Quality risk assessment and DoE – Practiced validated stability-indicating chromatographic method for quantification of Rivaroxaban in bulk and tablet dosage form

A systematic DoE and Analytical Quality by Design (AQbD) approach was utilized for the development and validation of a novel stability indicating high-performance thin–layer chromatographic (HPTLC) method for Rivaroxaban (RBN) estimation in bulk and marketed formulation. A D-optimal design was used to screen the effect of solvents, volume of solvents, time from spotting to development and time for development to scanning. ANOVA results and Pareto chart revealed that toluene, methanol, water and saturation time had an impact on retention time. The critical method and material attributes were further screened by Box-Behnken design (BBD) to achieve optimal chromatographic condition. A stress degradation study was carried out and structure of major alkaline degradant was elaborated. According to the design space, a control strategy was used with toluene: methanol: water (6:2:2) and the saturation time was 15 min. A retention factor (RF) of 0.59 ± 0.05 was achieved for RBN using chromatographic plate precoated with silica gel at detection wavelength 282 nm with optimized conditions. The linear calibration curve was achieved in the concentration range of 200–1,200 ng/band with r 2 > 0.998 suggesting good coordination between analyte concentration and peak areas. The quadratic model was demonstrated as the best fit model and no interaction was noted between CMAs. The optimized HPTLC method was validated critically as stated in International Conference on Harmonization (ICH) Q2 (R1) guideline and implemented successfully for stress degradation study of RBN. The developed HPTLC method obtained through AQbD application was potentially able to resolve all degradants of RBN achieved through forced degradation study. The obtained results demonstrate that a scientific AQbD approach implementation in HPTLC method development and stress degradation study drastically minimizes the number of trials in experiments, ultimately time and cost of analysis could be minimized.

LC-MS/MS CHARACTERIZATION OF FORCED DEGRADATION PRODUCTS OF TUCATINIB, A NOVEL TYROSINE KINASE INHIBITOR: DEVELOPMENT AND VALIDATION OF RP-HPLC METHOD

Objective: The current study focused on the development, validation, and characterization of forced degradation products using LC-MS/MS. Methods: A simple, selective, validated and well-defined isocratic HPLC methodology for the quantitative determination of Tucatinib at a wavelength of 239 nm. An isocratic elution of samples was performed on an Inertsil ODS (250x4.6 mm, 5m) column with a mobile phase of 70:30v/v Acetonitrile and formic acid (0.1%) delivered at a flow rate of 1.0 ml/min. MS/MS was used to characterize degradation products formed in the forced degradation study. The validation and characterization of forced degradation products were performed in accordance with ICH guidelines. Results: Over the concentration range of 5-100μg/ml, a good linear response was obtained. Tucatinib's LOD and LOQ were determined to be 0.05 and 0.5, respectively. According to standard guidelines, the method was quantitatively evaluated in terms of system suitability, linearity, precision, accuracy, and robustness, and the results were found to be within acceptable limits. The drug was degraded under acidic, alkaline, and reduction conditions in forced degradation studies. Conclusion: The method was found to be applicable for routine tucatinib analysis. Because no LC-MS/MS method for estimating tucatinib and its degradation products has been reported in the literature. There is a need to develop a method for studying the entire tucatinib degradation pathway.

MODERN INSTRUMENTAL METHODS FOR QUALITATIVE AND QUANTITATIVE ANALYSIS OF LAPATINIB IN BIOLOGICAL FLUIDS AND DOSAGE FORMS (REVIEW)

Lapatinib is a small molecule, a heterocyclic quinazoline derivative. The drug is used for targeted therapy of patients with breast cancer, in which there is overexpression of the human epidermal growth factor receptors (HER/ErbB). This review is devoted to studying modern instrumental methods of qualitative and quantitative analysis of lapatinib, which can be used both for quality control and standardization (of bulk pharmaceuticals and dosage forms) and pharmacokinetics studies of a drug. Reverse-phase high-performance liquid chromatography (RP-HPLC) is mainly used to identify lapatinib in tablets. Depending on the purpose of the study, various detectors are used (ultraviolet or diode-matrix detector), which makes it possible to determine not only the native compound but also the products of its degradation. Definition of lapatinib in the presence of degraded products is necessary for forced degradation studies to determine drug stability. When a drug is being developed, it is important to define and understand its pharmacokinetics. For such studies, high-performance liquid chromatography (HPLC) coupled with the mass selective detector is often used. It allows determining lapatinib in biological fluids. However, these methods are not applicable for identifying the drug directly in dosage forms and require further development and validation.

Management of validation of HPLC method for determination of acetylsalicylic acid impurities in a new pharmaceutical product

The work mainly focused on a validation of the method for determining the content of salicylic acid and individual unknown impurities in new pharmaceutical product—tablets containing: 75, 100 or 150 mg of acetylsalicylic acid and glycine in the amount of 40 mg for each dosage. The separation of the components was carried out by means of HPLC, using a Waters Symmetry C18 column (4.6 × 250 mm, 5 μm) as the stationary phase. The mobile phase consisted of a mixture of 85% orthophosphoric acid, acetonitrile and purified water (2:400:600 V/V/V). Detection was carried out at a wavelength of 237 nm, with a constant flow rate of 1.0 ml min−1. In order to verify the method, linearity, precision (repeatability and reproducibility), accuracy, specificity, range, robustness, system precision, stability of the test and standard solution, limit of quantification and forced degradation were determined. Validation tests were performed in accordance with ICH (International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use) guidelines. The method was validated successfully. It was confirmed that the method in a tested range of 0.005–0.40% salicylic acid with respect to acetylsalicylic acid content is linear, precise and accurate.

Development and validation of a novel stability indicating HPLC method for the separation and determination of darolutamide and its impurities in pharmaceutical formulations

This study reports for the first time about a stability indicating RP-HPLC method for analysis of darolutamide and its impurities 1, 2, and 3 in bulk and formulations. The separation was achieved on Phenomenex column with Luna C18 (250 mm × 4.6 mm, 5 μm) as stationary phase, and 50 mM ammonium acetate: methanol solution 15:80 (v/v) at pH 5.2 as mobile phase at 1.0 mL/min flow rate. UV detection was carried at wavelength of 239 nm. In these conditions the retention time of darolutamide and its impurities 1, 2, and 3 was 7.05, 8.90, 4.63 and 5.95 min, respectively. The method was validated for system suitability, range of analysis, precision, specificity, stability, and robustness. Forced degradation study was done through exposure of the analyte to five different stress conditions and the % degradation was small in all degradation condition. The proposed method can separate and estimate the drug and its impurities in pharmaceutical formulations. Hence, the developed method was suitable for the quantification of darolutamide and can separate and analyse impurities 1, 2, and 3

Stability-Indicating UHPLC Method for the Determination of Desvenlafaxine: Application to Degradation Kinetics

This study was aimed to investigate the degradation behavior and physicochemical stability of desvenlafaxine using reversed-phase ultra-high-performance liquid chromatography (RP-UHPLC) system. The chromatogram was developed on Eclipse XDB-C8 column (150 x 4.6 mm, 5μm). The eluents were monitored through a photo diode array plus (PDA+) detector at 210 nm using an isocratic method with a flow rate of 1.5 ml/min. Mobile phase composition was 30:70 v/v mixture of 0.1 % trifluoroacetic acid (TFA) in water and methanol. Forced degradation studies were performed on drug substance of desvenlafaxine as per International Conference on Harmonization (ICH) prescribed stressed conditions (Q1A(R2) and Q1B) using hydrolytic (acidic, basic, and neutral), oxidative and photolytic methods. The drug substance was found highly labile to acidic (0.5 N hydrochloric acid, 18.65 % degradation in 2 hours at 70°C), basic (1.0 N sodium hydroxide, 11.01 % degradation in 12 hours at 70°C) and oxidative (3 % hydrogen peroxide, 17.05 % degradation in 2 hours at 50°C) stressed conditions, but a great resistance was observed towards dry heat (maximum degradation 0.27 % in 10 days from ambient to higher temperature, 80°C), moist heat (maximum degradation 0.25 % in 2 hours at 80°C and 75 % relative humidity) as well as in photolytic degradation (maximum degradation 0.23 % in 10 days at UV light of 315 - 400 nm). A pseudo-first order kinetic was followed in acidic, basic and peroxide degradation methods which paved a way to calculate the half-life of the drug substance desvenlafaxine under ICH mentioned stressed conditions. The results were also statistically analyzed and the % RSD values were compared with recommended guidelines. Dhaka Univ. J. Pharm. Sci. 20(2): 167-176, 2021 (December)