Validation

A simple, precise, accurate and stability-indicating reverse phase high performance liquid chromatography (RP-HPLC) method is developed for estimation of Levosalbutamol sulphate and Budesonide in bulk and suspension for inhalation dosage form. The method employed, with reverse phase Inertsil® 5μ C18 (250 × 4.0 mm) column in an isocratic mode, with mobile phase of acetonitrile: buffer in the ratio 40:60 (%v/v). The flow rate was 0.8 ml/min and effluent was monitored at 266 nm. Retention time was found to be 3.16 min., 17.94 min. and 20.90 min. The method was validated in terms of linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ) etc. in accordance with ICH guidelines. Linear regression analysis data for the calibration plot showed that there was good linear relationship between response and concentration in the range of 25 – 150% of the working concentration (r2 > 0.999) respectively. The LOD and LOQ values for were found to be 0.43, 0.72, 0.97 and 1.24 µg/ml respectively. No chromatographic interference from placebo and degradants were found. The proposed method was successfully used for estimation of Levosalbutamol sulphate and Budesonide in bulk and suspension for inhalation dosage forms.

A specific, accurate and precise reverse phase high performance liquid chromatographic method was developed and validated for the estimation of vardenafil in bulk and pharmaceutical formulation. A Hypersil BDS C 8, 5μ column having 250 x 4.6 mm internal diameter in isocratic mode with mobile phase containing buffer and acetonitrile (60:40, v/v) was used. The flow rate was 1.0 ml/min and effluents were monitored at 240.0 nm over the concentration range of 50-250 µg/ml. The retention time obtained for vardenafil was 5.4 min. The method was validated for linearity, accuracy, repeatability, precision, specificity (in terms of acid, alkali, peroxide and thermal degradation), limit of detection, limit of quantification and ruggedness. Chromatographic peak purity data demonstrated specificity of the method to estimate vardenafil in presence of degradation products.  Limit of detection and limit of quantification were found 0.0125 µg/ml and 0.025 µg/ml respectively. Accuracy in terms of recovery of vardenafil from tablet formulation was found to be above 98 % indicating that vardenafil can be estimated from tablet dosage form without interference from the excipients. The validation data proves the potential utility of the proposed method for the quantitative determination of vardenafil in tablet formulation.

A simple, precise and accurate RP-HPLC method was developed and validated for rapid assay of Dosulepin tablet dosage form. Isocratic elution at a flow rate of 1mL/min was employed on a symmetry Chromosil C18 (250x4.6mm, 5µm in particle size) at ambient temperature. The mobile phase consisted Methanol: Acetonitrile: 0.01M Phosphate buffer in the ratio of 55:20:25 (v/v/v). The UV detection wavelength was 230nm and 20 μL sample was injected. The retention time for Dosulepin was 3.46min. The percentage RSD for precision and accuracy of the method was found to be less than 2%. The method was validated as per the ICH guidelines. The method was successfully applied for routine analysis of Dosulepin tablet dosage form and bulk drug.

A simple, fast and precise reverse phase, isocratic HPLC method was developed for the separation and quantification of Sildenafil and Duloxetine in pharmaceutical dosage form. The quantification was carried out using Symmetry C18-ODS 4.6X150mm, 3µm enhanced polar selectivity column and mobile phase comprised of potassium dihydrogen phosphate buffer and acetonitrile and water in proportion of ratio 30:65:5 and degassed under ultra-sonication. The flow rate was 0.6mL/min and the effluent was monitored at 244nm. The retention time of Sildenafil and Duloxetine were 4.3 and 3.4 respectively. The method was validated in terms of linearity, precision, accuracy, and specificity, limit of detection and limit of quantization. Linearity of Sildenafil and Duloxetine were in the range of 100 to 300µg/mL and 30 to 90µg/mL respectively. The percentage recoveries of both the drugs were 98.7% and 99.8% for Sildenafil and Duloxetine respectively from the tablet formulation.The method was found to be precise, accurate and specific during the study. The proposed method enables rapid quantification and simultaneous analysis of both drugs from commercial formulations without any excipients interference. The method can be used for routine analysis of marketed products of Sildenafil and Duloxetine in combined tablet formulation

A simple, precise, accurate and stability-indicating reverse phase high performance liquid chromatography (RP-HPLC) method is developed for estimation of Isosorbide 5-Mononitrate in bulk drug and tablet dosage form. The method employed, with reverse phase phenomenex® Luna 5µ C18 (2) 100A (250 × 4.60 mm) column in an isocratic mode, with mobile phase of methanol: water: acetonitrile in the ratio 55:28:17 (%v/v/v). The flow rate was 1.0 ml/min and effluent was monitored at 217 nm. Retention time was found to be 4.391±0.015 min. The method was validated in terms of linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ) etc. in accordance with ICH guidelines. Linear regression analysis data for the calibration plot showed that there was good linear relationship between response and concentration in the range of 1- 9 µg/ml respectively. The LOD and LOQ values for were found to be 2.5 and 10 ng/ml respectively. No chromatographic interference from tablet excipients and degradants were found. The proposed method was successfully used for estimation of Isosorbide 5-Mononitrate in tablet dosage form.

A simple, reliable, rapid, precise, sensitive and validated RP-HPLC method has been developed to determine Acetaminophen (AI) and Prasugrel (PRA) in synthetic mixture form. Chromatographic separation achieved isocratically on Luna C18 column (5 µm, 150mm x 4.60mm) and acetonitrile: 0.05M ammonium acetate buffer (pH 4.5) in the ratio of 75:25 (v/v) as the mobile phase, at a flow rate of 0.6 mL/min. Detection was carried out at 245 nm. Parameters such as linearity, precision, accuracy, recovery, specificity and ruggedness are studied as reported in the ICH guidelines. The retention times for AI and PRA was found to be 2.25 ± 0.5 and 8.72 ± 0.5 min respectively. Linearity for AI and PRA was in the range of 75-375 μg/mL and 10-50 μg/mL respectively.  The mean recoveries obtained for AI and PRA were 99.58 and 99.48% respectively and RSD was less than 2. The correlation coefficients for all components are close to 1. Developed method was found to be accurate, precise, selective and rapid for simultaneous estimation of AI and PRA.

The objective of the present research work is to develop a isocratic reversed-phase liquid chromatographic (RP-HPLC) method for the determination of Aspirin in pharmaceutical pharmaceutical dosage forms for its related impurities in presence of esomeprazole. The chromatographic separation was achieved on a RP 18 column (100mm×4.6mm, 5 µm). The isocratic LC method employs mixture of buffer methanol and isopropyl alcohol in the ratio of (84:13:3 v/v) solutions as mobile phase. The buffer solution contains 6.8g of Potassium dihydrogen orthophosphate adjusted to pH 2.5 with orthophosphoric acid .The flow rate was 1.5 ml/min and the detection wavelength was 275 nm. In the developed HPLC method, the resolution between Aspirin and its potential impurity salicylic acid was found to be greater than 4.0. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation in presence of esomeprazole. Considerable degradation was found to occur in basic medium and mild degradation observed in acid hydrolysis stress conditions. Degradation product formed during acidic hydrolysis was salicylic acid. The stress samples were assayed against a qualified reference standard and the mass balance was found close to 99.5%. The developed RP-HPLC method was validated with respect to linearity, accuracy, precision and robustness.

The objective of the present research work is to develop a gradient, reversed-phase liquid chromatographic (RP-UPLC) method for the determination of Dutasteride in pharmaceutical bulk drugs for assay and its related impurities. The chromatographic separation was achieved on a Waters ACQUITY TM UPLC C8 Column (100mm×2.1mm, 1.7µm),. The isocratic LC method employs mixture of buffer and Acetonitrile in the ratio of (50:50 v/v) solutions as mobile phase. The buffer solution contains 1.0mM potassium di hydrogen orthophosphate pH adjusted to 5.0 with dil.Potassium hydroxide solution (Buffer) .The flow rate was 0.4 ml/min and the detection wavelength was 210 nm. In the developed UPLC method, the resolution between Dutasteride and its potential impurities, namely Imp-1, Imp-2 and Imp-3 was found to be greater than 4.0. The drug was subjected to stress conditions of hydrolysis, oxidation, photolysis and thermal degradation. Considerable degradation was found to occur in Acidic medium and mild degradation observed in base hydrolysis stress conditions. Degradation product formed during acidic hydrolysis was found to be Unknown impurity. The stress samples were assayed against a qualified reference standard and the mass balance was found close to 99.5%. The developed RP-UPLC method was validated with respect to linearity, accuracy, precision and robustness. The developed method was found to be linear in the range of 2.5-15µg/mL with correlation coefficient of 0.999 for assay procedures and found to be linear in the range of 0.05-3µg/mL with correlation coefficient of 0.999 for related impurities

The present manuscript describes simple, sensitive, rapid, accurate, precise and economical Q-absorbance ratio method for the simultaneous determination of diclofenac sodium and tolperisone hydrochloride in bulk and synthetic mixture. Absorbance ratio method uses the ratio of absorbances at two selected wavelengths, one which is an isoabsorptive point and other being the λ-max of one of the two components. Tolperisone hydrochloride and diclofenac sodium show an isoabsorptive point at 267 nm in methanol. The second wavelength used is 255 nm, which is the λ-max of tolperisone hydrochloride in methanol. The linearity was obtained in the concentration range of 2-20 μg/ml for both tolperisone hydrochloride and diclofenac sodium. The concentrations of the drugs were determined by using ratio of absorbances at isoabsorptive point and at the λ-max of tolperisone hydrochloride. The method was successfully applied to pharmaceutical dosage form because no interference from the synthetic mixture excipients was found. The suitability of this method for the quantitative determination of tolperisone hydrochloride and diclofenac sodium was proved by validation. The proposed method was found to be simple and sensitive for the routine quality control application of tolperisone hydrochloride and diclofenac sodium in synthetic mixture or pharmaceutical dosage form. The results of analysis have been validated statistically and by recovery studies.

A rapid and sensitive High Performance liquid chromatography (HPLC) method has been developed and validated. The analyte was extracted from human plasma by simple precipitation technique. Nifedipine was used as the internal standard. A Princeton C18 column provided chromatographic separation of the analyte. A simple, selective, rapid, precise and economical reverse phase High Pressure Liquid Chromatographic method has been developed for the estimation of Lercanidipine in human Plasma. The method was carried out on a Princeton C18 (250 mm x 4.6 mm i.d., 5µ) column with a mobile phase consisting of acetonitrile: Water (adjusted to pH 3.5 using orthophosphoric acid) (55:45 v/v) at a flow rate of 1.0 ml/min. Detection was carried out at 235 nm. The retention time of Lercanidipine Nifedipine, was 5.31, 10.00 min, respectively. The proposed method has been validated with linear range of 5.0-250.0 ng/ml for Lercanidipine. The precision and accuracy values are within 10%. The overall recovery of Lercanidipine was 96.4 %. The developed and validated method was applicable for the pharmacokinetics studies.