Stability-indicating RP-HPLC method for simultaneous quantitation of tramadol and aceclofenac in presence of their major degradation products: Method development and validation

Primary objective of this study was to develop a stability-indicating reverse-phase high-performance liquid chromatography (HPLC) method for simultaneous quantitation of tramadol and aceclofenac in presence of their degradation products. The drugs were subjected to various International Conference on Harmonization recommended stress conditions, such as acid hydrolysis, alkaline hydrolysis, peroxide oxidation, thermolysis, and photolysis. The major degradation products got well resoluted from the analytes in HPLC analysis with a mobile phase composed of a mixture of 0.01?M ammonium acetate buffer (pH 6.5) and acetonitrile (65:35, v/v) through a Phenomenex Gemini C18 (250?mm?×?4.6?mm, 5?µm particle size) column. The method was linear over a range of 15–60?µg/mL for tramadol and 40–160?µg/mL for aceclofenac concentration. The analytes were detected at a wavelength of 270?nm. The method was validated and found to be specific, accurate, precise, stable, and robust for its intended use. The method can be recommended for its future use in routine quality control, accelerated and real-time stability analysis of the formulations containing tramadol and aceclofenac combination.     

Applying green analytical chemistry for development and validation of RP-HPLC stability indicating assay method for estimation of fenoverine in bulk and dosage form using quality by design approach

A green and robust reverse-phase liquid chromatographic method has been developed for the determination of fenoverine (FEN), by applying combined principles of green analytical chemistry and quality by design approaches on a Spherisorb C18 column (150?×?4.6?mm, 3?µm) with UV detection at 262?nm. A two level fractional factorial design (2^^7-3) Res IV was used for screening of influential chromatographic factors. The critical method parameters actively affecting critical quality attributes (CQAs) were identified and further optimized using Box–Behnken design. The predicted optimum assay conditions comprised of methanol and ammonium acetate buffer 20?mM, in an extent of 81:19% v/v individually having a flow rate of 1.0?mL/min with a column oven temperature of 33°C. The drug was stressed in hydrolytic, oxidative, reductive, thermal, and photolytic conditions. The developed method was validated successfully. The detector response was linear in the concentration of 0.5–160?µg/mL with a limit of detection (LOD) and limit of quantitation (LOQ) as 0.1 and 0.3?µg/mL, respectively. The % recovery was found to be 99.7%. The analytical method volume intensity value for developed method was 45?mL and the environment assessment tool (EAT) score was 41.07. The method is simple, environmentally benign, rapid, and robust for the determination of FEN in bulk and in its dosage form.     

Stability-indicating densitometric determination of some antidiabetic drugs in dosage forms, using TLC

The most commonly used antidiabetic sulfonylurea drugs, gliclazide, glipizide and glibenclamide, were determined using stability-indicating densitometric methods. The degradation products were prepared by acid hydrolysis of the intact drugs. Thin layer chromatography was carried out using silica gel 60 F₂₅₄ plates and different mobile phases, followed by scanning of the developed chromatograms. Mixtures of the investigated drugs and their degradation products were prepared and analysed using the proposed methods, with recovery in the range 100.4–101.0% and RSD in the range 0.6–0.7%. Pharmaceutical dosage forms were assayed and found to give results of the same accuracy and reproducibility as official or reference methods.     

Development,stability-indicating assessment,and evaluation of influential method conditions using a full factorial design for the determination of Nintedanib esylate-related impurities

The design of an appropriate analytical method for assessing the quality of pharmaceuticals requires a deep understanding of science, and risk evaluation approaches are appreciated. The current study discusses how a related substance method was developed for Nintedanib esylate. The best possible separation between the critical peak pairs was achieved using an X-Select charged surface hybrid Phenyl Hexyl (150 × 4.6) mm, 3.5 μm column. A mixture of water, acetonitrile, and methanol in mobile phase-A (70:20:10) and mobile phase-B (20:70:10), with 0.1% trifluoroacetic acid and 0.05% formic acid in both eluents. The set flow rate, wavelength, and injection volumes were 1.0 ml/min, 285 nm, and 5 μl, respectively, with gradient elution. The method conditions were validated as per regulatory requirements and United States Pharmacopeia general chapter < 1225 >. The correlation coefficient for all impurities from the linearity experiment was found to be > 0.999. The % relative standard deviation from the precision experiments ranged from 0.4 to 3.6. The mean %recovery from the accuracy study ranged from 92.5 to 106.5. Demonstrated the power of the stability-indicating method through degradation studies; the active drug component is more vulnerable to oxidation than other conditions. Final method conditions were further evaluated using a full-factorial design. The robust method conditions were identified using the graphical optimization from the design space.     

Development and Validation of a Stability-Indicating UPLC Method for the Determination of Hexoprenaline in Injectable Dosage Form Using AQbD Principles

A novel and efficient stability-indicating, reverse phase ultra-performance liquid chromatographic (UPLC^®) analytical method was developed and validated for the determination of hexoprenaline in an injectable dosage form. The development of the method was performed using analytical quality by design (AQbD) principles, which are aligned with the future requirements from the regulatory agencies using AQbD principles. The method was developed by assessing the impact of ion pairing, the chromatographic column, pH and gradient elution. The development was achieved with a Waters Acquity HSS T3 (50 × 2.1 mm i.d., 1.8 µm) column at ambient temperature, using sodium dihydrogen phosphate 5 mM + octane-1-sulphonic acid sodium salt 10 mM buffer pH 3.0 (Solution A) and acetonitrile (Solution B) as mobile phases in gradient elution (t = 0 min, 5% B; t = 1 min, 5% B; t = 5 min, 50% B; t = 7 min, 5% B; t = 10 min, 5% B) at a flow rate of 0.5 mL/min and UV detection of 280 nm. The linearity was proven for hexoprenaline over a concentration range of 3.50–6.50 µg/mL (R² = 0.9998). Forced degradation studies were performed by subjecting the samples to hydrolytic (acid and base), oxidative, and thermal stress conditions. Standard solution stability was also performed. The proposed validated method was successfully used for the quantitative analysis of bulk, stability and injectable dosage form samples of the desired drug product. Using the AQbD principles, it is possible to generate methodologies with enhanced knowledge, which can eventually lead to a reduced regulatory risk, high quality data and lower operational costs.     

Stability-indicating HPLC method for acyclovir and lidocaine in topical formulations

A simple, rapid and accurate stability-indicating HPLC assay was developed for the determination of acyclovir and lidocaine in topical formulations. Chromatographic separation of acyclovir and lidocaine was achieved using a reversed-phase C₁₈ column and a gradient mobile phase (20 mm ammonium acetate pH 3.5 in water and acetonitrile). The degradation products of acyclovir and lidocaine in the samples were analyzed by ultra performance liquid chromatography-time of flight mass spectrometry. The HPLC method successfully resolved the analytes from the impurities and degradation products in the topical formulation. Furthermore, the method detected the analytes from the human skin leachables following the extraction of the analytes in the skin homogenate samples. The method showed linearity over wide ranges of 5–500 and 10–200 μg/ml for acyclovir and lidocaine in the topical product, respectively, with a correlation coefficient (r²) >0.9995. The relative standard deviations for precision, repeatability, and robustness of the method validation assays were <2%. The skin extraction efficiency for acyclovir and lidocaine was 92.8 ± 0.7% and 91.3 ± 3.2%, respectively, with no interference from the skin leachables. Thus, simultaneous quantification of acyclovir and lidocaine in the topical formulations was achieved.     

White analytical chemistry-driven stability-indicating concomitant chromatographic estimation of thiocolchicoside and aceclofenac using response surface analysis and red,green, and blue model

White analytical chemistry is a novel concept for the assessment of analytical methods on basis of its validation efficiency, greenness power, and economical efficiency. White analytical chemistry-driven stability indicating chromatographic method has been developed for the concomitant analysis of thiocolchicoside and aceclofenac. The proposed chromatographic method has been developed using a safe and environmental-friendly organic solvents for the concomitant stability study of thiocolchicoside and aceclofenac. The analytical risk assessment was carried out for the identification of high-risk analytical risk factors and analytical method performance attributes. The mixture design was applied for the design of experiments-based response surface modeling of high-risk analytical risk factors and analytical method performance attributes. The degradation products were isolated and characterized using infrared, nuclear magnetic resonance, and mass spectral data. The proposed method was compared for its validation efficiency, greenness power, and cost-efficiency with published chromatographic methods using the red, green, and blue models. The white score of the proposed and reported method was calculated by averaging the red, green, and blue scores of the methods. The proposed method was found to be robust, green, and economical for the concomitant stability study of thiocolchicoside and aceclofenac.     

Quality-by-design-based development and validation of a stability-indicating UPLC method for quantification of teriflunomide in the presence of degradation products and its application to in-vitro dissolution

A systematic design-of-experiments was performed by applying quality-by-design concepts to determine design space for rapid quantification of teriflunomide by the ultraperformance liquid chromatography (UPLC) method in the presence of degradation products. Response surface and central composite quadratic were used for statistical evaluation of experimental data using a Design-Expert software. The response variables such as resolution, retention time, and peak tailing were analyzed statistically for the screening of suitable chromatographic conditions. During this process, various plots such as perturbation, contour, 3D, and design space were studied. The method was developed through UPLC BEH C18 2.1?×?100?mm, 1.7-µ column, mobile phase comprised of buffer (5?mM K₂HPO₄ containing 0.1% triethylamine, pH 6.8), and acetonitrile (40:60 v/v), the flow rate of 0.5?mL?min^?1 and UV detection at 250?nm. The method was developed with a short run time of 1?min. Forced degradation studies revealed that the method was stability-indicating, suitable for both assay and in-vitro dissolution of a drug product. The method was found to be linear in the range of 28–84?µg?mL^?1, 2.8–22.7?µg?mL^?1 with a correlation coefficient of 0.9999 and 1.000 for assay and dissolution, respectively. The recovery values were found in the range of 100.1–101.7%. The method was validated according to ICH guidelines.     

Development and Validation of Stability-Indicating HPLC Methods for Quantitative Determination of Pravastatin,Fluvastatin, Atorvastatin,and Rosuvastatin in Pharmaceuticals

Abstract

High-performance liquid-chromatographic (HPLC) methods were validated for determination of pravastatin sodium (PS), fluvastatin sodium (FVS), atorvastatin calcium (ATC), and rosuvastatin calcium (RC) in pharmaceuticals. Two stability-indicating HPLC methods were developed with a small change (10%) in the composition of the organic modifier in the mobile phase. The HPLC method for each statin was validated using isocratic elution. An RP-18 column was used with mobile phases consisting of methanol–water (60:40, v/v, for PS and RC and 70:30, v/v, for FVS and ATC). The pH of each mobile phase was adjusted to 3.0 with orthophosphoric acid, and the flow rate was 1.0 mL/min. Calibration plots showed correlation coefficients (r) > 0.999, which were calculated by the least square method. The detection limit (DL) and quantitation limit (QL) were 1.22 and 3.08 µg/mL for PS, 2.02 and 6.12 µg/mL for FVS, 0.44 and 1.34 µg/mL for ATC, and 1.55 and 4.70 µg/mL for RC. Intraday and interday relative standard deviations (RSDs) were <2.0%. The methods were applied successfully for quantitative determination of statins in pharmaceuticals.     

Validation of HPLC stability-indicating method for Vitamin C in semisolid pharmaceutical/cosmetic preparations with glutathione and sodium metabisulfite, as antioxidants

HPLC stability-indicating method was validated for Vitamin C (ascorbic acid) in semisolid pharmaceutical/cosmetic formulations containing glutathione and sodium metabisulfite, as antioxidants. The described procedure included a reliable, precise, accurate and specific method determination employing a 250 mm × 4.6 mm C₁₈ column, 0.2% metaphosphoric acid/methanol/acetonitrile (90:8:2, v/v/v) as the mobile phase and detection at 254 nm. Nicotinic and ascorbic acids were employed as standards, both presenting purity of 99.0%. Linearity was established for the ascorbic acid concentrations ranging form 1.0 to 12 μg mL⁻¹, accuracy/recovery percentage was 95.46-101.54%, precision values were 0.38 (intra-day) and 1.22% (inter-days), and LOD and LOQ were found to be 0.05 and 0.17 μg mL⁻¹, respectively. The working mobile phase elevated the ascorbic acid retention time to ≈3.5 min at a flow rate of 1.0 mL min⁻¹ and provided resolution of the active from the nicotinic acid (internal standard), degradation product (oxalic acid) and other excipients from the pharmaceutical/cosmetic preparations.