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Potential genotoxic impurities in medications are an increasing concern in the pharmaceutical industry and regulatory bodies because of the risk of human carcinogenesis. To prevent the emergence of these impurities, it is crucial to carefully examine not only the final product but also the intermediates and key starting material (KSM) used in drug synthesis. During the related substances analysis of KSM of Famotidine, an unknown impurity in the range of 0.5–1.0% was found prompting the need for isolation and characterization due to the possibility of its to infiltrate into the final product. In this study, the impurity was isolated and characterized as 5-(2-chloroethyl)-3,3-dimethyl-3,4-dihydro-2H-1,2,4,6-thiatriazine 1,1-dioxide using multiple instrumental analysis, uncovering a structural alert that raises concern. Considering the potential impact of impurity on human health, an in silico genotoxicity assessment was established using Derek and Sarah tool in accordance with ICH M7 guideline. Furthermore, molecular docking and molecular dynamics simulation were performed to evaluate the specific interaction of the impurity with DNA. The findings reveal consistent interaction of the impurity with the dG-rich region of the DNA duplex and binding at the minor groove. Both in silico prediction and molecular dynamic study confirmed the genotoxic character of the impurity. The newly discovered impurity in famotidine has not been reported previously, and there is currently no analytical method available for its identification and control. A highly sensitive HPLC-UV method was developed and validated in accordance with ICH requirements, enabling quantification of the impurity at trace level in famotidine ensuring its safe release. [ABSTRACT FROM AUTHOR]

The objective of the present research work was to demonstrate the application of liquid chromatography coupled with tandem mass spectrometry (LC‐MS/MS) for the separation, identification, and characterization of degradation products (DPs) of the drug lobeglitazone. The drug was subjected to various stress conditions such as oxidation, hydrolysis, thermal, and photolytic degradation as per the guidelines of the International Conference on Harmonization Q1A (R2). The drug was stable under thermal stress conditions, whereas it was susceptible to degradation in the other stress conditions forming four DPs. DPs were separated using a high‐performance LC system equipped with Zorbax SB C18 column (250 × 4.6 mm, 5 µm particles) using isocratic elution mode. The DPs were identified and characterized using MS and MS/MS. The chemical structure and fragmentation pattern were predicted for each degradant from the data obtained by mass studies. The drug and identified DPs were assessed by in‐silico approaches for predicting absorption, distribution, metabolism, and toxicity behavior. The in‐silico tools used for prediction were the pkCSM web server, ToxTree, and OSIRIS property explorer. [ABSTRACT FROM AUTHOR]

Indacaterol, glycopyrronium, and mometasone furoate triple combination inhalable fixed-dose medicines are effectively used to treat asthma and various chronic pulmonary disorders. The study aimed to develop and validate a simple single-run RP-HPLC impurity quantitation method. The chromatographic separation was accomplished using gradient elution mode of mobile phase A (potassium dihydrogen phosphate buffer pH 2.2) and mobile phase B (mixture of acetonitrile and methanol), with a flow rate of 0.8 mL/min using YMC Triart, C18 (250 × 4.6 mm, 5 µm) HPLC column at 45 °C and the detection wavelength of 210 nm (for indacaterol, glycopyrronium and their impurities) and 248 nm (for mometasone furoate and its impurities). Water and methanol (20:80) were used as a diluent. Quantitation of 10 known and several unknown impurities was successfully performed with the determination of relative response factors for all the known impurities. The developed method was validated as per the ICH Q2(R1) guidelines. The stability indicating the nature of the method was proved by performing stress study on the sample and placebo. The linearity and range of the method were proved by calculating the r2 values (> 0.998). The overall precision was found to be within 1.82 − 7.76% RSD. The recovery for all the actives and known impurities were within 90 − 115% with 0.4 − 12% RSD. The sample solution was stable for 2 days at room temperature. The developed method can be successfully used for the impurity analysis of routine, stability, and commercial samples in a quality control laboratory of the pharmaceutical industry. [ABSTRACT FROM AUTHOR]

The topical lotion of clobetasol propionate was approved for the treatment of psoriasis. Commercially, it is available in the various dosage forms like ointment, cream, foam gel, solution, in various quantities ranges from 0.05 %-0.5 % w/w. The drug is reported in all official pharmacopoeias with their 13 related substances. For the purpose of estimating clobetasol propionate in bulk and formulations using the reversed-phase high performance liquid chromatography technology, an accurate and reliable method was devised. Agilent 1260 Infinity II type high performance liquid chromatography with diode array detector and Phenomenex Luna-C18 column, measuring 250×4.6 mm, 5 µm was utilized in the method. Ammonium acetate buffer, acetonitrile and methanol (60:20:20) made up the mobile phase A combination; acetonitrile and ammonium acetate buffer (20:80) made up the mobile phase B combination. The optimum peak was obtained at retention duration of 15.73 min by maintaining a flow rate of 1.0 ml/min at a wavelength of 240 nm throughout. The percentage error for the instrument, method and intermediate precision was 0.01 %, 0.01 % and 0.02 % respectively. For both method and intermediate precision, the total percentage relative standard deviation was 0.02 %. The procedure was linear and accurate with correlation coefficient of 0.9997 for concentration ranges of 0.05-120 µg/ml with accuracy levels of 0.08 %, 0.02 % and 0.03 % for relative standard deviation of 80 %, 100 % and 120 %. When the drug's stress stability was examined, it was discovered to be unstable under basic conditions, degrading at 99.34 % and under heat conditions, degrading at 14.40 %. Since the established approach is relatively linear and the limits of detection and quantification for clobetasol propionate are very low at 0.93 µg/ml and 2.81 µg/ml respectively, it may be employed in a commercial setting. Retentions were verified and impurities were found for injecting the reference standard. Impurities A, B, J, L and M were found and their retention times were 7.91, 9.28, 18.75, 21.35 and 27.07 min. It was discovered that the approach was accurate and robust. [ABSTRACT FROM AUTHOR]

Ganirelix, a peptide‐based drug used to treat female infertility, has been in high market demand, which attracted generic formulation. A hitherto unknown impurity of ganirelix was observed in our formulation process, which reached ~0.3% in 6 months and led to a detailed investigation of its structure. In‐depth analysis of ESI‐MS/MS data of this impurity coupled with an artificial intelligence prediction tool led to a highly unusual putative structure, that is, N‐(2‐carboxyethyl)‐ganirelix (NCE‐GA), which was authenticated by chemical synthesis from ganirelix and NMR analysis and via corroborated HPLC and MS/MS data with the formulation‐derived impurity. [ABSTRACT FROM AUTHOR]

The present research aimed to comprehensively investigate the forced degradation of tepotinib through a combined in‐silico and experimental analysis. Tepotinib is a recently approved drug for metastatic non‐small cell lung cancer. A short and precise ultra‐high‐performance liquid chromatography (UHPLC) method utilizing a CSH C18 column (100 × 2.1 mm, 1.7 μm) was developed, using a mobile phase A as 0.1% formic acid and mobile phase B as HPLC‐grade acetonitrile. The flow rate was set at 0.2 mL/min, and detection was carried out at 260 nm. Forced degradation was carried out under acidic, basic, oxidative, and photolytic environments and also virtually with Zeneth software. Structural characterization of degradation products (DPs) was carried out using hyphenated techniques such as LC‐quadrupole time‐of‐flight‐tandem mass spectrometry in positive electrospray ionization mode. Further, toxicity was assessed with Derek and Sarah software. Analysis revealed the formation of five distinct forced DPs: DP‐1 (m/z 511), DP‐2 (m/z 527), DP‐3 (m/z 525), DP‐4 (m/z 512), and DP‐5 (m/z 509) under experimental conditions and eight in Zeneth prediction. Furthermore, nephrotoxicity alerts were observed for DP‐3 and DP‐5, whereas DP‐3 also showed mutagenicity alerts. This integrated blend of in‐silico and real experimental analyses was found to be effective in quality control for routing monitoring of degradation products of tepotinib. [ABSTRACT FROM AUTHOR]

Background: The goal of the proposed study was to develop and validate stability indicating mass compatible reverse phase UPLC method for impurity profiling of Darunavir Ethanolate and Ritonavir degradation impurities in fixed-dose drug combination products. Materials and Methods: The optimized chromatographic condition includes use of Zorbax Bonus C18 column (150x2.1 mm, 1.8 µm) with mobile phase A (Buffer (55): Methanol (45)) and mobile phase B (Acetonitrile: (30) and Methanol (70)), flow rate of 0.22 mL/min and detection at 240 nm with gradient program of 50 min. The method was validated as per ICH quality guideline Q2 (R1) including specificity by forced degradation study to confirm suitability for intended use. Results: The observed retention time for Darunavir was 11.4 min and for Ritonavir was 30.0 min. The Limit of Quantitation (LOQ) for all degradation impurities found is 0.05%, equivalent to the reporting threshold level. The method was found linear and accurate in the range of LOQ to 150% with an observed range of % accuracy of 90.1 to 106.3 for all known impurities. The method was found precise based on less than 10.0% RSD and robust for deliberate changes. Considering the use of a volatile mobile phase, the method can be applied for LC-MS-based analysis for mass identification. Conclusion: The developed UPLC method was applied for impurity profiling during the release and stability study of Darunavir Ethanolate and Ritonavir in fixed-dose drug combination products. [ABSTRACT FROM AUTHOR]

Propranolol hydrochloride (PPH) is a medication of beta‐adrenergic receptors. It is used to treat pediatric hemangiomas that are growing and need systemic therapy. A reversed‐phase high‐performance liquid chromatography (HPLC) was made to separate and estimate the amounts of ten known organic impurities of propranolol in bulk drugs, tablets, and capsule dosage forms. The chromatographic separation was achieved on a C18 column (100 × 4.6 mm, 2.7 μm) using a binary gradient mixture of pH 2.3 phosphate buffer and organic modifiers of acetonitrile, methanol, and isopropyl alcohol as the mobile phase. The stability‐indicating character of the proposed method was proven using stress testing study. The test method was validated for specificity, limit of detection, limit of quantitation, linearity, precision, accuracy, and robustness. For the propranolol and its ten organic impurities, the limit of quantitation, linearity, and recoveries were found in a range of 0.0123–0.4266 μg/mL (R2 > 0.9982) and 89.2–98.9%, respectively. The proposed liquid chromatography method is found to be highly suitable for impurity profiling of propranolol in bulk drugs and pharmaceutical formulations (tablets and capsules). [ABSTRACT FROM AUTHOR]

Abstract Background Since long, triamcinolone and fluocinolone, fluorinated derivatives, have been used for treating various types of dermatitis. Rosacea and perioral dermatitis are the most common side effects observed with prolonged use of fluorinated derivatives. Recent studies have shown that these adverse effects are more severe at low doses of fluorinated derivatives compared to low doses of non-fluorinated derivatives. Therefore, this study focused on impurity profiling through force degradation studies of hydrocortisone butyrate in a 0.1% lotion. Results A precise and robust method with low concentration was established for the estimation of hydrocortisone butyrate in bulk and formulations was done using the RP-HPLC technique. The mobile phase consisted of a combination of acetonitrile and purified water (30:70% v/v) and acetonitrile and purified water (95:5% v/v), with detection at a wavelength of 254 nm and a total run time of 20 min. The method demonstrated linearity and accuracy within the concentration range of 0.1–250 µg/mL and 50–125 µg/mL, respectively, with an r2 value of 0.999. Stress stability studies were conducted on hydrocortisone butyrate, revealing 11% degradation in alkaline conditions and 18% degradation in photolytic conditions. Conclusion The established method can be commercially used as it exhibits excellent linearity. Impurities were identified by injecting the reference standard, and their retentions were confirmed. The identified impurities included hydrocortisone, hydrocortisone-21-butyrate, hydrocortisone 3-methyl enol ether 17-butyrate, and hydrocortisone 17, 21-methylorthobutyrate, with retention times of 2.89, 9.14, 13.70, and 16.25 min, respectively. This precise method can be utilized in commercial applications for the accurate identification of hydrocortisone butyrate at low concentrations.

Abstract Cardiovascular disorders are among the leading causes of death worldwide, especially hypertension, a silent killer syndrome requiring multiple drug therapy for appropriate management. Hydrochlorothiazide is an extensively utilized thiazide diuretic that combines with several antihypertensive drugs for effective treatment of hypertension. In this study, sustainable, innovative and accurate high performance liquid chromatographic methods with diode array and tandem mass detectors (HPLC–DAD and LC–MS/MS) were developed, optimized and validated for the concurrent determination of Hydrochlorothiazide (HCT) along with five antihypertensive drugs, namely; Valsartan (VAL), Amlodipine besylate (AML), Atenolol (ATN), Amiloride hydrochloride (AMI), and Candesartan cilextil (CAN) in their diverse pharmaceutical dosage forms and in the presence of Chlorothiazide (CT) and Salamide (DSA) as HCT officially identified impurities. The HPLC–DAD separation was achieved utilizing Inertsil ODS-3 C18 column (250 × 4.6 mm, 5 μm) attached with photodiode array detection at 225.0 nm. Gradient elution was performed utilizing a mixture of solvent A (20.0 mM potassium dihydrogen phosphate, pH 3.0 ± 0.2, adjusted with phosphoric acid) and solvent B (acetonitrile) at ambient temperature. Linearity ranges were 0.1–100.0 µg/mL for HCT, VAL, AML and CAN, 0.05 –100.0 µg/mL for both ATN and AMI and 0.05–8.0 µg/mL for both CT and DSA. Additionally, this work describes the use of liquid chromatography–electrospray–tandem mass spectrometry for the accurate detection and quantification of the impurities; CT and DSA in the negative mode utilizing triple quadrupole mass spectrometry. The linearity ranges for those impurities were 1.0–200.0 ng/mL and 5.0–200.0 ng/mL for CT and DSA, respectively. Developed methods’ validation was achieved in accordance with International Conference on Harmonization (ICH) guidelines. Upon applying liquid chromatographic techniques for the drug analysis, a green and sustainable assessment have to be handled due to the consumption of energy and many solvents. Through the use of the HEXAGON, Analytical Greenness (AGREE) and White Analytical Chemistry (WAC) tools, greenness and sustainability have been statistically assessed. The optimized HPLC–DAD and LC–MS/MS methods were fast, accurate, precise, and sensitive, and consequently could be applied for conventional analysis and quality control of the proposed drugs in their miscellaneous dosage forms for the purpose of reducing laboratory wastes, time of the analysis time, effort, and cost. Graphical Abstract

A systematic literature survey published in several journals of pharmaceutical chemistry and of chromatography used to analyze impurities for most of the drugs that have been reviewed. This article covers the period from 2016 to 2020, in which almost of chromatographic techniques have been used for drug impurity analysis. These chromatography techniques are important in the analysis and description of drug impurities. Moreover, some recent developments in forced impurity profiling have been discussed, such as buffer solutions, mobile phase, columns, elution modes, and detectors are highlighted in drugs used for the study. This primarily focuses on thorough updating of different analytical methods which include hyphenated techniques for detecting and quantifying impurity and degradation levels in various pharmaceutical matrices. [ABSTRACT FROM AUTHOR]

Nearly a decade ago, fentanyl reappeared in the United States illicit drug market. In the years since, overdose deaths have continued to rise as well as the amount of fentanyl seized by law enforcement agencies. Research surrounding fentanyl production has been beneficial to regulatory actions and understanding illicit fentanyl production. In 2017, the Drug Enforcement Administration (DEA) began collecting seized fentanyl samples from throughout the United States to track purity, adulteration trends, and synthetic impurity profiles for intelligence purposes. The appearance of a specific organic impurity, phenethyl‐4‐anilino‐N‐phenethylpiperidine (phenethyl‐4‐ANPP) indicates a shift in fentanyl production from the traditional Siegfried and Janssen routes to the Gupta‐patent route. Through a collaboration between the DEA and the US Army's Combat Capabilities Development Command Chemical Biological Center (DEVCOM CBC), the synthesis of fentanyl was investigated via six synthetic routes, and the impurity profiles were compared to those of seized samples. The synthetic impurity phenethyl‐4‐ANPP was reliably observed in the Gupta‐patent route published in 2013, and its structure was confirmed through isolation and structure elucidation. Organic impurity profiling results for illicit fentanyl samples seized in late 2021 have indicated yet another change in processing with the appearance of the impurity ethyl‐4‐anilino‐N‐phenethylpiperidine (ethyl‐4‐ANPP). Through altering reagents traditionally used in the Gupta‐patent route, the formation of this impurity was determined to occur through a modification of the route as originally described in the Gupta patent. [ABSTRACT FROM AUTHOR]

Impurities found in active pharmaceutical ingredients (APIs) and pharmaceutical products are of ever‐increasing interest. According to several regulatory agencies, purity and impurity profiles are essential. An impurity is defined as any additional inorganic or organic material, residual solvents other than the medicinal components, or undesired compounds that remain with APIs. Impurities and degradation products in bulk drug materials and pharmaceutical formulations are identified, their structures are clarified, and their quantitative determination is part of impurity profiling. Unrecognized, poisonous impurities are dangerous to health and should be identified by selective procedures to increase the safety of drug therapy, and impurity profiling has become more significant in pharmaceutical analysis. This review briefly introduces process and product‐related impurities and emphasizes the creation of cutting‐edge analytical techniques for identifying them. It discusses the use of analytical methods, particularly high‐performance thin‐layer chromatography, liquid chromatography with mass spectrometry (MS), ultrahigh‐performance liquid chromatography, gas chromatography–MS, and nuclear magnetic resonance spectroscopy for the identification of contaminants and degradation products. It has discussed the importance of the quality, efficacy, and safety of drug substances and products, including the origin, types, and quality control of impurities, the need for the development of impurity profiling methods, impurity identification, and regulatory aspects. [ABSTRACT FROM AUTHOR]

Cardiovascular disorders are among the leading causes of death worldwide, especially hypertension, a silent killer syndrome requiring multiple drug therapy for appropriate management. Hydrochlorothiazide is an extensively utilized thiazide diuretic that combines with several antihypertensive drugs for effective treatment of hypertension. In this study, sustainable, innovative and accurate high performance liquid chromatographic methods with diode array and tandem mass detectors (HPLC–DAD and LC–MS/MS) were developed, optimized and validated for the concurrent determination of Hydrochlorothiazide (HCT) along with five antihypertensive drugs, namely; Valsartan (VAL), Amlodipine besylate (AML), Atenolol (ATN), Amiloride hydrochloride (AMI), and Candesartan cilextil (CAN) in their diverse pharmaceutical dosage forms and in the presence of Chlorothiazide (CT) and Salamide (DSA) as HCT officially identified impurities. The HPLC–DAD separation was achieved utilizing Inertsil ODS-3 C18 column (250 × 4.6 mm, 5 μm) attached with photodiode array detection at 225.0 nm. Gradient elution was performed utilizing a mixture of solvent A (20.0 mM potassium dihydrogen phosphate, pH 3.0 ± 0.2, adjusted with phosphoric acid) and solvent B (acetonitrile) at ambient temperature. Linearity ranges were 0.1–100.0 µg/mL for HCT, VAL, AML and CAN, 0.05 –100.0 µg/mL for both ATN and AMI and 0.05–8.0 µg/mL for both CT and DSA. Additionally, this work describes the use of liquid chromatography–electrospray–tandem mass spectrometry for the accurate detection and quantification of the impurities; CT and DSA in the negative mode utilizing triple quadrupole mass spectrometry. The linearity ranges for those impurities were 1.0–200.0 ng/mL and 5.0–200.0 ng/mL for CT and DSA, respectively. Developed methods' validation was achieved in accordance with International Conference on Harmonization (ICH) guidelines. Upon applying liquid chromatographic techniques for the drug analysis, a green and sustainable assessment have to be handled due to the consumption of energy and many solvents. Through the use of the HEXAGON, Analytical Greenness (AGREE) and White Analytical Chemistry (WAC) tools, greenness and sustainability have been statistically assessed. The optimized HPLC–DAD and LC–MS/MS methods were fast, accurate, precise, and sensitive, and consequently could be applied for conventional analysis and quality control of the proposed drugs in their miscellaneous dosage forms for the purpose of reducing laboratory wastes, time of the analysis time, effort, and cost. [ABSTRACT FROM AUTHOR]

Two unknown impurities were detected during the related substance analysis of the Silodosin crude sample using High- Performance Liquid Chromatography. These impurities were isolated using preparative HPLC and then subjected to LCMS for mass determination. Structural elucidation has been carried out using NMR (¹H NMR, 13C NMR) spectroscopy. These impurities were discovered to have a structural alert, which was confirmed by the Insilico prediction model indicating that they were Potentially Genotoxic Impurities (PGI's). A genotoxic assessment of these impurities was done using the Insilico prediction model for genotoxicity (VEGA and Toxtree). These impurities have not been reported elsewhere, to the best of our knowledge. Thus, a HPLC-UV method was developed and validated for the determination of these impurities at the TTC level. The proposed method was validated in terms of Specificity, Precision, Accuracy, Linearity and Robustness as per ICH guidelines. The LOD and LOQ of the method were established to be 0.0018 ppm and 0.055 ppm with respect to test concentration (1000 µg mL-1 silodosin). [ABSTRACT FROM AUTHOR]

Forced degradation studies have been used to simplify analytical methodology development and achieve a deeper knowledge about the inherent stability of active pharmaceutical ingredients (API) and drug products. This provides insight into degradation species and pathways. Identification of impurities in pharmaceutical products is closely related to the selection of the most appropriate analytical methods like HPLC-UV, LC-MS/MS, LC-NMR, GC-MS, and capillary electrophoresis. Herein, recent trends in analytical perspectives during 2018–April 14, 2021, are discussed based on forced and impurity degradation profiling of pharmaceuticals. Literature review showed that several methods have been used for experimental design and analysis conditions such as matrix type, column type, mobile phase, elution modes, detection wavelengths, and therapeutic category. Thus, since these factors influence the separation and identification of the impurities and degradation products, we attempted to perform a statistical analysis for the developed methods according to the abovementioned factors. [ABSTRACT FROM AUTHOR]

A previously developed high‐performance liquid chromatography method combined with pulsed amperometric detection allowed to separate many impurities of paromomycin. However, due to the presence of ion pairing agents and sodium hydroxide in the mobile phase, direct coupling to mass spectrometry for the identification of the chemical structures of the impurities was not an option. Indeed, ion suppression was encountered by trifluoroacetic acid and pentafluoroproponic acid in the mobile phase. A cation self‐regenerating suppressor, which was originally designed for increasing analyte conductivity of ammonia and amines analysis in ion chromatography, was coupled between the liquid chromatography and ion trap‐time of flight‐mass spectrometry and almost all trifluoroacetic acid and pentafluoroproponic acid in the mobile phase was removed. The limit of detection of paromomycin in this integrated system improved significantly to 20 ng/ml (0.4 ng). The chemical structures of 19 impurities were elucidated and seven impurities were reported for the first time. [ABSTRACT FROM AUTHOR]

Identification and characterization of biproducts/ impurities present in agrochemicals are critical in view of their efficacy and safety towards public health. We herein present our study on identification and characterization of an impurity, 5‐chloro‐2‐cyano‐N,N‐dimethyl‐4‐p‐tolylimidazole‐1‐sulfonamide (2) present in the fungicide, "cyazofamid". Intermittent HPLC analysis of the reaction of substituted imidazole (1) with N,N‐dimethylsulfamoyl chloride suggested that 2 is formed during the reaction. Isolation by preparative HPLC and characterization by NMR, LC/HRMS, MS/MS and single crystal XRD analysis confirmed 2 as an isomer of cyazofamid, wherein the N,N‐dimethyl sulfonamide group was positioned on the other nitrogen of imidazole in close proximity to chloride group. Computational studies further supported the formation of 2 and ruled out the other possible isomeric structures. [ABSTRACT FROM AUTHOR]

Apalutamide, an androgen receptor inhibitor, is used to treat prostate cancer. A stability‐indicating high‐performance liquid chromatography method was developed for the estimation of assay and organic impurities of apalutamide in drug substance and in tablet dosages using Design of Experiments. The chromatographic separation was achieved within 30 min using Atlantis dC18, 100 × 4.6 mm, 3.0 μm and the binary gradient program (10 mm KH2PO4, pH 3.5; acetonitrile). The detection wavelength, flow rate, column temperature and injection volume used were 270 nm, 1.0 ml/min, 45°C and 10 μl, respectively. The interaction of independent variables (pH, column temperature and flow rate) and their influences on HPLC parameters were studied using a central composite design, and then the peak separation and elution behaviors between apalutamide and its seven impurities were determined. The method validation was performed for linearity, detection limit, quantitation limit, accuracy, precision and robustness as per the International Conference on Harmonization. A high‐quality recovery with good precision (91.7–106.0%) and correlations (r2 > 0.997) within a linear range of 0.12–2.24 μg/ml (0.05–0.3%, w/w) were achieved consistently for assay and organic impurities of apalutamide. The stability‐indicating characteristics of the proposed method were assessed through forced degradation and mass balance studies. An effort was made to figure out the chemical structures of newly formed degradation products (DP1–DP5) using LC–MS/MS. [ABSTRACT FROM AUTHOR]

Omeprazole (OME) is a proton pump inhibitor used to treat gastroesophageal reflux disease associated conditions. The current study presents an Analytical Quality by Design-based approach for the development of a CE method for OME impurity profiling. The scouting experiments suggested the selection of solvent modified Micellar ElectroKinetic Chromatography operative mode using a pseudostationary phase composed of sodium dodecyl sulfate (SDS) micelles and n -butanol as organic modifier in borate buffer. A symmetric three-level screening matrix 37//16 was used to evaluate the effect of Critical Method Parameters, including Background Electrolyte composition and instrumental settings, on Critical Method Attributes (critical resolution values, OME peak width and analysis time). The analytical procedure was optimized using Response Surface Methodology through a Central Composite Orthogonal Design. Risk of failure maps made it possible to define the Method Operable Design Region, within which the following optimized conditions were selected: 72 mM borate buffer pH 10.0, 96 mM SDS, 1.45 %v/v n -butanol, capillary temperature 21 °C, applied voltage 25 kV. The method was validated according to ICH guidelines and robustness was evaluated using a Plackett-Burman design. The developed procedure enables the simultaneous determination of OME and seven related impurities, and has been successfully applied to the analysis of pharmaceutical formulations. [Display omitted] • A CE method for the analysis of Omeprazole and its impurities was set up. • Analytical Quality by Design principles were employed to develop the procedure. • The knowledge space was explored through a 37//16 symmetric screening matrix. • In Response Surface Methodology a Central Composite Orthogonal Design was used. • Probability maps allowed the Method Operable Design Region to be defined. [ABSTRACT FROM AUTHOR]

Capillary electrophoresis (CE) is widely used for the impurity profiling of drugs that contain stereochemical centers in their structures, analysis of biomolecules, and characterization of biopharmaceuticals. Currently, CE is the method of choice for the analysis of foodstuffs and the determination of adulterants. This article discusses the general theory and instrumentation of CE as well as the classification of various CE techniques. It also presents an overview of research on the applications of different CE techniques in the impurity profiling of drugs in the past decade. The review briefly presents a comparison between CE and liquid chromatography methods and highlights the strengths of CE using drug compounds as examples. This review will help scientists, fellow researchers, and students to understand the applications of CE techniques in the impurity profiling of drugs.

Amitriptyline hydrochloride is an antidepressant drug with sedative effects used to treat the symptoms of anxiety, agitation with depression and schizophrenia with depression. A reversed‐phase high‐performance liquid chromatography method was developed to separate and quantitatively determine the assay and four organic impurities of amitriptyline in tablet dosage form and bulk drugs using a C18 column in an isocratic elution mode with mobile phase consisting of a mixture of pH 7.5 phosphate buffer and methanol. The pH conditions used in the chromatographic separation are discussed. The stability‐indicating characteristics of the proposed method were proved using stress testing [5 m HCl at 80°C/1 h, 5 m NaOH at 80°C/1 h, H2O (v/w) at 80°C/1 h, 6% H2O2 (v/v) at 25°C/1 h, dry heat at 105°C/24 h and UV–vis light/4 days] and validated for specificity, detection limit, quantitation limit, linearity, precision, accuracy and robustness. For amitriptyline and its four known organic impurities, the quantitation limits, linearity and recoveries were in the ranges 0.25–3.0 μg/ml (r2 > 0.999) and 87.9–107.6%, respectively. The mass (m/z) spectral data of amitriptyline hydrochloride and its impurity are discussed. The proposed LC method is also suitable for impurity profiling and assay determination of amitriptyline in bulk drugs and pharmaceutical formulations. [ABSTRACT FROM AUTHOR]

This article described the development and validation of a method for spiramycin related substances based on hybrid particle column. The chromatographic conditions were as follows: water - 0.2 mol/L dipotassium hydrogen phosphate (the pH value adjusted to 9.5 using a 1 mol/L KOH solution) - acetonitrile - methanol (10: 60: 28.5: 1.5, v/v/v/v) as mobile phase A, water - 0.2 mol/L dipotassium hydrogen phosphate (pH 9.5) - acetonitrile - methanol (10: 30: 57: 3, v/v/v/v) as mobile phase B and gradient elution was performed. Compared with previous analytical methods, this method has strong specificity, excellent sensitivity and stability, which could be used for the daily testing of related substances of spiramycin. Furthermore, impurities above 0.1 % were characterized using two-dimensional liquid chromatography coupled with quadrupole time-of-flight mass spectrometer (2D LC-QTOF-MS/MS) and there were 6 impurities reported for the first time., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Abstract Background An accurate, precise and robust analytical method was developed for the impurity profiling in the metformin hydrochloride and teneligliptin hydrobromide hydrate tablet. The gradient was optimized for better separation of impurities by using BDS Hypersil C18 250 × 4.6 mm, 5µ column operated at 35 °C. The octane sulfonic acid and phosphate buffer with triethylamine at pH 3.0 were used as mobile phase A, and acetonitrile was used as mobile phase B. The mobile phase was pumped at 1.0 mL/min. The gradient was optimized for better resolution, and the chromatogram was monitored at 210 nm. Results The % recovery of teneligliptin and metformin HCL observed was above 90% from LOQ level to 150%. The correlation coefficient r 2 was 0.999 for metformin HCl, teneligliptin, melamine, cyanocobalamin, teneligliptin impurity A and 0.998 for teneligliptin impurity B. The method was found unaffected by change in method variance during the robustness study. During the stress study with acid, base, peroxide and temperature, maximum degradation was observed with peroxide indicating the sensitivity of the molecule toward oxidative stress. Conclusions The developed method is precise, accurate, robust and linear and hence can be routinely used for the related substance analysis of metformin hydrochloride and teneligliptin hydrobromide hydrate tablet in the quality control laboratory at manufacturing site during the commercial manufacturing.

Dinotefuran (DNT) is a neonicotinoid insecticide widely used in pest control. Identification of structurally related impurities is indispensable during material purification and pesticide registration and certified reference material development, and therefore needs to be carefully characterized. In this study, a combined strategy with liquid chromatography high-resolution mass spectrometry and SIRIUS has been developed to elucidate impurities from DNT material. MS and MS/MS spectra were used to score the impurity candidates by isotope score and fragment tree in the computer assisted tool, SIRIUS. DNT, the main component, worked as an anchor for formula identification and impurity structure elucidation. With this strategy, two by-product impurities and one stereoisomer were identified. Their fragmentation pathways were concluded, and the mechanism for impurity formation was also proposed. This result showed a successful application for combined human intelligence and machine learning, in the identification of pesticide impurities. [ABSTRACT FROM AUTHOR]

The investigation's objective was to access the stability of the drug substance Pimozide under the stress conditions specified in the International Conference on Harmonization Q1A(R2) guideline and to separate, identify, and characterize the degradation products by using liquid chromatography‐tandem mass spectrometry studies without isolating them from the reaction mixture. It was observed that the drug was susceptible to oxidative degradation (15% hydrogen peroxide, 48 h, room temperature) forming four novel degradation products while being stable under hydrolytic, Photolytic, and thermal conditions. The separation of the drug and the degradation products was achieved with good resolution on a Phenomenex C18 column (150 mm × 4.6 mm, 5 µm) using gradient elution. The fragmentation pattern constructed from the liquid chromatography‐quadrupole‐time of flight mass spectrometry data was used for structural characterization of the degradation products. Lastly, the in‐silico absorption, distribution, metabolism, excretion, and toxicity properties prediction was performed by using in‐silico tools like the pKCSM webserver, ToxTree, and OSIRIS property explorer. The primary screening through the pKCSM webserver exposed the hepatoxic and the mutagenic potential of Pimozide, 1‐(1‐(4‐(4‐fluorophenyl)‐4‐phenylbutyl) piperidin‐4‐yl)‐1,3‐dihydro‐2H‐benzo[d]imidazol‐2‐one (degradation product 3) and 1‐(4,4‐bis(4‐fluorophenyl)butyl)‐4‐(2‐oxo‐2,3‐dihydro‐1H‐benzo[d]imidazol‐1‐yl)piperidine 1‐oxide (degradation product 5) whereas, 1‐(1‐(4,4‐diphenylbutyl) piperidin‐4‐yl)‐2,3‐dihydro‐1H‐benzo[d]imidazol‐2‐ol (degradation product 1) was predicted to only possess mutagenic potential. Secondary screening for mutagenicity was done by using ToxTree and OSIRIS property explorer. [ABSTRACT FROM AUTHOR]

The group of P2P precursors including α‐phenylacetoacetonitrile (APAAN), α‐phenylacetoamide (APAA) and methyl α‐acetylphenylacetate (MAPA) has become increasingly popular in Europe and other parts of the world in the last decade. Previous investigations have reported the use of APAAN in the synthesis of amphetamine and methamphetamine and identified a range of characteristic impurities. This research has expanded upon the current literature by investigating the use of MAPA in the synthesis of methamphetamine. In this study methamphetamine was synthesized via three common clandestine methods: the Leuckart synthesis and two reductive amination methods. We report the identification of seven impurities, four of which are methyl ester equivalents of impurities previously reported for the detection of APAAN. These are methyl 2‐phenylbut‐2‐enoate, methyl 2‐phenyl‐3‐hydroxybutanoate, methyl 3‐(methylamino)‐2‐phenylbut‐2‐enoate and methyl 3‐(methylamino)‐2‐phenylbutanoate. The other impurities identified are ethyl ester compounds formed via transesterification of the methyl ester due to the reaction solvent. This susceptibility for transesterification suggests that identification of the pre‐precursor used may not always be straightforward and may be dependent on the reaction conditions employed. Of the impurities reported, methyl 3‐(methylamino)‐2‐phenylbutanoate was deemed to be a potentially reliable impurity for detection of the use of MAPA; however, it is expected that lower levels of characteristic impurities may be detected in methamphetamine synthesized from MAPA than that from APAAN. [ABSTRACT FROM AUTHOR]

In 2018, the discovery of carcinogenic nitrosamine process related impurities (PRIs) in a group of widely used drugs led to the recall and complete withdrawal of several medications that were consumed for a long time, unaware of the presence of these genotoxic PRIs. Since then, PRIs that arise during the manufacturing process of the active pharmaceutical ingredients (APIs), together with their degradation impurities, have gained the attention of analytical chemistry researchers. In 2020, favipiravir (FVR) was found to have an effective antiviral activity against the SARS-COVID-19 virus. Therefore, it was included in the COVID-19 treatment protocols and was consequently globally manufactured at large-scales during the pandemic. There is information indigence about FVR impurity profiling, and until now, no method has been reported for the simultaneous determination of FVR together with its PRIs. In this study, five advanced multi-level design models were developed and validated for the simultaneous determination of FVR and two PRIs, namely; (6-chloro-3-hydroxypyrazine-2-carboxamide) and (3,6-dichloro-pyrazine-2-carbonitrile). The five developed models were classical least square (CLS), principal component regression (PCR), partial least squares (PLS), genetic algorithm-partial least squares (GA-PLS), and artificial neural networks (ANN). Five concentration levels of each compound, chosen according to the linearity range of the target analytes, were used to construct a five-level, three-factor chemometric design, giving rise to twenty-five mixtures. The models resolved the strong spectral overlap in the UV-spectra of the FVR and its PRIs. The PCR and PLS models exhibited the best performances, while PLS proved the highest sensitivity relative to the other models. [ABSTRACT FROM AUTHOR]

Technical advances in the field of quality analysis allow an increasingly deeper look into the impurity profile of drugs. The ability to detect unexpected impurities in addition to known impurities ensures the supply of high-quality drugs and can prevent recalls due to the detection of harmful unexpected impurities, as has happened recently with the N -nitrosamine and azido impurities in losartan (LOS) drug products. In the present study, the LC-MS/HRMS approach described by Backer et al. was applied to an even more complex system, being the investigation of 35 LOS drug products and combination preparations purchased in 2018 and 2022 in German pharmacies. The film-coated tablets were analysed by means of four LC-MS/HRMS method variants. For the separation a Zorbax RR StableBond C18 column (3.0 ×100 mm, particle size of 3.5 µm, pore size of 80 Å), a gradient elution and for mass spectrometric detection a qTOF mass spectrometer with electrospray ionization in positive and negative mode was used. An information-dependent acquisition method was applied for the acquisition of high-resolution mass spectrometry data. The combination of an untargeted and a targeted screening approach revealed the finding of eight impurities in total. Beside the five LOS related compounds, LOS impurity F, J, K, L, M, and related compound D from amlodipine besilate, LOS azide and an unknown derivative thereof were detected. Identification and structure elucidation, respectively, were successfully performed using in silico fragmentation. Differences in the impurity profiles of drug products from 2018 and 2022 could be observed. This study shows that broad screening approaches like this are applicable to the analysis of drug products and can be an important enhancement of the quality assurance of medicinal products. [Display omitted] • Analysis of losartan finished dosage forms by means of HRMS. • Structural elucidation by means of MS and MS/MS data. • Verification of proposed structural formulae by in silico fragmentation. • Comparison of the impurity profiles of losartan tablets purchased in 2018 and 2022. • Development of a procedure which provides a broad look into the impurity profile of drugs. [ABSTRACT FROM AUTHOR]

Sarin is a highly toxic nerve agent classified by the Chemical Weapon Convention as a Schedule 1 chemical with no use other than to kill or injure. Moreover, in recent times, chemical warfare agents have been deployed against both military and civilian populations. Chemical warfare agents always contain minor impurities that can provide important chemical attribution signatures (CAS) that can aid in forensic investigations. In order to understand the trace molecular composition of sarin, various analytical approaches including GC–MS, LC–MS and NMR were used to determine the chemical markers of a set of sarin samples. Precursor materials were studied and the full characterisation of a synthetic process was undertaken in order to provide new insights into potential chemical attribution signatures for this agent. Several compounds that were identified in the precursor were also found in the sarin samples linking it to its method of preparation. The identification of these CAS contributes critical information about a synthetic route to sarin, and has potential for translation to related nerve agents. [ABSTRACT FROM AUTHOR]

Montelukast sodium (MLS) is a leukotriene receptor antagonist drug used in the treatment of asthma, bronchospasm, allergic rhinitis and urticaria. A reversed‐phase high performance liquid chromatography method was developed to separate, identify and quantitative determination of MLS and its eight known organic impurities in tablet dosage form using a C18 column and mobile phases consisting of a gradient mixture of pH 2.5 phosphate buffer and acetonitrile. The stability‐indicating character of the developed method was proven using stress testing (1 m HCl at 80°C/30 min, 1 m NaOH at 80°C/30 min, H2O at 80°C/30 min, 3% H2O2 at 25°C/1 min, dry heat at 105°C/10 h and UV–vis light/4 days) and was validated for specificity, quantitation limit, linearity, precision, accuracy and robustness. For MLS and its eight known impurities, the quantitation limits, linearity and recoveries were 0.015–0.03 μg/ml, correlation coefficient > 0.997 (R2 > 0.995) and 85.5–107.0%, respectively. The developed chromatographic method is suitable for impurity profiling and also for assay determination of MLS in bulk drugs and pharmaceutical formulations. The mass values (m/z) of newly formed degradation products (DP1 and DP2) of montelukast sodium were identified using liquid chromatography–mass spectrometry. [ABSTRACT FROM AUTHOR]

An HPLC method has been described in the European Pharmacopoeia and United States Pharmacopeia for the determination of nine organic impurities (imp A–I) in fingolimod hydrochloride, a synthetic sphingosine‐1‐phosphate receptor modulator. The manufacturing process of fingolimod hydrochloride consists of multistep chemical synthesis wherein controls of precursors, intermediates and process steps should be performed to assure the final quality of the drug substance. We synthesized and isolated eight process‐related impurities (FINI imp A–H) of fingolimod, which were different from the pharmacopoeial impurities. One unknown process‐related impurity was found as a key intermediate (FINI) and was identified by LC–MS. Characterization of all of the impurities were done using spectroscopic techniques (1H and 13C NMR, FTIR, MS), and the mechanistic pathways to the formation of these impurities were also discussed. Two of these impurities were evaluated as potential genotoxic impurities owing to their alerting structures and alkylating properties (alkyl sulfonates and alkyl halides, class 3, ICH M7). We also developed and validated an RP‐UPLC method in line with ICH Q2 guidelines for control these impurities (FINI imp A–H) and to assure the pharmacopoeial quality drug substance. [ABSTRACT FROM AUTHOR]

Capillary electrophoresis (CE) is widely used for the impurity profiling of drugs that contain stereochemical centers in their structures, analysis of biomolecules, and characterization of biopharmaceuticals. Currently, CE is the method of choice for the analysis of foodstuffs and the determination of adulterants. This article discusses the general theory and instrumentation of CE as well as the classification of various CE techniques. It also presents an overview of research on the applications of different CE techniques in the impurity profiling of drugs in the past decade. The review briefly presents a comparison between CE and liquid chromatography methods and highlights the strengths of CE using drug compounds as examples. This review will help scientists, fellow researchers, and students to understand the applications of CE techniques in the impurity profiling of drugs. [Display omitted] • An overview of research related to the use of capillary electrophoresis in the impurity profiling of drugs is presented. • The principle, instrumentation, and different types of capillary electrophoresis (CE) methods are outlined here. • Applications of different of CE methods with the chemical structures of drugs and their impurities are highlighted. • A brief description is also provided on the analysis of Pharmacopeial monographs using CE methods. • A comparison of CE with liquid chromatography for impurity profiling and analysis of drugs is presented in this review. [ABSTRACT FROM AUTHOR]

• Impurities and adulterants in cocaine are assessed as candidates for drug profiling. • Temporal and spatial variation in occurrence allows for forensic intelligence. • A selection of candidates are detectable in routinely available GC-MS data. • Retrospective analysis may lead to additional insights out of already existing data. Cocaine is one of the most widely used illicit drugs worldwide. Cocaine powders seized by the Police may contain numerous other substances besides the drug itself. These can be impurities originating from the coca plant or the production process, or be purposely added to the drug formulation as adulterants and cutting agents. In forensic laboratories, identification of cocaine is routinely done through GC-MS analysis, but other components are often ignored even if the method allows for their detection. Yet, they can provide valuable insight into the history of a seizure and its potential connection to other samples. To explore this idea, an extensive review of common impurities and adulterants encountered in cocaine is presented. Based on their incidence, concentration in the end product and compatibility with GC-MS methods, their overall usefulness as candidates for the statistical investigation of existing forensic data is evaluated. The impurities cis- and trans- cinnamoylcocaine, tropacocaine, norcocaine and N -benzoylnormethylecgonine as well as the adulterants lidocaine, procaine, tetracaine, benzocaine, caffeine, acetylsalicylic acid, phenacetin, ibuprofen, levamisole, hydroxyzine and diltiazem are promising candidates to provide additional forensic intelligence. Future research on optimized routine GC-MS methods, signal reproducibility, comparison, statistics and databases is suggested to facilitate this concept. Ultimately, such an approach may significantly advance the amount of information that is extracted from routine casework data, elucidate developments in the cocaine markets in the past and facilitate Police work in the future. Preliminary assessment of existing data from the forensic laboratory of the Amsterdam Police has been included to show that the detection of the identified target impurities is feasible, and that small adjustments to the analysis method could significantly increase the detectability of these analytes in prospective drug screenings. Forensic intelligence based on retrospective data mining of cocaine containing casework samples may thus be realized with minimal additional laboratory efforts by using already available instrumentation, samples and data. [ABSTRACT FROM AUTHOR]

Background: Stability studies, stress testing of drug substances and finished drug product is obligatory due to the guidelines by the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) and other Regulatory Authorities. Materials and Methods: In the present work the selective Beta-1 receptor blocker, Bisoprolol fumarate was subjected to the forced degradation studies which includes hydrolysis (acidic, basic and neutral), oxidative, photolytic and thermal degradation. The drug was stable under light and basic and neutral medium. Results: The major findings in the presented work are the degradation products formed in the course of thermal degradation. The complete thermal degradation products were identified by High Performance Liquid Chromatography, Quadrupole-Time of Flight micromass (Electronspray Ionization-Mass Spectroscopy). The chromatographic conditions opted for the study includes Cosmosil C-18 column (4.6ID*250mm) with mobile phase consisting of methanol: phosphate buffer (pH 3.5 adjusted with orthophosphoric acid): Acetonitrile (45:35:20). Conclusion: The stability indicating method was validated as per the International Conference on Harmonization guidelines. The use of superior statistical tools for validation ensures the efficiency, quality and reproducibility of the presented method. Results obtained were agreeable and ensured the quality and reproducibility of the method. [ABSTRACT FROM AUTHOR]

Aim: The purpose of this research study, is to develop and validate a reverse phase HPLC test method for detecting relevant impurities in Testosterone cypionate (TCY). Materials and Methods: The chromatographic system for separation of related impurities were achieved in Zorbax XDB-C8 (15 cm x 4.6 mm), 5 micron HPLC column utilising gradient elution technique. Water was selected as solvent-A and Acetonitrile was preferred as solvent-B for mobile phase. The method is gradient technique. Column heater was kept constant at 35°C; the rate of flow was 1.2 mL per min; volume of injection was 20 µL and 240 nm was set for detector wavelength. Results: The % recovery was in the range of 95.6% to 108.7% for all impurities. The result of correlation coefficient were higher than 0.98. Testosterone is the major degradants obtained from forced degradation study. Conclusion: The created method can be utilise in quality control testing on a regular basis for the analysis of Testosterone cypionate. [ABSTRACT FROM AUTHOR]

The rise in popularity of 'designer' precursor compounds for the synthesis of amphetamine‐type stimulants poses a significant challenge to law enforcement agencies. One such precursor is α‐phenylacetoacetonitrile (APAAN). APAAN emerged in Europe in 2010 and quickly became one of the most popular precursors for amphetamine synthesis in that region. Previous literature has identified four APAAN‐specific impurities formed in the synthesis of amphetamine; however, there is currently no research on the use of APAAN in the synthesis of methamphetamine, which is more likely to be employed in a non‐European market. In this study methamphetamine was synthesised via three common clandestine methods: the Leuckart method and two reductive amination methods. We report the identification of five new impurities and two previously identified impurities characteristic for the use of APAAN in the synthesis of methamphetamine. The newly identified impurities were characterised by MS and NMR and determined to be (E)‐3‐(methylamino)‐2‐phenylbut‐2‐enenitrile, 3‐(methylamino)‐2‐phenylbutanenitrile, 3‐methyl‐2,4‐diphenylpentanedinitrile, 2‐phenylbutyronitrile and 3‐hydroxy‐2‐phenylbutanenitrile. [ABSTRACT FROM AUTHOR]

Midostaurin (MDS) is used for the treatment of acute myeloid leukemia, myelodysplastic syndrome, and advanced systemic mastocytosis. MDS softgel capsule samples were subjected to stress testing per International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use guidelines for impurity profiling study. MDS underwent extensive degradation under stress testing (acid, alkaline, oxidative, photolytic, thermolytic, and hydrolysis conditions) and formed four degradation products (DPs). MDS and its DPs were separated well from one another with good resolution using reserved‐phase HPLC using an Inertsil ODS‐3V column (250 × 4.6 mm, 5 μm) and a mobile phase of ammonium formate (40 mM) and acetonitrile. The stability‐indicating characteristic of the newly developed method was proven for the estimation of MDS assay, and its organic impurities were free from interference. The validated method exhibited excellent linearity, accuracy, precision, specificity, detection limit, and quantitation limit within 25 min run time. Stress testing, robustness, and solution stability were performed to ensure the continuous performance of the developed method. The peak fractions of DPs formed under stress testing were isolated and characterized using LC–MS, 1H and 13C NMR, IR, and UV–Vis. The structure of the major DPs was predicted as DP1 based on the spectral data. The proposed method is effectively used for MDS in bulk drug and finished formulations in the pharmaceutical industry. [ABSTRACT FROM AUTHOR]

A new RP‐HPLC method with a quick, sensitive and stable indication for the quantitative measurement of selexipag and its associated substances was developed and validated in the present study. In this new method, using the impurity‐spiked solution, the chromatographic approach was optimized. Similarly, using the X‐bridge phenyl column with isocratic elution of mobile phase containing acetonitrile and formic acid, selexipag and its impurities were separated. Recovery experiments obtained were satisfactory, and also the calibration graphs plotted for selexipag and its five impurities were found to be linear. The system validation parameters such as specificity, linearity, precision, accuracy and robustness were determined successfully. The obtained results indicated that the developed method was found to be useful for analyzing selexipag from its impurities. Further, using stress tests against acid, alkali, peroxide, reduction, thermal, hydrolysis and UV conditions, the present established method of HPLC was assessed and validated as per ICH Q2(R1) guidelines. [ABSTRACT FROM AUTHOR]

The GABAB receptor agonist baclofen is a medication commonly used for the treatment of muscle spasticity. It is an amino acid and related to the neurotransmitter GABA. In this study, we developed a new, gradient high-performance liquid chromatography (HPLC) method for the impurity assessment of baclofen, which is appropriate for pharmacopoeial purposes. Since the impurities related to the synthesis pathway are acids, zwitterionic, or neutral, the method development is challenging. However, the separation of all components was achieved on a C18 stationary phase using a water–acetonitrile–trifluoroacetic acid gradient. A limit of detection (LOD) of at least 0.02% was registered for all specified impurities. Additionally, CAD detection was performed to detect potential impurities lacking off a chromophore. The baclofen batches analyzed are far more pure than expected. All impurities were found below the specification limit, and thus, they can be regarded as unspecified. Moreover, the required runtime could be significantly reduced compared to the current USP or Ph. Eur. method. [ABSTRACT FROM AUTHOR]

Two selective, sensitive and environmentally safe LC methods were developed and validated for determination of paracetamol, caffeine, ergotamine tartrate and metoclopramide in coformulated antimigraine tablets along with p‐aminophenol, p‐nitrophenol and theophylline as officially specified impurities. The first is based on high‐performance thin‐layer chromatography (HPTLC) coupled with densitometric quantitation. Separation was achieved on HPTLC silica gel 60 F254 plates as stationary phase using ethyl acetate:aqueous ammonium hydroxide solution:glacial acetic acid (10.0:0.4:0.1, by volume) as a developing system followed by scanning of the separated bands at 210.0 nm. The subsequent method depends on HPLC with diode array detection. The LC separation was accomplished on a Scharlau C18 (250 × 4.6 mm, 5 μm) column using a mixture of 20.0 mm sodium dihydrogen phosphate, pH 3.0, adjusted with o‐phosphoric acid and methanol, at a flow rate of 1.3 mL/min in a gradient elution program. The separated peaks were detected at 210.0 nm. The proposed methods have been validated and proven to meet the requirements outlined in the International Council for Harmonisation (ICH) guidelines. The greenness profile evaluation was carried out using three tools, namely, the National Environmental Method Index, the Analytical EcoScale and the Green Analytical Procedure Index tool, and a comparative study was then conducted. Successful application of the developed methods for determination of the cited quaternary mixture in Metograine tablets confirms their suitability regarding the analytical performance and ecological impact in quality control assay and impurity profiling purposes. [ABSTRACT FROM AUTHOR]

Several examples of the emergence of unexpected impurities in medicinal products in the past, e.g. the 2018 valsartan scandal, disclosed the need for sophisticated approaches in impurity profiling. Advanced techniques in mass spectrometry offer the possibility to detect impurities in untargeted approaches complementing the targeted search for potential impurities. In this study, a combination of targeted and untargeted approaches using LC-MS/HRMS was applied in creating an impurity profile of bisoprolol fumarate. In the targeted approach, a reaction matrix was used to predict potential impurities, which were searched for in addition to the related substances stated in the European Pharmacopoeia. For the untargeted analytics, general unknown comparative screening was performed to detect unexpected impurities. To cover a maximum range of detectable analytes, two complementary mobile phases, i.e. buffered to acidic and basic pH, were combined with four mass spectrometric ionization conditions, i.e. electrospray ionization and atmospheric pressure chemical ionization in both positive and negative mode. Information-dependent acquisition was used to generate MS and MS/MS data in a single run. The targeted and the untargeted approach revealed the presence of 18 and 17 impurities, respectively. The plausibility of proposed/elucidated structural formulae was checked by in silico fragmentation and assignment of characteristic fragments/neutral losses. Quantification of the impurities was performed with respect to the internal standard metoprolol. The analyzed batches showed contents of up to 0.05% of single impurities. A thorough procedure for the development of a complete impurity profile for drug substances was demonstrated. To ensure a maximum of patient safety, the described approach is prototypical and should be implemented during drug development and after relevant changes in manufacturing processes.

A simple, rapid, precise and rugged reverse phase high performance liquid chromatographic (RP-HPLC) method has been developed for identification and quantitation of Bilastine and Montelukast active pharmaceutical ingredients along with related impurities from bulk and tablet dosage form. The separation of analytes and impurities peaks were achieved using a HPLC system equipped with photo diode array detector. A Hypersil BDS C-18 column used as stationary phase and 2mM ammonium acetate buffer: Acetonitrile used as a mobile phase operated in a gradient mode at flow rate of 0.8ml/min to 1.0ml/min with 30°C column temperature. Injection volume was 10 µl and chromatograms were analyzed at 275 nm with run time of 25 min. Under above conditions the retention time (RT) of Bilastine and Montelukast was 9.4 and 16.5 min respectively. Spiked Bilastine and Montelukast impurities i.e. BLS imidazole imp, BLS DEO imp, BLS TSO imp and MTK sulphoxide impurities were well separated at RT 1.1 min, 4.6 min, 13.5 min, and 14.3 min respectively. The calibration curve of Bilastine, Montelukast, Bilastine DEO impurity and Montelukast sulphoxide impurities were found linear with regression coefficient more than 0.999. Accuracy was determined by recovery studies and found between 95-105%. The sample was found stable in aqueous solution over 23 hours. In all the forced degradation conditions, Bilastine and Montelukast were detected along with known and unknown impurities, which were well separated from each others with high resolution. The recorded observations were proved that the proposed analytical method is reliable for quantitation and impurity profiling of Bilastine and Montelukast from bulk mixture and tablet dosage form. [ABSTRACT FROM AUTHOR]

A novel high-performance thin-layer chromatographic (HPTLC) analytical method has been developed and optimized for the quantification of quetiapine fumarate (QF) and its two genotoxic impurities in drug substance and drug product. The desired separation was achieved on 60F254 pre-coated HPTLC plates using combination of green solvents, ethyl acetate‒ethanol‒n-heptane (5:1:4, V/V) as developing solvents. The detection wavelength used for quantification was 229 nm. QF and its two related genotoxic impurities, namely, 2-chloroaniline and 2-aminodiphenylsulfide, were well resolved from one another with retention factor values of 0.13 ± 0.02, 0.57 ± 0.02 and 0.76 ± 0.02, respectively. The optimized method was validated according to the guidelines laid down by the International Council for Harmonisation. The linearity was determined in the range of 100–600 ng/spot for QF and 10‒60 ng/spot for its two related genotoxic impurities; R2 ≥ 0.993. The method exhibited precision along with good accuracy, where 0.51, 0.86 and 1.86. The percentage recoveries obtained for 2-chloroaniline and 2-aminodiphenylsulfide were 99.04‒101.04%. The developed method can be successfully used for the analysis of drug samples. [ABSTRACT FROM AUTHOR]

[Display omitted] • An Advanced, environmentally friendly stability indicating analytical method for Pretomanid analysis integrating Quality by Design (QbD) and Green Chemistry Principles. • The regulatory focus on quality in drug development emphasised for the drug stability in different circumstances and PMD found highly sensitive towards hydrolysis, oxidation, and photolytic conditions. • The eco-friendly method developed for PMD degradation products in stress studies unveils vital insights into its stability profile, contributing to the development of robust pharmaceutical formulations. • The developed method's eco-friendliness is confirmed through complex GAPI and AGREE assessments, aligning with the increasing focus on green chemistry and highlighting its broader impact on environmentally conscious drug development. Regulatory agencies prioritize the safety and effectiveness of pharmaceuticals. The focus here is to highlight the crucial need for control and oversight of Pretomanid (PMD), a synthetic nitroimidazooxazine utilized in treating multidrug-resistant tuberculosis. This study aims to develop a simple, sensitive, LCMS compatible, and environmentally friendly HPLC Stability Indicating Analytical Method following ICH Q1A (R2) and Q3 guidelines. The approach involves the application of Quality by Design (QbD) principles and adherence to green chemistry practices. A comprehensive risk assessment was conducted using cause-and-effect diagram and risk assessment tool. The QbD approach includes screening and optimization phases. Plackett-Burman design was performed for screening selected 11 primary parameters. Central composite design was used for optimisation of selected critical variables based on critical method parameters (CMPs) and Critical method attributes (CMAs). The method was optimised using Contour plot, 3D plot, ANOVA, Desirability value and Design space. Superior component separation was achieved through an isocratic elution using Kromasil C18 (250 × 4.6 mm; 5 μm) column, mobile phase composition of (50:50; v/v) ACN: 10 mM Ammonium Acetate pH 4.5 adjusted with Glacial Acetic acid and a flow rate of 0.9 mL/min having runtime of 30 mins. The stress studies revealed that the drug is highly sensitive to hydrolysis, oxidation, and photolytic conditions. Experimentation results demonstrated that acetonitrile served as the most suitable diluent. Additionally, the correlation coefficient (r2) exceeded 0.996, while the RSD values (n = 6) varied from 0.76 % to 1.93 % across the LOQ–150 % range. Specificity studies revealed no interference between peaks from impurities and known active analytes. Through stress studies, major degradation products were identified. The method's eco-friendliness was assessed using ComplexGAPI and AGREE, confirming its environmentally friendly nature. [ABSTRACT FROM AUTHOR]

• Effects of ion-pairing agent and counter ion on peak capacity were investigated. • The system configuration was fine-tuned to enhance resolving power. • ACN in IP-RPLC provides better chromatographic resolution than MeOH. • Column comparison was conducted at the kinetic performance limit (1000 bar). • Sub-minute separation and high-resolution analysis with coupled columns are demonstrated. A generic performance comparison strategy has been developed to evaluate the impact of mobile-phase additives (ion-pairing agent / counter ion systems), distinct stationary phases on resulting resolving power, and MS detectability of oligonucleotides and their critical impurities in gradient IP-RPLC. Stationary-phase considerations included particle type (core-shell vs. fully porous particles), particle diameter, and pore size. Separations were carried out at 60°C to optimize mass transfer (C-term). The incorporation of an active column preheater mitigated thermal mismatches, leading to narrower peaks and overcoming peak splitting. Acetonitrile as organic modifier outweighed methanol in terms of peak-capacity generation and yielded a 30% lower back pressure. Performance screening experiments were conducted varying ion-pairing agents and counter ions, while adjusting gradient span achieved an equivalent effective retention window. Hexafluoromethylisopropanol yielded superior chromatographic resolution, whereas hexafluoroisopropanol yielded significantly higher MS detection sensitivity. The 1.7 µm core-shell particle columns with 100 Å pores provided maximum resolving power for small (15–35 mers) oligonucleotides. Sub-min analysis for 15–35 polyT ladders was achieved operating a 50 mm long column at the kinetic performance limits. High-resolution separations between a 21-mer modified RNA sequence oligonucleotides and its related (shortmer and phosphodiester) impurities and complementary strand were obtained using a coupled column set-up with a total length of 450 mm. [ABSTRACT FROM AUTHOR]

[Display omitted] • A triple-action cramp relief over-the-counter pharmaceutical formulation of paracetamol, caffeine and pyrilamine maleate. • Novel eco-friendly HPTLC-densitometry and HPLC-UV methods for mixture analysis and purity assessment. • Successful applicability in assaying and quality control of the market tablets. • Dissolution profiling of Midol® Complete caplets as an in-vivo efficiency indicator. • Affirming methods sustainability via implementing various greenness and whiteness assessment tools. Over the years, several single-pill combinations have been produced and utilized as an alternative option for treatment of the signs and symptoms of Primary dysmenorrhea. The presence of multiple drugs together poses a challenge for analysts regarding analysis, making it a significant hurdle to overcome. A combination of paracetamol (PAR), caffeine (CAF), and pyrilamine maleate (PYR) is commonly recommended as an over-the-counter triple-action treatment to alleviate mild to moderate premenstrual syndrome symptoms. This study presented two liquid chromatographic methods that simultaneously determine PAR, CAF, and PYR in Midol® Complete caplets along with three officially recognized impurities, namely, p-aminophenol, theophylline, and p-anisaldehyde. The initial approach employed high-performance thin-layer chromatography alongside a densitometric scanning technique. The experimental setup utilized silica gel HPTLC plates as the stationary phase, and ethyl acetate: aqueous ammonium hydroxide (10.0:0.1, v/v) mixture as the developing system. The densitometric scanning was conducted at a wavelength of 210.0 nm. The second approach, developing high-performance liquid chromatographic method coupled with ultraviolet detection. Inertsil ODS-3 C 18 column (250 × 4.6 mm, 5 μm) was used to separate the six compounds effectively. For the experimental procedure, an isocratic elution method was chosen with a mobile phase consisting of a mixture of 0.02 M potassium dihydrogen phosphate buffer acidified with o -phosphoric acid (pH 3.0), and methanol (80:20, v/v) at a flow rate of 1.0 mL/min. UV detection was carried out at a wavelength of 210.0 nm. The two proposed methods have been validated in accordance with ICH recommendations. In addition, the suggested HPLC-UV technique was used to track the release profile of the cited drugs during the in-vitro study. The developed methods were assessed and compared for their environmental impact using various greenness evaluation tools. Additionally, the recently introduced RGB12 model was used to study the whiteness features of the suggested methods. The proposed methods yielded desirable outcomes and are sustainable, simple, affordable, and low-cost, which highly recommends their use in quality control labs. [ABSTRACT FROM AUTHOR]

Dinotefuran (DNT) is a neonicotinoid insecticide widely used in pest control. Identification of structurally related impurities is indispensable during material purification and pesticide registration and certified reference material development, and therefore needs to be carefully characterized. In this study, a combined strategy with liquid chromatography high-resolution mass spectrometry and SIRIUS has been developed to elucidate impurities from DNT material. MS and MS/MS spectra were used to score the impurity candidates by isotope score and fragment tree in the computer assisted tool, SIRIUS. DNT, the main component, worked as an anchor for formula identification and impurity structure elucidation. With this strategy, two by-product impurities and one stereoisomer were identified. Their fragmentation pathways were concluded, and the mechanism for impurity formation was also proposed. This result showed a successful application for combined human intelligence and machine learning, in the identification of pesticide impurities.

In 2018, the discovery of carcinogenic nitrosamine process related impurities (PRIs) in a group of widely used drugs led to the recall and complete withdrawal of several medications that were consumed for a long time, unaware of the presence of these genotoxic PRIs. Since then, PRIs that arise during the manufacturing process of the active pharmaceutical ingredients (APIs), together with their degradation impurities, have gained the attention of analytical chemistry researchers. In 2020, favipiravir (FVR) was found to have an effective antiviral activity against the SARS-COVID-19 virus. Therefore, it was included in the COVID-19 treatment protocols and was consequently globally manufactured at large-scales during the pandemic. There is information indigence about FVR impurity profiling, and until now, no method has been reported for the simultaneous determination of FVR together with its PRIs. In this study, five advanced multi-level design models were developed and validated for the simultaneous determination of FVR and two PRIs, namely; (6-chloro-3-hydroxypyrazine-2-carboxamide) and (3,6-dichloro-pyrazine-2-carbonitrile). The five developed models were classical least square (CLS), principal component regression (PCR), partial least squares (PLS), genetic algorithm-partial least squares (GA-PLS), and artificial neural networks (ANN). Five concentration levels of each compound, chosen according to the linearity range of the target analytes, were used to construct a five-level, three-factor chemometric design, giving rise to twenty-five mixtures. The models resolved the strong spectral overlap in the UV-spectra of the FVR and its PRIs. The PCR and PLS models exhibited the best performances, while PLS proved the highest sensitivity relative to the other models.