Quantification and Stability Indicating Method Development and Validation of Vismodegib in Bulk and Pharmaceutical Dosage Form by Ultra Performance Liquid Chromatography

Background: Vismodegib (VMD) is a drug of choice for the treatment of basal-cell carcinoma. Present studies carried out to estimate VMD by RP-UPLC technique and to develop a simple, précised, accurate method for routine analysis. Methods: For this purpose Chromatographic conditions used were stationary phase STD BEH C18 column (100 mm x 2.1 mm, 1.8m), a mixture of Methanol:KH2PO4 taken in the ratio 50:50%v/v as a mobile phase with a pH 7.4 and flow rate was maintained at 0.3ml/min, detection wave length was Acquity TUV 254nm, column temperature was set to 30oC and diluent was mobile phase, Conditions were finalized as optimized method. Results: System suitability parameters were studied by injecting the standard six times. Linearity study was carried out between 25% to150% (37.5-225µg/ml) levels, R2 value was found to be as 0.9992. Precision was found to be 0.6 for repeatability and 0.4 for intermediate precision. LOD and LOQ are 0.33 µg/ml and 0.99 µg/ml respectively and results were well under the acceptance criteria. Conclusion: By using above method assay of marketed formulation was carried out and was found 100.12%. Degradation studies of VMD were done, in all conditions purity threshold was more than purity angle and within the acceptable range. The developed method was simple and can be used for routine analysis.

DEVELOPMENT OF FORCED DEGRADATION STUDIES OF FAVIPIRAVIR BY RP-HPLC

Forced degradation studies and stability indicating method were developed for the estimation of Favipiravir by reverse phase High performance liquid chromatography in active Pharmaceutical ingredient and its tablet dosage form. The method was achieved by using C18 column (250 X 4.6mm X 4µm) with mobile phase mixture ortho phosphoric acid and acetonitrile in the ratio 60:40. The mobile phase was allowed to pump with the flow rate 1ml/min by maintaining detection wavelength at 324nm using ultra-violet detector. Favipiravir drug was subjected to various stress conditions according to International Conference of Harmonization Q1A(R2) guidelines to establish stability indicating method. Favipiravir drug was found to be sensitive at peroxide degradation. The impurity peak was characterized by mass spectral studies. The method was validated for analytical standards such as linearity, accuracy, Precision, sensitivity and robustness. A rapid and sensitive method was developed for the estimation of favipiravir which indicates its stability indicating behavior.

Stability-indicating method development and validation for the concurrent determination of darunavir, cobicistat, emtricitabine and tenofovir alafenamide by UPLC in bulk and tablet dosage forms

Background Tablet dosage forms containing combination of darunavir a protease inhibitor, cobicistat a cytochrome P450 3A inhibitor, emtricitabine and tenofovir alafenamide which were nucleoside reverse transcriptase inhibitors were approved by USFDA on 1st July 2018 to suppress the viral load in HIV patients. It can be used as a complete regimen for the treatment of HIV-1 infection in adults and paediatric patients weighing at least 40 kg. An UPLC method was developed, and separation was done on SB C8 column of dimensions 50 × 2.1 × 1.8 μ with mobile phase 0.01 N potassium dihydrogen ortho phosphate (pH-4.8) and acetonitrile in 60:40 ratio, at a flow rate of 0.3 mL/min and an injection volume of 2 μL. The column temperature was maintained at 30 °C, and detection wavelength was 267 nm. The method was validated according to ICH guidelines. Results The retention times were 1.031, 1.341, 1.630 and 2.153 min, and they were linear in the concentration range of 1.25–7.5 μg/mL, 18.75–112.5 μg/mL, 25–150 μg/mL and 100–600 μg/mL for tenofovir alafenamide, cobicistat, emtricitabine and darunavir, respectively. The intraday and interday precisions were found to be within acceptable limits. LOD was found to be 0.06 μg/mL, 0.51 μg/mL, 1.31 μg/mL and 3.01 μg/mL, and LOQ was 0.19 μg/mL, 1.54 μg/mL, 3.96 μg/mL and 9.13 μg/mL for tenofovir alafenamide, cobicistat, emtricitabine and darunavir. The correlation coefficients were found to be more than 0.999, and recovery was more than 99.52% indicating the method was accurate. Forced degradation studies reveal that the drugs are unstable under acidic conditions. The method was simple, accurate, precise, stable and can be analysed in less runtime of 4 min. Conclusions The flexibility, accuracy and precision of the developed method ensure its applicability in routine analysis of tablet dosage forms. Graphical Abstract

A Stability Indicating Method Development and Validation for The Simultaneous Estimation of Emtricitabine, Tenofovir Alafenamide and Doultegravir in Pharmaceutical Formulation by Ultra Performance Liquid Chromatography

A Precise, specific, linear, accurate and robust technique used to be developed for the simultaneous estimation of the, Emtricitabine, Tenofovir alafenamide and Doultegravir in pill dosage structure and Validated as per ICH Validation guidelines. Method was once optimised with the useful resource of Acquity BEH C18 125A° (100 × 2.1 mm, 1.7μ) column at a flow rate of 0.6ml/min, Mobile part was as soon as pH 3.8 Buffer, Methanol and Acetonitrile (50:40:10 respectively). The Column Oven temperature used to be maintained at 35°C and working wave dimension used to be selected at 274nm. The retention times of Emtricitabine, Tenofovir Alafenamide and Doultegravir have been determined to be 3.77, 6.34 and 7.67minutes respectively. The % RSD of the Emtricitabine, Tenofovir Alafenamide and Doultegravir had been and located to be 0.27%, 1.24% and 0.11% respectivelyIn Method precision Parameter, % Assay had been determined ninety five to 105.0% and p.c Recovery had been acquired as 98.9%, 100.4% and 100.0% for Emtricitabine, Tenofovir Alafenamide and Doultegravir respectively. Linearity was as soon as acquired as 0.999, 0.999 and 0.999 for Emtricitabine, Tenofovir Alafenamide and Doultegravir respectively. Analytical Range used to be determined from the linearity and accuracy for Emitricitabine used to be 50µg/mL to 150µg/mL, Tenofovir Alafenamide used to be as soon as 6.25µg/mL to 18.75µg/mL and Doultegravir was as soon as 12.5µg/mL to 37.5µg/mL.

STABILITY INDICATING RP-HPLC METHOD FOR SEPARATION AND QUANTIFICATION OF RELATED SUBSTANCES OF TICLOPIDINE IN BULK AND PHARMACEUTICAL FORMULATIONS

A simple, specific, accurate and stable reverse phase liquid chromatographic method was developed for the simultaneous determination of ticlopidine and its related impurities A and B in bulk drug and tablet dosage forms. The analysis has been performed on XTerra C18 column (250 mm×4.6 mm; 5 µ id) and mobile phase containing of methanol and pH 6.8 phosphate buffer in the ratio of 80:20 (V/V). The detection was carried at 228 nm with a flow rate of 1.0 mL/min. The retention times were found to be 8.9, 5.98 and 4.62 min for ticlopidine, impurities A and B, respectively. The method was validated according to ICH guidelines. The method was validated for specificity, precision, linearity, accuracy and robustness. The linearity range of 50-200 µg/mL for ticlopidine and 0.5-2.0 µg/mL for impurity A and B. The recoveries of ticlopidine and impurities were found to be within the range of 98-102 and the % RSD in each spiked level was found to be less than 2. The stress degradation studies confirmed that the method was effectively separate the degradation products and impurities formed in the stress studies and hence the method was found to be stability indicating method. The method can effectively quantify the standard drug ticlopidine and its impurities A and B in bulk drug and pharmaceutical formulations.

A novel stability-indicating method for known and unknown impurities profiling for diltiazem hydrochloride pharmaceutical dosage form (tablets)

Background A novel gradient, high-sensitive and specific stability-indicating reverse-phase HPLC method was developed and validated for quantitative purpose of known, unknown and degradant impurities profiling for diltiazem hydrochloride tablets. The impurities were separated on the Zorbax RX C8 column (150 mm × 4.6 mm, 5 μm) with mobile phase-A consisting of a mixture of 0.05 M sodium dihydrogen phosphate monohydrate buffer pH 3.0 and methanol in the ratio 800:200v/v and mobile phase-B consisting of acetonitrile with a flow rate of 1.0 mL min−1. The column compartment was maintained at 35 °C, and the detection wavelength was 240 nm. Diltiazem hydrochloride, its known impurities and unknown impurities have been well resolved from each other. Results The linearity of the method has been demonstrated across the concentration range of 0.18 to 5.65 µg mL−1 for EP impurity-F with correlation coefficient R2 greater than 0.99. Recovery of method was proved from LOQ to 150% for known and unknown impurities with respect to test concentration and found in between 80 and 120%. Forced degradation study and specificity experiment results with mass balance proved the stability-indicating nature of the method and separated all known, unknown impurities and degradants from each other as well as from main drug component (diltiazem hydrochloride). The mass balance for stress study was found in between 95 and 105%. Conclusion Newly developed analytical method was validated as per ICH Q2 (R1) guidelines “Validation of analytical procedure” and found linear, accurate, specific, robust and precise in the established working range.