin (TAM) in bulk and tablet formulation. Validation was carried out in compliance with International Conference on Harmonization guidelines. The method employed thin-layer chromatography aluminium plates pre-coated with silica gel 60F₂₅₄ as the stationary phase and the mobile phase consisted of acetonitrile/methanol/dichloromethane (2.0: 1.0: 2.0, v/v/v). This solvent system was found to give compact spots for tamsulosin (R _f = 0.27 ± 0.02). Densitometric analysis of TAM was carried out in the absorbance mode at 286 nm. Linear regression analysis showed good linearity (r ² = 0.9993) with respect to peak area in the concentration range of 300–800 ng per band. The method was validated for precision, accuracy, ruggedness and recovery. Limits of detection and quantitation were 8.49 and 25.72 ng per band, respectively. TAM was subjected to acid and alkali hydrolysis, oxidation, photo degradation, dry heat and wet heat treatment. The drug underwent degradation under acidic, basic and photolytic conditions. The degraded products were well separated from the pure drug. Statistical analysis proved that the developed method, used for quantification of TAM as a bulk drug and present in pharmaceutical tablets, was reproducible and selective.     

Validation of an HPLC Method for the Determination of Valdecoxib and its Degradation Product: a Mixture of α- and β-n-Lactosyl Sulfonamide Anomers

An HPLC method has been developed for the separation of valdecoxib and a degradation product consisting of α and β-N-lactosyl sulfonamide, i.e. α and β anomers (SC-77852). Best results were achieved with a Chromolith Performance RP-18e column (100 mm × 4.6 mm), macropore size 2 μm, mesopore size 13 nm, with an eluent of methanol:water containing a 1% solution of TEA (36:64 v/v), pH 7.4 (adjusted with 85% orthophosphoric acid), at 22 °C. Detection was at 220 nm. The method was validated for its selectivity, linearity, precision (repeatability) and robustness. Quantitation and detection limits were determined for both valdecoxib and SC-77852. Method robustness was further evaluated by performing 2³ full factorial design experiments. The final step, optimisation of the variables, was performed using response surface design. The validated method was used for assay of valdecoxib and SC-77852 in Bextra^® film-coated tablets.     

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.     

Separation and Characterization of New Forced Degradation Products of Macitentan: A Dual Endothelin Receptor Antagonist

Macitentan (MCT) is an endothelin receptor antagonist used for the treatment of pulmonary arterial hypertension. In the present study, MCT was subjected to forced degradation as per ICH guidelines. The drug degraded extensively in acidic, basic as well as neutral hydrolytic conditions and seven degradation products (DPs) were formed. All these DPs were selectively separated using high-performance liquid chromatography (HPLC) with a stationary phase of Inertsil C₁₈ column (150 × 4.6 mm, 5 μm) and a mobile phase consisting of gradient mixture of 0.02% trifluoroacetic acid (TFA) and acetonitrile (ACN). The developed HPLC method was transferred to LC–ESI–QTOF–MS/MS for identification of DPs. The final mass spectrometric conditions were optimized for better ionization of drug and DPs with optimum mass signal sensitivity. All the formed DPs were new and well separated with sufficient resolution. The developed HPLC method was validated as per ICH-guidelines and can be used in drug testing labs for determination of quality of MCT in bulk and finished formulations.     

Stress Degradation Studies on Aripiprazole and Development of a Validated Stability Indicating LC Method

The present paper describes the development of a stability indicating reversed phase column liquid chromatographic method for aripiprazole in the presence of its impurities and degradation products generated from forced decomposition studies. The drug substance was subjected to stress conditions of aqueous hydrolysis, oxidative, photolytic and thermal stress degradation. The degradation of aripiprazole was observed under acid hydrolysis and peroxide. The drug was found to be stable to other stress conditions attempted. Successful separation of the drug from the synthetic impurities and degradation products formed under stress conditions was achieved on an Inertsil phenyl column using a mixture of 0.2% trifluoroacetic acid and acetonitrile (55:45, v/v). The developed LC method was validated with respect to linearity, accuracy, precision, specificity and robustness. The assay method was found linear in the range of 25–200 μg mL^?1 with a correlation coefficient of 0.9999 and the linearity of the impurities were established from LOQ to 0.3%. Recoveries of the assay and impurities were found between 97.2 and 104.6%. The developed LC method for the related substances and assay determination of aripiprazole can be used to evaluate the quality of regular production samples. It can also be used to test the stability samples of aripiprazole. To the best of our knowledge, the validated stability indicating LC method which separates all the impurities disclosed in this investigation was not published elsewhere.     

Stress Degradation Behavior of Desipramine Hydrochloride and Development of Suitable Stability-Indicating LC Method

A simple reverse phase liquid chromatographic method was developed for the quantitative determination of desipramine hydrochloride and its related impurities in bulk drugs which is also stability-indicating. During the forced degradation at hydrolysis, oxidative, photolytic and thermal stressed conditions, the degradation results were only observed in the oxidative stress condition. The blend of the degradation product and potential impurities were used to optimize the method by an YMC Pack Pro C₁₈ stationary phase. The LC method employs a linear gradient elution with the water–acetonitrile–trifluoroacetic acid as mobile phase. The flow rate was 1.0 mL min^?1 and the detection wavelength 215 nm. The stressed samples were quantified against a qualified reference standard and the mass balance was found close to 99.0% (w/w) when the response of the degradant was considered to be equal to the analyte (i.e. desipramine). The developed RP-LC method was validated in agreement with ICH requirements.     

A Simple and Rapid Stability Indicating LC Determination of Ciprofloxacin Hydrochloride as a Bulk Drug and from Formulations

The present research work discusses the development of a stability indicating reversed phase LC method for determination of ciprofloxacin hydrochloride as a bulk drug and from formulations. The mobile phase selected was water-acetonitrile-triethylamine 75:25:0.1 (v/v/v) adjusted to pH 4.0 with o-phosphoric acid. The calibration curve of the drug was linear in the range 0.25–15 μg mL^?1. The method was accurate and precise with limits of detection and quantitation of 8.01 and 26.7 ng, respectively. Mean percent recovery was 100.71%. The method was used for analysis of ciprofloxacin hydrochloride from pharmaceutical formulations in the presence of its degradation products and commonly used excipients.     

Stress Degradation Behavior of Entacapone and Development of LC Stability-Indicating Related Substances and Assay Method

A new, sensitive, stability indicating gradient RP-LC related substances and assay method has been developed for the quantitative determination of entacapone in bulk drugs. Efficient chromatographic separation was achieved on a C18 stationary phase with simple mobile phase combination of buffer and acetonitrile. Buffer consisted of 0.1% orthophosphoric acid, delivered in a gradient mode and quantitation was carried out using ultraviolet detection at 220 nm with a flow rate of 1.5 mL min⁻¹. In the developed LC method the resolution (R _s ) between entacapone and its three potential process impurities were found to be >2.0. Regression analysis showed an r ² value (correlation coefficient) >0.99 for entacapone and its three potential impurities. This method was capable to detect all three process impurities of entacapone at a level of 0.003% with respect to test concentration of 0.5 mg mL⁻¹ for a 20 μL injection volume. The inter- and intra-day precision values for all three impurities and for entacapone was found to be within 2.0% RSD. The method has shown good and consistent recoveries for entacapone in bulk drugs (99.2–101.5%) and its three impurities (99.5–102.2%). The test solution was found to be stable in diluent for 48 h. The drug substances were subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. Considerable degradation was found to occur in acid stress, base stress and oxidative conditions. The stressed test solutions were assayed against the qualified working standard of entacapone and the mass balance in each case was close to 99.7% indicating that the developed method was stability-indicating. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness.

     

Stability-Indicating LC Determination of a New Antihypertensive, Olmesartan Medoxomil in Tablets

A stability-indicating liquid chromatographic method was developed and validated for quantitative determination of olmesartan medoxomil (OLM) in coated tablets in the presence of degradation products generated under stress conditions. An isocratic LC separation was performed using a Phenomenex RP-18 column using a mobile phase consisting of water:triethylamine:acetonitrile (60:0.3:40 v/v/v, pH adjusted to 6.3 with phosphoric acid). The flow rate was 1.2 mL min^?1 and the detection was achieved with a photodiode array detector set at 257 nm. The response was linear over a range of 10.0 to 30.0 μg mL^?1 (r = 0.9999). The specificity and stability-indicating capability of the method was verified subjecting the reference substance and drug product to hydrolytic, oxidative, photolytic, and thermal stress conditions. The method showed a good and consistent recovery (100.2%) with low intra- and inter-day relative standard deviation (RSD) (≤1.0%). A considerable degradation occurred in all stress conditions and the degradation product was well resolved from the main peak. There was no interference of the excipients in the determination of the active pharmaceutical ingredient. Thus, the proposed method was found to be stability-indicating and can be used for routine analysis for quantitative determination of OLM in coated tablets without the interference of major degradation products.     

Development and Validation of a New Analytical Method for the Determination of Related Components in Tolterodine Tartarate Using LC

A novel liquid chromatographic method has been developed, and validated for the determination of tolterodine tartarate, for its potential three impurities in drug substances and drug products. Efficient chromatographic separation was achieved on a C8 stationary phase (150 × 4.6 mm, 3.5 μm particles) with a simple mobile phase combination delivered in an isocratic mode at a flow rate of 0.8 mL min^?1 and quantitation was carried out using ultraviolet detection. Microwave assisted degradation procedure was employed for stress testing studies in addition to the conventional way of a refluxing method. The results of both studies were compared. In the developed LC method, the resolution between tolterodine and its three potential impurities was found to be greater than 2.0. Regression analysis shows an r value (correlation coefficient) greater than 0.999 for tolterodine and for its three impurities. This method was capable to detect all three impurities of tolterodine at a level below 0.0038% with respect to a test concentration of 0.5 mg mL^?1 for a 10 μL injection volume. The inter- and intra-day precisions for all three impurities and for tolterodine were found to be within 1.1% RSD at its specification level. The method has shown good, consistent recoveries for tolterodine (98.9–101.6%) and for its three impurities (94.5–103.0%). The test solution was found to be stable in the diluent for 48 h. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation, as prescribed by ICH. Degradation was found to occur in alkaline stress condition, while the drug was stable to water hydrolysis, acid hydrolysis, oxidative stress, photolytic and thermal stress. The assay of stressed samples was calculated against a qualified reference standard and the mass balance was found close to 99.5%. Microwave degradations were very fast and comparable to the conventional way of the refluxing method. Robustness studies were carried out and suggested that system suitability parameters were unaffected by small changes in critical factors. The validated method was successfully applied for the determination of tolterodine tartarate in drug substances and drug products.     

Stability-Indicating HPLC and RP-TLC Determination of Cefpirome Sulfate with Kinetic Study

Two sensitive and selective stability-indicating methods were developed for the determination of the antibiotic cefpirome sulfate in bulk powder, pharmaceutical formulation and in presence of its acid, alkaline, photo- and oxidative degradation products. Method A was based on HPLC separation of cefpirome sulfate in the presence of its degradation products on a reversed phase column C18, 250 × 4.6 mm, 5-μm particle size and mobile phase consisting of 0.1 M disodium hydrogen phosphate dihydrate pH 3.9 adjusted with phosphoric acid–acetonitrile (85:15, v/v). Quantitation was achieved with UV detection at 270 nm. The linear calibration curve was in the range 5.0–50.0 μg mL^?1. Method B was based on reversed phase TLC separation of the cited drug in the presence of its degradation products followed by densitometric measurement of the intact drug at 270 nm. The separation was carried out using disodium hydrogen phosphate dihydrate 2.0 g %w/v, at pH 3.5 adjusted with phosphoric acid–acetone (15:10, v/v) as a developing system. The calibration curve was in the range of 1.0–10.0 μg/spot. The HPLC method was used to study the kinetic of cefpirome sulfate acid degradation. The results obtained were statistically analyzed and compared with those obtained by applying the official Japanese method.