Mass Spectrometric Techniques Research Articles

Composition and Sources of Organic Aerosol in Two Megacities in Western China Using Complementary Mass Spectrometric and Statistical Techniques

Composition and Sources of Organic Aerosol in Two Megacities in Western China Using Complementary Mass Spectrometric and Statistical Techniques

Obscured Contribution of Oxygenated Intermediate-Volatility Organic Compounds to Secondary Organic Aerosol Formation from Gasoline Vehicle Emissions.

Secondary organic aerosol (SOA) formation from gasoline vehicles spanning a wide range of emission types was investigated using an oxidation flow reactor (OFR) by conducting chassis dynamometer tests. Aided by advanced mass spectrometric techniques, SOA precursors, including volatile organic compounds (VOCs) and intermediate/semivolatile organic compounds (I/SVOCs), were comprehensively characterized. The reconstructed SOA produced from the speciated VOCs and I/SVOCs can explain 69% of the SOA measured downstream of an OFR upon 0.5-3 days' OH exposure. While VOCs can only explain 10% of total SOA production, the contribution from I/SVOCs is 59%, with oxygenated I/SVOCs (O-I/SVOCs) taking up 20% of that contribution. O-I/SVOCs (e.g., benzylic or aliphatic aldehydes and ketones), as an obscured source, account for 16% of total nonmethane organic gas (NMOG) emission. More importantly, with the improvement in emission standards, the NMOG is effectively mitigated by 35% from China 4 to China 6, which is predominantly attributed to the decrease of VOCs. Real-time measurements of different NMOG components as well as SOA production further reveal that the current emission control measures, such as advances in engine and three-way catalytic converter (TWC) techniques, are effective in reducing the "light" SOA precursors (i.e., single-ring aromatics) but not for the I/SVOC emissions. Our results also highlight greater effects of O-I/SVOCs to SOA formation than previously observed and the urgent need for further investigation into their origins, i.e., incomplete combustion, lubricating oil, etc., which requires improvements in real-time molecular-level characterization of I/SVOC molecules and in turn will benefit the future design of control measures.

Marine Bacillus haynesii chitinase: Purification, characterization and antifungal potential for sustainable chitin bioconversion

The development of chitinase tailored for the bioconversion of chitin to chitin oligosaccharides has attracted significant attention due to its potential to alleviate environmental pollution associated with chemical conversion processes. In this present investigation, we purified extracellular chitinase derived from marine Bacillus haynesii to homogeneity and subsequently characterized it. The molecular weight of BhChi was approximately 35 kDa. BhChi displayed its peak catalytic activity at pH 6.0, with an optimal temperature of 37 °C. It exhibited stability across a pH range of 6.0–9.0. In addition, BhChi showed activation in the presence of Mn2+ with the improved activity of 105 U mL−1. Ca2+ and Fe2+ metal ions did not have any significant impact on enzyme activity. Under the optimized enzymatic conditions, there was a notable enhancement in catalytic activity on colloidal chitin with Km of 0.01 mg mL−1 and Vmax of 5.75 mmol min−1. Kcat and catalytic efficiency were measured at 1.91 s−1 and 191 mL mg−1 s−1, respectively. The product profiling of BhChi using thin layer chromatography and Mass spectrometric techniques hinted an exochitinase mode of action with chitobiose and N-Acetyl glucosamine as the products. This study represents the first report on an exochitinase from Bacillus haynesii. Furthermore, the chitinase showcased promising antifungal properties against key pathogens, Fusarium oxysporum and Penicillium chrysogenum, reinforcing its potential as a potent biocontrol agent.

Pyrene based AIE-active probe for selective detection of picric acid through the inner filter effect channel in aqueous medium

The development of pyrene derivatives possessing the AIE feature have emerged as an intriguing area of research with potential applications in several domains. In the present study, we designed and developed an AIE active pyrene derivative for picric acid sensing in an aqueous medium. The design based on a single step condensation reaction and the product has been meticulously characterized by using NMR and Mass spectrometric techniques. In comparison to pristine pyrene, the synthesized pyrene derivative has a lower bandgap energy as a result of the intramolecular charge transfer between pyrene and 4-(methylthio)-2-oxo-2H-pyran-3-carbonitrile. The synthesized probe demonstrates a moderate level of fluorescence when dissolved in a solution, and it has exhibited a remarkable emission caused by aggregation at higher water percentages. The aggregates were meticulously characterized by DLS, SEM and TEM measurements. The spherically shaped aggregates have shown enhanced fluorescence quantum yield and lifetime, rendering them a more suitable option for sensor applications. The aggregates were effectively employed as a fluorescent probe for the detection of picric acid in an aqueous medium. The probe exhibited a notable level of selectivity and sensitivity towards picric acid, as evidenced by its low detection limit of 4.47 nM. Moreover, the quenching mechanism was ascribed to the inner filter effect, providing supporting evidence from both experimental and mathematical calculations. The findings indicate that the probe has potential as a feasible choice for a picric acid sensor.

Improving the therapeutic window of anticancer agents by β-cyclodextrin encapsulation: Experimental and theoretical insights

3-Nitro-2H-chromene derivatives exhibit promising anticancer activities, but their clinical translation is hindered by poor aqueous solubility. Herein, we report a cyclodextrin complexation strategy to improve the solubility and therapeutic efficacy of 8-methoxy-2-(4-methoxy phenyl)-3-nitro-2H-chromene (MNC), a potent anticancer agent. Solid inclusion complexes of MNC: β-cyclodextrin (β-CD) were synthesized by co-precipitation and kneading methods, and comprehensively characterized using UV–vis, Fluorescence, Fourier Transform Infrared (FT-IR), Differential scanning calorimetry (DSC), X-ray diffraction (XRD), Scanning electron microscope (SEM), 1H NMR, and mass spectrometric techniques. A 1:1 host–guest stoichiometry was verified by phase solubility and UV–Vis spectroscopic studies. β-CD complexation remarkably enhanced the aqueous solubility of hydrophobic MNC. Cytotoxicity evaluation against different human cancer cell lines such as breast cancer (MCF-7, MDA-MB-231), lung cancer (A549), and liver cancer (HepG2) demonstrated superior anticancer activity of the co-precipitated MNC: β-CD complex compared to free MNC and the chemotherapeutic 5-fluorouracil. Although the host–guest interaction energy obtained from molecular docking was modest (−5.1 kcal/mol), it facilitated effective MNC encapsulation and delivery by β-CD. Density functional theory (DFT) calculations provided further insights into the inclusion phenomenon. This cyclodextrin complexation approach paves the way for developing an effective MNC formulation with improved bioavailability and anticancer therapeutic efficacy.

GPR39-mediated ERK1/2 signaling reduces permethrin-induced proliferation of estrogen receptor α-negative cells

Certain insecticides are known to have estrogenic effects by activating estrogen receptors through genomic transcription. This has led researchers to associate specific insecticide use with an increased breast cancer risk. However, it is unclear if estrogen receptor-dependent pathways are the only way in which these compounds induce carcinogenic effects. The objective of this study was to determine the impact of the pyrethroid insecticide permethrin on the growth of estrogen receptor negative breast cancer cells MDA-MB-231. Using tandem mass spectrometric techniques, the effect of permethrin on cellular protein expression was investigated, and gene ontology and pathway function enrichment analyses were performed on the deregulated proteins. Finally, molecular docking simulations of permethrin with the candidate target protein was performed and the functionality of the protein was confirmed through gene knockdown experiments. Our findings demonstrate that exposure to 10–40 μM permethrin for 48 h enhanced cell proliferation and cell cycle progression in MDA-MB-231. We observed deregulated expression in 83 upregulated proteins and 34 downregulated proteins due to permethrin exposure. These deregulated proteins are primarily linked to transmembrane signaling and chemical carcinogenesis. Molecular docking simulations revealed that the overexpressed transmembrane signaling protein, G protein-coupled receptor 39 (GPR39), has the potential to bind to permethrin. Knockdown of GPR39 partially impeded permethrin-induced cellular proliferation and altered the expression of proliferation marker protein PCNA and cell cycle-associated protein cyclin D1 via the ERK1/2 signaling pathway. These findings offer novel evidence for permethrin as an environmental breast cancer risk factor, displaying its potential to impact breast cancer cell proliferation via an estrogen receptor-independent pathway.

Phytochemical screening, gas chromatograph/mass spectrometer (GCMS) analysis and molecular toxicological potential of Hunteria umbellata aqueous fruit extract against Staphylococcus aureus in accessory gene regulators (AGRs)

BackgroundFruits of Hunteria umbellata (HU) have been utilized in folk medicine as potent against Staphylococcus aureus (SA) infections, particularly skin and nasal related conditions. However, there is scarcity of literature concerning toxicological evaluation of graded doses of HU fruit, especially at molecular level, specifically targeting the accessory gene regulator (AGR) system to prevent abuse of doses in the treatment of bacterial infections. This research evaluated molecular toxicological property of SA exposed to varied concentrations of aqueous HU fruit extract ranging from 500 to 3.90625 µg/µL using broth microdilution method and quantification of AGR I and II genes’ expression employing two-step reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). We first identified quality and quantity of chemical compounds in aqueous fruit of HU using phytochemistry and hybrid gas chromatograph–mass spectrometer (GC–MS) technique. Additionally, preliminary bactericidal potential of HU was assessed before molecular toxicology.ResultsResults revealed six phytochemicals and twenty analytical grade compounds from a standard library were identified from chromatograms of HU fruit extract. Some main compounds detected are n-Hexadecanoic acid (25.24%), 2-Pentadecanone, 6,10,14-trimethyl (16.08%), Cuparene (16.63%), Tetradecanoic acid (6.21%) and 9-Octadecenoic acid, (E)- (5.70%). Bactericidal activity shows significant (p < 0.05) toxicity in the tested (8) concentrations of HU fruits in a dose-response relationship compared to the controls (positive and negative). The quantified expression of AGR I and II genes in SA was most significantly increased (p < 0.05) at both 250 and 500 µg/µL of HU fruit extract while least significant increase (p < 0.05) was recorded at 125 µg/µL compared to control.ConclusionsNotably, the study highlighted a potential risk of augmented bacterial infection especially with higher doses of HU extracts during boils’ treatment and other epidermal infections instigated by Staph. Expression of both AGR genes at higher doses (250 and 500 µg/µL) is indicative of further expression of several other genes responsible for virulence factors in a variety of human bacterial infections. Thus, consumption of HU fruit to treat boils and blisters should be with great caution especially at high doses.

Assessment of energy consumption and environmental safety measures in a plasma pyrolysis plant for eco-friendly waste treatment

A plasma pyrolysis plant with a 50 kg/h installed capacity is investigated for its performance in terms of energy consumption, carbon emission, and environmental safety. Municipal Solid Waste (MSW), precisely the combustible fraction, is processed at approximately 1000 °C. A temperature shoot study is conducted to reduce the energy required for waste processing, revealing a total energy consumption of 0.586 kWh per kg of waste, leading to a reduction of ∼15%. A mass and energy balance has been done to get a clear understanding of different products, by-product generation, and heat losses during waste treatment. The gas analysis identified CO and H2 as dominant components, comprising around 30% of the produced gases, with a lower heating value (LHV) of 4.7 MJ/Nm3. The cold gas efficiency ranges from 44 to 53%. The Energy Dispersive X-ray (EDX) and Inductively Coupled Plasma Mass Spectrometer (ICP-MS) techniques characterize the ash generated during the process and are found carbonless and non-toxic in nature. The wastewater produced during gas cleaning exhibited a neutral acidic nature, with total dissolved solids (TDS) ranging from 1200 to 1500 mg/l. These findings contribute to the plant's promising role in sustainable waste management practices, highlighting its potential as an environmentally sound solution for processing waste.

Joint forces of mass spectrometric techniques (ICP-MS and MALDI-TOF-MS) and fluorescence spectrometry in the study of platinum-based cytostatic drugs interactions with metallothionein MT2 and MT3

Herby, the interaction of metallothioneins with commonly used Pt-based anticancer drugs – cisplatin, carboplatin, and oxaliplatin - was investigated using the combined power of elemental (i.e. LA-ICP-MS, CE-ICP-MS) and molecular (i.e. MALDI-TOF-MS) analytical techniques providing not only required information about the interaction, but also the benefit of low sample consumption. The amount of Cd and Pt incorporated within the protein was determined for protein monomers and dimer/oligomers formed by non-oxidative dimerization. Moreover, fluorescence spectrometry using Zn2+-selective fluorescent indicator – FluoZin3 - was employed to monitor the ability of Pt drugs to release natively occurring Zn from the protein molecule. The investigation was carried out using two protein isoforms (i.e. MT2, MT3), and significant differences in behaviour of these two isoforms were observed.The main attention was paid to elucidating whether the protein dimerization/oligomerization may be the reason for the potential failure of the anticancer therapy based on these drugs. Based on the results, it was demonstrated that the interaction of MT2 (both monomers and dimers) interacted with Pt drugs significantly less compared to MT3 (both monomers and dimers). Also, a significant difference between monomeric and dimeric forms (both MT2 and MT3) was not observed. This may suggest that dimer formation is not the key factor leading to the inactivation of Pt drugs.

High-resolution mass spectrometric elucidation of electron ionization induced fragmentation pathways of methylated warfarin and selected hydroxylated species

The plant secondary metabolite families of coumarin and 4-hydroxy coumarin have a broad pharmacological spectrum ranging from antibacterial to anticancer properties. One prominent member of this substance class is the synthetic but naturally inspired anticoagulant drug and rodenticide warfarin (coumadin). A vast number of publications focus on the identification of warfarin and its major cytochrome P450-mediated phase I metabolites by liquid chromatography (LC) with mass spectrometry (MS) and tandem mass spectrometric (MS/MS) detection techniques. For the first time, electron ionization (EI) induced high-resolution quadrupole time-of-flight mass spectrometric (HR-qToF-MS) data of in-liner derivatized warfarin and selected hydroxylated species is provided in this study as an alternative to LC-MS/MS approaches. Furthermore, the characteristic fragments and fragmentation pathways of the analyzed methyl ethers are concluded. The obtained data of analytical standards, specific deuterated and 13C-labeled compounds prove inductive cleavage of the acyl or acetonyl side chain, methyl migration, and H-migration, along with consequential inductive cleavage as predominant fragmentation routes. Based on the HR-spectral data, commonalities and differences between the analyzed compounds and fragment groups were evaluated with future applicability in structure elucidation and spectra prediction of related compounds.

Pyrroloquinolones B–F: Five unusual alkaloids from Vernonia glabra (Steetz) Vatke (Asteraceae)

Five unusual alkaloids featuring a pyrrolo[1,2-a]quinolone skeleton (pyrroloquinolones B–F, 1–5) were isolated from the ethanol extract of the whole plant of Vernonia glabra (Steetz) Vatke, along with sixteen known compounds. Their structures were established by means of spectroscopic (1D and 2D NMR, UV, IR, and ECD) and high resolution mass spectrometric techniques as well as by comparison of their spectroscopic data with those reported in the literature. The ethanol extract and some isolated compounds were assessed for their antibacterial activity against four bacterial strains. The extract was significantly active against Staphylococcus aureus ATCC1026 and S. epidermidis ATCC35984 (MIC = 64 μg/mL). All the tested compounds showed moderate activity against S. epidermidis (16 ≤ MIC ≤ 64 μg/mL). Furthermore, this is the first report on tricyclic pyrrolo[1,2-a]quinolone alkaloids from a plant source. A biosynthetic pathway for the formation of these compounds is also proposed.

Neutralization of Anthropogenic Acidic Particles by NH3 From Wildfire Over Tropical Peatland

AbstractAcidity is an essential characteristic of aerosol particles that influences chemical and physical properties. Aerosol particles in tropical Asia, such as Singapore, have been reported as highly acidic due to intense anthropogenic sulfur emissions. At the same time, the region has also been experiencing wildfire over tropical peatlands, which is recognized as an intense source of NH3. Here, we investigated the role of NH3 from wildfire on aerosol particles in Singapore by employing the aerosol mass spectrometric technique. The observation was conducted both during wildfire haze (2015 October and 2019 September–October) and non‐haze (2018 October and 2019 April) periods. The observation result demonstrated that inorganic ionic species in Singapore were neutralized by NH4+ during the haze periods. Namely, the degree of neutralization of aerosol particles (i.e., measured NH4+ concentration/predicted NH4+ concentration by assuming that NH4+ fully neutralized SO42−, NO3−, and Cl−) was lower than 0.77during the non‐haze periods. On the other hand, the corresponding values were higher than 0.93 during the haze periods. In addition, NO3− concentration during the daytime of the haze period in 2015 was higher than that in other observation periods. A thermodynamic model calculation suggested that the regime shifts from the “NH3 sensitive region” to the “NH3 and HNO3 sensitive region” or “HNO3 sensitive region” might have occurred during the haze period. In the future, continuous monitoring of both gas‐ and particle‐phase inorganic chemical species will need to be conducted to investigate the impact of wildfire haze on atmospheric chemical processes in more detail.

Rapid and Bifunctional Chemoselective Metabolome Analysis of Liver Disease Plasma Using the Reagent 4‐Nitrophenyl‐2H‐azirine

AbstractPrimary sclerosing cholangitis (PSC) is a chronic inflammatory disease of the bile ducts that has been associated with diverse metabolic carboxylic acids. Mass spectrometric techniques are the method of choice for their analysis. However, the broad investigation of this metabolite class remains challenging. Derivatization of carboxylic acids represents a strategy to overcome these limitations but available methods suffer from diverse analytical challenges. Herein, we have designed a novel strategy introducing 4‐nitrophenyl‐2H‐azirine as a new chemoselective moiety for the first time for carboxylic acid metabolites. This moiety was selected as it rapidly forms a stable amide bond and also generates a new ketone, which can be analyzed by our recently developed quant‐SCHEMA method specific for carbonyl metabolites. Optimization of this new method revealed a high reproducibility and robustness, which was utilized to validate 102 metabolic carboxylic acids using authentic synthetic standard conjugates in human plasma samples including nine metabolites that were newly detected. Using this sequential analysis of the carbonyl‐ and carboxylic acid‐metabolomes revealed alterations of the ketogenesis pathway, which demonstrates the vast benefit of our unique methodology. We anticipate that the developed azirine moiety with rapid functional group transformation will find broad application in diverse chemical biology research fields.

Synthesis, Antimicrobial Activity and Molecular Docking of Some Novel 2-Aryl-4H-[1,3]-thiazolo[4,5-b]indoles

In present study, six novel 2-aryl-4H-[1,3]-thiazolo[4,5-b]indoles (4a-f) were synthesized by integrating thiazole and indole motifs through a three-component one-pot condensation involving isatin, ammonium thiocyanate and arylaldehydes. Structural confirmation was achieved using IR, 1H NMR and mass spectrometric techniques. Molecular targets, bioactivity scores, drug-likeness, anti- tuberculosis and antibacterial properties and toxicity were all predicted using various computational techniques. To calculate binding affinities to Mycobacterium tuberculosis enoyl-ACP reductase and Escherichia coli Topoisomerase IV, in vitro bioactivity studies were performed and Schrödinger docking simulations were performed. Anti-tubercular efficacy was evaluated using the MABA method, while antibacterial effectiveness against both Gram-positive and Gram-negative bacteria was assessed through the agar plate method. The results highlight the potential of these heterocyclic molecular hybrids, positioning them as promising candidates for further lead optimization in the development of more potent and safer pharmaceutical agents.

Rapid and Bifunctional Chemoselective Metabolome Analysis of Liver Disease Plasma Using the Reagent 4-Nitrophenyl-2H-azirine.

Primary sclerosing cholangitis (PSC) is a chronic inflammatory disease of the bile ducts that has been associated with diverse metabolic carboxylic acids. Mass spectrometric techniques are the method of choice for their analysis. However, the broad investigation of this metabolite class remains challenging. Derivatization of carboxylic acids represents a strategy to overcome these limitations but available methods suffer from diverse analytical challenges. Herein, we have designed a novel strategy introducing 4-nitrophenyl-2H-azirine as a new chemoselective moiety for the first time for carboxylic acid metabolites. This moiety was selected as it rapidly forms a stable amide bond and also generates a new ketone, which can be analyzed by our recently developed quant-SCHEMA method specific for carbonyl metabolites. Optimization of this new method revealed a high reproducibility and robustness, which was utilized to validate 102 metabolic carboxylic acids using authentic synthetic standard conjugates in human plasma samples including nine metabolites that were newly detected. Using this sequential analysis of the carbonyl- and carboxylic acid-metabolomes revealed alterations of the ketogenesis pathway, which demonstrates the vast benefit of our unique methodology. We anticipate that the developed azirine moiety with rapid functional group transformation will find broad application in diverse chemical biology research fields.

Exploring the Potential of Bioactive Peptides: From Natural Sources to Therapeutics.

Bioactive peptides, specific protein fragments with positive health effects, are gaining traction in drug development for advantages like enhanced penetration, low toxicity, and rapid clearance. This comprehensive review navigates the intricate landscape of peptide science, covering discovery to functional characterization. Beginning with a peptidomic exploration of natural sources, the review emphasizes the search for novel peptides. Extraction approaches, including enzymatic hydrolysis, microbial fermentation, and specialized methods for disulfide-linked peptides, are extensively covered. Mass spectrometric analysis techniques for data acquisition and identification, such as liquid chromatography, capillary electrophoresis, untargeted peptide analysis, and bioinformatics, are thoroughly outlined. The exploration of peptide bioactivity incorporates various methodologies, from in vitro assays to in silico techniques, including advanced approaches like phage display and cell-based assays. The review also discusses the structure-activity relationship in the context of antimicrobial peptides (AMPs), ACE-inhibitory peptides (ACEs), and antioxidative peptides (AOPs). Concluding with key findings and future research directions, this interdisciplinary review serves as a comprehensive reference, offering a holistic understanding of peptides and their potential therapeutic applications.

AN LC-ESI-MS/MS METHOD DEVELOPMENT AND VALIDATION FOR THE QUANTIFICATION OF INFIGRATINIB IN BIOLOGICAL MATRICES

Objective: The study was aimed to develop a precise and simple liquid chromatographic electrospray ionization tandem mass spectrometric (LC-ESI-MSMS) technique is essential for the quantification of Infigratinib in biological matrices. Methods: Chromatographic resolution was attained with PhenominexC18 (50 mm×2.6 mm, 3 µm) stationary column and a mobile solvent composition of 0.1% HCOOH, methyl alcohol and acetonitrile in the proportion of 10:10:80. Chromatograms were resolved by an isocratic separation with a flowing rate of 0.50 ml/min at 40 °C. Results: Quantitation was executed by monitoring the transitions of m/z. 560.19/189.13 for Infigratinib and 494.5→394.5 for Imatinib internal standard in multiple reaction monitoring. The standard curve regression line was y = 0.0016x+0.0062 and the correction coefficient (r2) was 0.9994. The % CV outcomes for matrix effect at Lower-QC and Higher-QC were 4.95% and 3.61% respectively. The percentage average recoveries for Infigratinib in Higher-QC (900ng/ml), MQC (600ng/ml) and Lower-QC (3ng/ml) were 93.27%, 94.69% and 97.24% respectively. The intra and interday precisions of analytical procedure was estimated by assessing the %CV outcomes and were in between 1.88 to 5.93% for the QC samples. Conclusion: The developed procedure can be useful for the assessment of Infigratinib in biological matrices in quality control, forensic and bioavailability studies.

New triazole based oxadiazolo/thiadiazolo–phthalazines as potent antimycobacterial agents: Design, synthesis, molecular modelling and in silico ADMET profiles

AbstractThe study of the chemical and biological properties of molecules simultaneously comprising various heterocycles, such as fused 1,3,4–oxadiazolyl–1,2,3–triazoles and 1,2,4–triazolyl–1,3,4–thiadiazole have been conducted as part of our ongoing research in the field of medicinal and organic chemistry. In this study, we performed ligand–based pharmacophore modelling as a promising design strategy of substituted phthalazinone derivatives (5 and 7). These were synthesized from key intermediate mercapto‐containing oxadiazo‐phthalazinone derivative 2. The synthesized compounds were characterized by IR, 1H and 13C NMR, elemental analysis and mass spectrometric techniques. In preliminary antibacterial studies, analogues 5 d, 5 f, 7 a and 7 d were found to be most effective against S. epidermidis, whereas 5 d displayed excellent activity against P. aeruginosa microorganism. Later, explored the probable key active site and inhibitors of Cytochrome P450 CYP121A1 interactions from M. tuberculosis H37Rv based antitubercular drug candidate with 5O4K protein and ligand 7 b possesses highest hydrophobic amino acid residues. The physicochemical and medicinal properties were also evaluated using SwissADME and ADMETlab2.0 web to function as effective oral bioavailability medications.

Application of advanced high resolution mass spectrometric techniques for the analysis of losartan potassium regarding known and unknown impurities

Recalls of medicinal products can cause supply bottlenecks. This is often due to the findings of unexpected impurities that pose a health risk to patients. A recent example is losartan potassium which was contaminated with azido-impurities. The choice of the analytical method determines which substances can be detected and thus controlled. In this study a combination of an untargeted screening approach for impurities and a targeted evaluation of high-resolution mass spectrometry data was applied to search for impurities not described so far, leaving out a precise quantification. Six losartan potassium samples were studied regarding known and unknown impurities and hence highlight the applicability and capability of the approach. For 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 an electrospray ionization in positive and negative mode for mass spectrometric detection was used. An information-dependent acquisition method was applied for the measurement of losartan potassium samples. The untargeted data evaluation using general unknown comparative screening revealed the presence of N-methyl-2-pyrrolidone (NMP) and another impurity from synthesis. The identity of NMP was corroborated by a spiking experiment and the amount was estimated by means of standard addition. A targeted data evaluation by generating extracted ion chromatograms resulted in finding of four additional impurities. Combined approaches like this are needed to detect and respond to changes in the quality of drugs precociously.

Collision induced unfolding and molecular dynamics simulations of norovirus capsid dimers reveal strain-specific stability profiles.

Collision induced unfolding (CIU) is a method used with ion mobility mass spectrometry to examine protein structures and their stability. Such experiments yield information about higher order protein structures, yet are unable to provide details about the underlying processes. That information can however be provided using molecular dynamics simulations. Here, we investigate the gas-phase unfolding of norovirus capsid dimers from the Norwalk and Kawasaki strains by employing molecular dynamics simulations over a range of temperatures, representing different levels of activation, together with CIU experiments. The dimers have highly similar structures, but their CIU reveals different stability that can be explained by the different dynamics that arises in response to the activation seen in the simulations, including a part of the sequence with previously observed strain-specific dynamics in solution. Our findings show how similar protein variants can be examined using mass spectrometric techniques in conjunction with atomistic molecular dynamics simulations to reveal differences in stability as well as differences in how and where unfolding takes place upon activation.