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This article describes the first simple, fast, time-saving, and cost-effective UHPLC method that was developed and validated for simultaneous quantification of abiraterone acetate, its four degradation products, and six specified process impurities in bulk and tablet form. Moreover, when coupled with a mass spectrometer detector, the proposed method provides additional advantages for confirmation of peak and correct identification based on molecular weight. The eleven peaks were separated on a Water Acquity BEH C18, (150 mm length, 2.1 mm internal diameter, 1.7 µm particle size) column maintained at a 50.0 °C temperature. Using 0.05% formic acid in 10 mM ammonium formate, acetonitrile, and methanol as mobile phases in gradient elution at a flow rate of 0.40 mL/min. provides excellent separation at 260 nm. The linearity curves of all analytes showed promising results with a correlation coefficient of 0.999 with a lower limit of detection and quantification. A forced degradation study on solid abiraterone acetate proved its specificity with improvements and significance. This proposed method provides improved separation with a lower flow rate, which offers faster analysis, reduces wastage and cost, and specifies the greener advantages compared to reported methods. The outcome of the specificity, linearity, precision, and trueness as per ICH guidelines proved that the proposed method is fast, time-saving, and cost-effective for the intended purpose.

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Analytical methods for the drug substance and degradation products (DPs) are validated by performing forced degradation studies. Forced degradation studies of Velpatasvir (VEL) drug substance and Velpatasvir copovidone solid dispersion (VEL-CSD) were performed under the stressed alkaline, acidic, oxidative and thermal conditions according to ICH guidelines ICH Q1A (R2). VEL is labile to degrade in stressed alkaline, acidic, and oxidative conditions. It is also photolabile and degraded during photostability studies as described by ICH Q1B, and showed no degradation on exposure to extreme temperature when protected from light. A sensitive stability indicating HPLC-UV method was developed and validated for the separation of VEL and eight DPs. The DPs of VEL are separated using gradient elution of mobile phase containing 0.05% Trifluoroacetic acid (TFA) and methanol over symmetry analytical column C18 (250 mm × 4.6 mm, 5 µm) with a flow rate of 0.8 mL min-1. Simultaneous detection of all DPs and VEL was performed on UV detector at 305 nm. The performance parameters like precision, specificity and linearity of the method were validated using reference standards as prescribed by ICHQ2 (R1). Limits of quantification and limits of detection were determined from calibration curve using the expression 10δ/slope and 3δ/slope respectively. The proposed method is stability-indicating and effectively applied to the analysis of process impurities and DPs in VEL drug substance and VEL-CSD.

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The enantiomeric purity of escitalopram oxalate ESC and its "in-process impurities," namely, ESC-N-oxide, ESC-citadiol, and R(-)-enantiomer were studied in drug substance and products using high-performance liquid chromatography (HPLC)-UV (Method I), synchronous fluorescence spectroscopy (SFS) (Method IIA), and first derivative SFS (Method IIB). Method I describes as an isocratic HPLC-UV for the direct resolution and determination of enantiomeric purity of ESC and its "in-process impurities." The proposed method involved the use of αl -acid glycoprotein (AGP) chiral stationary phase. The regression plots revealed good linear relationships of concentration range of 0.25 to 100 and 0.25 to 10 µg mL-1 for ESC and its impurities. The limits of detection and quantifications for ESC were 0.075 and 0.235 µg mL-1 , respectively. Method II involves the significant enhancement of the fluorescence intensities of ESC and its impurities through inclusion complexes formation with hydroxyl propyl-ß-cyclodextrin as a chiral selector in Micliavain buffer. Method IIA describes SFS technique for assay of ESC at 225 nm in presence of its impurities: R(-)-enantiomer, citadiol, and N-oxide at ∆λ of 100 nm. This method was extended to (Method IIB) to apply first derivative SFS for the simultaneous determination of ESC at 236 nm and its impurities: the R(-)-enantiomer, citadiol, and N-oxide at 308, 275, and 280 nm, respectively. Linearity ranges were found to be 0.01 to 1.0 µg mL-1 for ESC and its impurities with lower detection and quantification limits of 0.033/0.011 and 0.038/0.013 µg mL-1 for SFS and first derivative synchronous fluorescence spectra (FDSFS), respectively. The methods were used to investigate the enantiomeric purity of escitalopram.

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During the formation of a tetrazole ring on an investigational drug, two in-process impurities were detected and analyzed by LC-MS, which suggested that both impurities were drug-related with the same mass-to-charge ratio. To understand and control their formation, both impurities were isolated from the mother liquor of the reaction using a multi-step isolation procedure to obtain a sufficient amount for high-resolution mass spectrometry (HRMS) and NMR structural analysis. HRMS suggested a protonated mass of 577.32 Da for both impurities; however, MS fragmentation patterns provided limited information on their structures. NMR analysis indicated the presence on an additional NH functional group in both isolates with similar spatial and bond correlations to one of the dimethylcarbamoyl moieties and the corresponding aromatic ring. A phenyldimethylcarbamoylamino moiety was supported by the NMR and HRMS data and could be explained based on the 'Schmidt-like' reaction mechanism, which was an unexpected reaction pathway. Because the reaction conditions were fixed because of safety concerns, the crystallization protocol was redesigned to reduce the levels of these impurities significantly. Copyright © 2016 John Wiley & Sons, Ltd.

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A novel ultra high performance liquid chromatography method development strategy was ameliorated by applying quality by design approach. The developed systematic approach was divided into five steps (i) Analytical Target Profile, (ii) Critical Quality Attributes, (iii) Risk Assessments of Critical parameters using design of experiments (screening and optimization phases), (iv) Generation of design space, and (v) Process Capability Analysis (Cp) for robustness study using Monte Carlo simulation. The complete quality-by-design-based method development was made automated and expedited by employing sub-2 µm particles column with an ultra high performance liquid chromatography system. Successful chromatographic separation of the Coenzyme Q10 from its biotechnological process related impurities was achieved on a Waters Acquity phenyl hexyl (100 mm × 2.1 mm, 1.7 µm) column with gradient elution of 10 mM ammonium acetate buffer (pH 4.0) and a mixture of acetonitrile/2-propanol (1:1) as the mobile phase. Through this study, fast and organized method development workflow was developed and robustness of the method was also demonstrated. The method was validated for specificity, linearity, accuracy, precision, and robustness in compliance to the International Conference on Harmonization, Q2 (R1) guidelines. The impurities were identified by atmospheric pressure chemical ionization-mass spectrometry technique. Further, the in silico toxicity of impurities was analyzed using TOPKAT and DEREK software.

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The present study reports the degradation behaviour of a new prokinetic agent, Prucalopride succinate, under various stress conditions as per International Conference on Harmonization guidelines (ICH, Q1A (R2)). The investigation involved monitoring decomposition of the drug under hydrolytic (acidic, basic and neutral), oxidative, photolytic and thermal stress conditions followed by characterization of the degradation products (DPs) and process related impurities (IMPs). A rapid, precise, accurate and robust reverse phase high performance liquid chromatography (RP-HPLC) method has been developed involving mobile phase of 20mM ammonium bicarbonate buffer and acetonitrile: methanol (80:20v/v) on a Waters Xbridge-C8 (150mm×4.6mm i.d., 3.5µm) column using gradient elution. The drug was found to be degraded in hydrolytic (acidic) and oxidative conditions, whereas it was stable under basic and neutral hydrolytic, photolytic and thermal stress conditions. The method was extended to LC-ESI-QTOF-MS/MS for the structural characterization of DPs and process related IMPs. Structural characterization was carried out based on the generated molecular formula of DPs and its fragment ions. It has been observed that two major DPs were formed under each acid hydrolysis and oxidative stress conditions. The most probable mechanisms involved in the formation of DPs were also proposed. Finally, the method was validated in the term of specificity, linearity, accuracy, precision, and robustness as per ICH guidelines, Q2 (R1).

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In this study, a stability-indicating reversed-phase liquid chromatographic electrospray mass spectrometric method was developed and validated for the determination of process-related impurities and forced degradants of Efavirenz in bulk drugs. Efavirenz was subjected to acid, alkaline hydrolysis, H2O2 oxidation, photolysis, and thermal stress. Significant degradation was observed during alkaline hydrolysis, and the degradants were isolated on a mass-based purification system and characterized by high-resolution mass spectrometry, positive electrospray ionization tandem mass spectrometry, and (1)H and (13)C NMR spectroscopy. Accurate mass measurement and NMR spectroscopy revealed the possible structure of process-related impurities and degradant under stress conditions. The acceptable separation was accomplished on Waters bondapak C18 column (250 mm × 4.6 mm; 5 µm), using 5 mM ammonium acetate and acetonitrile as a mobile phase in a gradient elution mode at a flow rate of 1.0 mL/min. The eluents were monitored by diode array detector at 247 nm and quantitation limits were obtained in the range of 0.1-2.5 µg/mL for Efavirenz, degradants, and process-related impurities. The liquid chromatography method was validated with respect to accuracy, precision, linearity, robustness, and limits of detection and quantification as per International Conference on Harmonization guidelines.

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Following the underlying principles of quality by design mentioned in the ICH Q8 guidance, systematic approaches for the control of process-related impurities have been taken in the manufacturing process of fasudil hydrochloride, a potent Rho-kinase inhibitor and vasodilator. Three related impurities were found in fasudil hydrochloride lab samples by a newly developed RP-HPLC with volatile mobile phase gradient elution and UV detection method. The elemental compositions of the impurities were determined by positive ESI high-resolution TOF-MS analysis of their [M + H](+) ions and their structures were identified through the elucidation of the product mass spectra obtained by a triple quadrupole mass spectrometer. The key impurity was further verified through synthesis and organic spectroscopy including NMR and IR spectroscopy. The origins of these impurities were located and the effective approaches to eliminate them were proposed based on the redesign of the synthetic conditions. The results obtained are important for quality control in the manufacture of fasudil hydrochloride bulk drug substance and injection.

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