In-situ formation and evaluation of N-nitrosamine drug substance related impurities in glycopeptides implying orbitrap mass spectrometry.

Nitrosamines, a class of N-nitroso compounds, have raised significant health concerns due to their established carcinogenicity. ICH M7 enlisted N-nitroso compounds in the so called cohorts of concern due to their carcinogenic effects. Glycopeptides (GPs) are complex molecules composed of peptide and glycan moieties.

Reactivity of N terminal histidine of peptides towards excipients/impurity of excipients: A case study of liraglutide excipient compatibility study.

The selection of quality excipients is a crucial step in peptide formulation development. Apart from excipient incompatibility, process-related impurities or degradants of an excipient can interact with peptide-active pharmaceutical ingredients, forming the interaction products. The formaldehyde has been reported as an impurity of excipient in polyethylene glycol, glycerol, magnesium stearate, microcrystalline cellulose, mannitol, etc.

Identification and characterization of GSK-9089 metabolites through high resolution-mass spectrometry based in vitro and in vivo rat biological sample analysis.

Estrogen related receptors (ERRs) agonist GSK-9089 (DY-131) reported to pose a potential in increasing exercise endurance. High resolution mass spectrometry (HRMS) based analysis has utmost importance in the detection, identification, or characterization of a molecule including its metabolites in human body. In this study, in vitro metabolism profile of GSK-9089 was investigated after incubation with liver microsomes and S9 fractions.

A comprehensive study on the identification and characterization of degradation products of lipoglycopeptide Dalbavancin using LC and LC-HRMS/MS.

Dalbavancin is the second-generation approved semisynthetic lipoglycopeptide by the United States Food and Drug Administration (USFDA) for the treatment of acute bacterial skin and skin-structure infections. Unlike other lipoglycopeptides, the stability behavior of Dalbavancin was least explored, which is a prerequisite. The current study endeavors to elucidate the oxidative and hydrolytic stability behavior of Dalbavancin by exposing the drug to oxidative, acidic, and basic stress conditions.

Bioanalysis of Stress Biomarkers through Sensitive HILIC-MS/MS Method: A Stride toward Accurate Quantification of MDA, ACR, and CTA.

Quantifying reactive aldehyde biomarkers, such as malondialdehyde, acrolein, and crotonaldehyde, is the most preferred approach to determine oxidative stress. However, reported analytical methods lack specificity for accurately quantifying these aldehydes as certain methodologies may produce false positive results due to harsh experimental conditions. Thus, in this research work, a novel HILIC-MS/MS method with endogenous histidine derivatization is developed, which proves to have higher specificity and reproducibility in quantifying these aldehydes from the biological matrix.

Cell-Engineered Recombinant α-Synuclein: A Gage R&R Validated Protocol.

α-Synuclein (α-Syn) misfolding and its presence in Lewy bodies are observed in almost all Parkinson's disease (PD) patients. Basic biomedical research would benefit from a quick, low-cost approach to purifying α-Syn and developing models for PD. Several research groups utilize PFF-based models, yet the production of α-Syn PFFs is inconsistent, resulting in nonconclusive findings.

A critical assessment on stability behaviour of Vorinostat using LC-MS-QTOF with H/D exchange and NMR.

Vorinostat is the first USFDA-approved HDAC inhibitor for the treatment of cutaneous t-cell lymphoma. Vorinostat was exposed to ICH-recommended hydrolytic (acid, base, and neutral), oxidative, thermal, and photolytic stress conditions to understand the degradation behaviour. A Stability indicating LC method was developed and validated for separating and identifying forced degradation products.

Drug-Excipient Compatibility Study Through a Novel Vial-in-Vial Experimental Setup: A Benchmark Study.

Drug-excipient compatibility study (DECS) is one of the critical steps during pre-formulation studies to select the appropriate excipient to obtain a stable formulation/dosage form. As such, there is no recommended guideline for DECS. Further, the previously reported studies and protocols followed by various pharmaceutical industries are very lengthy and laborious.

A rapid discriminative hydrogen-deuterium exchange and LC-HRMS/MS strategy for primary and higher order structural mapping of therapeutic proteins: a case study using filgrastim.

A vast number of therapeutic proteins are approved and available on the market. However, there are very limited analytical approaches available for the rapid determination of primary and higher-order structures which can be utilized for counterfeit identification. In the present study, filgrastim biosimilar products from different manufacturers were considered for developing discriminative orthogonal analytical techniques to determine structural variations.

Modified NAP test: A simple and Responsive Nitrosating Methodology for Risk Evaluation of NDSRIs.

N-Nitroso compounds have been listed as one of the cohorts of concern as per ICH M7. In recent years, the regulatory focus has shifted from common nitrosamines to nitroso-impurities of drug products. Thus, the detection and quantification of unacceptable levels of nitrosamine drug substance-related impurities are of great concern for analytical scientists during drug development.

Hyphenated liquid chromatography - diode array detection - charged aerosol detection - high resolution - multistage mass spectrometry with online hydrogen/deuterium exchange: One stop solution for pharmaceutical impurity profiling.

Hyphenation of different analytical techniques has always been advantageous in structural characterization as it saves time, money and resources. In the pharmaceutical sector, chromatography-based impurity profiling, including identification, characterization, and quantification in drug substances or finished products, is of utmost importance to comply with quality, patient safety and regulatory requirements. These impurities are monitored using LC-UV/DAD and identified and/or characterized using HRMS and MS/MS.

Cutting-edge strategies and critical advancements in characterization and quantification of metabolites concerning translational metabolomics.

Despite remarkable progress in drug discovery strategies, significant challenges are still remaining in translating new insights into clinical applications. Scientists are devising creative approaches to bridge the gap between scientific and translational research. Metabolomics is a unique field among other omics techniques for identifying novel metabolites and biomarkers.

Extrinsic hyaluronic acid induction differentially modulates intracellular glutamine metabolism in breast cancer stem cells.

The effect of low and high molecular weight hyaluronic acid on glutamine metabolism in luminal and basal breast cancer and cancer stem cells is being investigated. In luminal cell lines (MCF-7), HA enhances the intracellular utilization of gln in redox metabolism and decreases its use in TCA. On the contrary, in MDAMB-231 cells, HA induces the uptake of gln to be utilized in anaplerosis rather than ROS maintenance.

Synthetic pharmaceutical peptides characterization by chromatography principles and method development.

As per the United States Food and Drug Administration, any polymer/chain composed of 40 or fewer amino acids is called a peptide, where more than 40 amino acids are considered proteins. On many occasions, there is a change in the source of manufacturing of the peptide active pharmaceutical ingredient, where one has to prove the sameness of that product with the existing formulation by considering several aspects like the presence of impurities/degradation products, the extent of aggregations, and so forth. For the same, several chromatographic characterization techniques such as reversed-phase high-performance liquid chromatography-ultraviolet/high-resolution mass spectrometry, supercritical fluid chromatography, size-exclusion chromatography, ion-exchange chromatography, and so forth, are widely used in the pharmaceutical industry.

LC-HRMS studies on ruxolitinib degradation: a comprehensive approach during drug development.

Ruxolitinib, a kinase inhibitor, was subjected to stress studies as described in the ICH Q1A(R2) guidelines. Solution state hydrolytic and solid state oxidative and thermal stress studies were carried out to understand its degradation behaviour. The drug showed significant instability in the hydrolytic condition in comparison with other conditions.

A systematic UHPLC-Q-TOF-MS/MS based analytical approach for characterization of flibanserin metabolites and establishment of biotransformation pathway.

A systematic metabolite profiling approach has paramount importance in detecting, identifying, and characterizing drug metabolites. Till date, there is no report published on the comprehensive metabolic fate of flibanserin (FLB). In this study, the structure of entire potential metabolites of FLB has been elucidated by execution of in silico tool and high resolution mass spectrometry based metabolite profiling strategy employing data-dependent and data-independent approaches.

Cross-linking of poly (vinyl alcohol) films under acidic and thermal stress.

Poly (vinyl alcohol), PVA, a commonly used excipient to coat tablets, forms insoluble films in the presence of acids and thermal stress. This may lead to drug products failing to meet dissolution specifications over time. Studies were conducted to understand the effect of acid strength, processing conditions, and storage stress on the mechanism of insoluble film formation using PVA and Opadry II as model systems.

Amalgamation of stress degradation and metabolite profiling in rat urine and feces for characterization of oxidative metabolites of flibanserin using UHPLC-Q-TOF-MS/MS, H/D exchange and NMR technique.

Flibanserin (FLB) is the first FDA approved drug showed to have significant activity against sexual desire disorder of premenopausal and postmenopausal women. Unfortunately, FLB is used as an adulterant in dietary supplement products as a performance enhancer in sports. Identification of FLB and its metabolites in the biological samples requires an authenticated analytical technique.

Practical and economical implementation of online H/D exchange in LC-MS.

Structural elucidation is an integral part of drug discovery and development. In recent years, due to acceleration of the drug discovery and development process, there is a significant need for highly efficient methodologies for structural elucidation. In this work, we devised and standardized a simple and economical online hydrogen-deuterium exchange methodology, which can be used for structure elucidation purposes.

Critical practical aspects in the application of liquid chromatography-mass spectrometric studies for the characterization of impurities and degradation products.

Liquid chromatography-mass spectrometry (LC-MS) is considered today as a mainstay tool for the structure characterization of minor components like impurities (IMPs) and degradation products (DPs) in drug substances and products. A multi-step systematic strategy for the purpose involves high resolution mass and multi-stage mass studies on both the drug and IMPs/DPs, followed by comparison of their fragmentation profiles. Its successful application requires consideration of many practical aspects at each step.

LC-MS/TOF, LC-MSn, on-line H/D exchange and LC-NMR studies on rosuvastatin degradation and in silico determination of toxicity of its degradation products: a comprehensive approach during drug development.

The present study dealt with the forced degradation behaviour of rosuvastatin under ICH prescribed stress conditions. The drug was found to be labile under acid hydrolytic and photolytic conditions, while it was stable to base/neutral hydrolytic, oxidative and thermal stress. In total, 11 degradation products were formed, which were separated on a C-18 column using a stability-indicating method.

A critical review on the use of modern sophisticated hyphenated tools in the characterization of impurities and degradation products.

With ever increasing regulatory and compendial stringency on the control of impurities (IMPs) and degradation products (DPs) (including genotoxic impurities) in drug substances and finished pharmaceutical formulations, a profound emphasis is being paid on their characterization and analysis at trace levels. Fortunately, there have been parallel tremendous advancements in the instrumental techniques that allow rapid characterization of IMPs and/or DPs at the prescribed levels of ∼0.1%.

ICH guidance in practice: degradation behaviour of oseltamivir phosphate under stress conditions.

Oseltamivir phosphate was subjected to stress degradation conditions prescribed by ICH guideline Q1A (R2). A total of five degradation products (Os I to Os V) were generated under hydrolytic (acid and alkaline) stress conditions. Their unambiguous structural elucidation was carried out using LC-MS, LC-NMR and HR-NMR data.