Feed aggregator

Mitapivat is a novel, first-in-class, allosteric activator of pyruvate kinase enzyme. It has been approved by the US FDA in February 2022 for disease modifying treatment of haemolytic anaemia in adults. In the current study, the in vivo metabolites of mitapivat in the rat model were identified using quadrupole-time-of-flight mass spectrometry. A total 20 metabolites were identified, out of which nine metabolites were found to be novel and reported first time in the literature. The study also further refined the chemical structures of some of the reported metabolites. Oxidation, N-dealkylation, oxidation followed by dehydrogenation, and hydrolysis were the major Phase I metabolic pathways of mitapivat. The chief Phase II metabolism pathway was glucuronide conjugation of oxidised and amide hydrolysed metabolites of mitapivat.

Matrix-assisted laser desorption/ionization–imaging mass spectrometry (MALDI–IMS) is applied in drug discovery and development. A high-throughput quantitative MALDI–IMS methodology was developed to confirm whether epertinib is superior to lapatinib in penetrating brain metastases using intraventricular injection mouse models (IVMs) of human EGFR2 (HER2)-positive breast or T790M–EGFR-positive lung cancer cells. A simple calibration curve was prepared for each compound via spotting standard solutions without using blank tissue sections or blank tissues onto the same glass slide as the epertinib or lapatinib brain section samples. Quantitative MALDI–IMS was performed via coating a glass slide with a MALDI matrix solution containing each internal standard solution. The samples of calibration curve and brain section were analyzed using a linear ion trap mass spectrometer with a MALDI ion source. Epertinib and lapatinib responses were strongly linear, with a wide dynamic range and low variation (relative standard deviation [RSD] < 20%) among the individual concentrations. Epertinib and lapatinib were sufficiently extracted from brain sections after oral administration in a breast cancer IVM. The quantitative MALDI–IMS results revealed that the epertinib concentrations administered to the brain sections in the lung cancer IVM were similar to those measured using liquid chromatography–tandem mass spectrometry (LC–MS/MS). Quantitative MALDI–IMS, owing to its high reproducibility and throughput, is useful for selecting drug candidates in the early stages of discovery and development, enabling efficient and rapid screening of candidate compounds as well as an understanding of the mechanisms of drug efficacy, toxicity, and pharmacokinetics/pharmacodynamics.

Proximity labeling (PL) proteomics has emerged as a powerful tool to capture both stable and transient protein interactions and subcellular networks. Despite the wide biological applications, PL still faces technical challenges in robustness, reproducibility, specificity, and sensitivity. Here, we discuss major analytical challenges in PL proteomics and highlight how the field is advancing to address these challenges by refining study design, tackling interferences, overcoming variation, developing novel tools, and establishing more robust platforms. We also provide our perspectives on best practices and the need for more robust, scalable, and quantitative PL technologies.

In this study, the components of Moringa oleifera (Lam) seed were extracted using an ultrasonic-assisted extraction technique with 70% methanol solution as the solvent. The antioxidant properties of the extract were evaluated through its scavenging abilities against DPPH. and ABTS. The antiproliferative effects of this extract on breast cancer MDA-MB-231, colon cancer SW480, leukemia HL-60, liver cancer SMMC-7721, and lung cancer A549 were assessed using the MTS assay. The distribution of MA, D-trehalose, and rbGlu in the heart, kidney, liver, and spleen of mice in various intervals was visually investigated using MALDI-IMS. In particular, the metabolic pathways of rbGlu were further elucidated. The result shows that rbGlu is metabolized to rbot-Glu in mice within 1 h. This approach further substantiates the use of MALDI-MSI technique in situ for studying the pharmacological mechanisms of bioactive component in natural products.

The fragmentation pathways for amines dissolved in methanol (CH3OH) or deuterated methanol (CD3OD) have been investigated by high-resolution accurate mass gas chromatography mass spectrometry (HRAM-GCMS) or GC-Orbitrap. Primary and secondary amines used in this study were 1,3-dimethylamylamine (1,3-DMAA) and ephedrine hydrochloride (Eph), respectively. For isotopic labeling experiment, 1S, 2R (+) ephedrine-D3 hydrochloride (D3-Eph) was used. Under splitless injection mode at an inlet temperature of 250°C, formaldehyde and its deuterated form were generated from CH3OH and CD3OD, respectively. This was evidenced by the oxonium ions generated from each solvent. When 1,3-DMAA was dissolved in CH3OH or CD3OD, distinct separation between the unreacted amine and condensation product fragments was observed, specifically methylene-imine (M + 12) and deuteromethylene-imine (M + 14) artifacts. More complex condensation patterns for Eph and D3-Eph were observed, attributed to the labile hydrogen/deuterium exchange and gradual deuteration from CH3OH to CD3OD. The fragmentation pathways were supported by the presence of oxazolidine intermediates before forming smaller condensation product fragments. Despite their close retention time and mass, the HRAM data distinguished the isobaric unreacted amine and condensation product fragments produced by Eph and D3-Eph in the coeluting region.

No abstract is available for this article.

Adequate vitamin K is crucial for optimal health. Although vitamin K detection methods have been established using liquid chromatography–tandem mass spectrometry (LC–MS/MS), some limitations remain. Therefore, we aimed to establish a stable and rapid LC–MS/MS method that can quantify phylloquinone (VK1), menaquinones-4 (MK-4), and menaquinones-7 (MK-7) in serum and cerebrospinal fluid and explore its clinical applications. We developed an LC–MS/MS method with atmospheric pressure chemical ionization to quantify and validate its performance according to Clinical Laboratory and Standard Institution standards (C62-Ed2). Serums from 50 healthy individuals and cerebrospinal fluid from 15 patients were collected for clinical application. Sample preparation involved lipase incubation, protein precipitation with ethanol, and liquid–liquid extraction with hexane/ethyl; optimization was performed for sample preparation and LC separation. Linearity was 50–10 000 pg/mL for VK1, MK-4, and MK-7. The total coefficient of variation (%) for VK1, MK-4, and MK-7 ranged from 8.5% to 10.4%, 8.0% to 10.4%, and 7.0% to 11.1%, respectively. Recovery of VK1, MK-4, and MK-7 was 82.3%–110.6%, 92.3%–110.6%, and 89.5%–117.8%, respectively. VK1 and MK-7 were detected in the serum of all 50 healthy subjects, whereas MK-4 was detected in only 13 (26%) subjects. Approximately 53.3% (8/15) had no detectable vitamin K in their cerebrospinal fluid. The developed method exhibited satisfactory performance and was applicable for detecting VK1, MK-4, and MK-7 in serum and cerebrospinal fluid.

Electrospray ionization (ESI) is often the ionization method of choice, particularly for high-mass polar molecules and complexes. However, when analyzing mixtures of analytes, charge state ambiguities and overlap in mass-to-charge (m/z) can arise from species with different masses and charges. While solution-phase conditions can sometimes be optimized to produce relatively low charge states—thereby reducing charge-state ambiguity and m/z overlap—gas-phase methods offer greater control over charge state reduction. For complex mixtures, however, charge state reduction alone often fails to resolve individual components in the mixture. Incorporating a mass-selection step prior to charge state manipulation can simplify the mixture and significantly improve the separation of the components. This general tandem mass spectrometry approach is referred to here as precursor resolution via ion z-state manipulation (PRIZM). Examples of variations of PRIZM experiments date back roughly 25 years and have involved ion/molecule proton transfer reactions, ion/ion proton transfer reactions, ion/ion electron transfer reactions, electron capture reactions, and multiply-charged ion attachment reactions. This tutorial review describes the PRIZM approach and provides illustrative examples using each of the charge state manipulation approaches mentioned above.

In the present work, we have first applied the fast nonlinear damping identification (NDI) method to nondestructively and simultaneously determine the values of mass, size, and charge of biological micro-objects trapped in a quadrupole Paul trap. Here, we used well-studied Lycopodium clavatum individual spores as a test object. For the nondestructive determination of spore parameters, we analyzed their extended orbit trajectory implemented in the nonlinear regime of viscous friction while trapping in a quadrupole Paul trap. The article discusses the prospects of the method proposed for investigating a wide class of biological objects.

Fungal infections are caused by opportunistic pathogens that can be life threatening and have been growing in prevalence. Many clinically relevant pathogens have resistance to or are developing resistance to the commonly used antifungal treatments. Occidiofungin (OCF) is a unique cyclic lipoglycopeptide with a novel structure that includes noncanonical amino acid in its covalent structure. It exhibits broad spectrum antifungal activity and has activity against drug resistant Candida species. Occidiofungin is a fungicidal compound that has a novel mechanism of action in which it disrupts higher order actin structures. Currently, occidiofungin is being developed for use in treating vulvovaginal candidiasis (VVC) and recurrent vulvovaginal candidiasis (RVVC). This study describes the development and application of a bioanalytical method for the quantification of occidiofungin in rabbit plasma. Method development was performed to quantify occidiofungin in rabbit plasma after intravaginal administration of a hydrogel containing occidiofungin. The method was validated with a linear range of 30–15 000 ng/mL in rabbit plasma. Precision, accuracy, calibration curve linearity, and stability of drug in plasma were established in quality controls. Extract stability, matrix effects, and recovery of drug in the extract were also determined. This study supported a repeat dose toxicity study in rabbits to determine occidiofungin pharmacokinetics and toxicokinetics. The pharmacokinetic and toxicokinetic primarily showed plasma concentrations of occidiofungin below the limit of quantification (BLOQ), suggesting that OCF-B does not readily cross the vaginal epithelial membrane.

Four 2,2,6,6-tetramethylpiperidinyloxyl (TEMPO) derivatives with molecular weights of 339–1131 Da were synthesized to investigate their ionization behaviors in fast atom bombardment (FAB), electron ionization (EI), direct analysis in real time (DART), electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), atmospheric pressure photoionization (APPI), and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). These include 4-dodecanoylamino-2,2,6,6-tetramethyl-1-piperidinyloxy (1), 4-dodecylamino-2,2,6,6-tetramethyl-1-piperidinyloxy (2), N,N′-bis(2,2,6,6-tetramethyl-1-oxyl-4-piperidinyl)dodecanediamide (3), and bis-cholesterol TEMPO derivative 4. For TEMPO derivatives 1 and 2 (molecular weights: 353 Da and 339 Da), [M]+, [M+H]+•, and [M+2H]+ peaks were detected. TEMPO derivative 3, containing two TEMPO moieties, showed [M]+•, [M+H]+, [M+2H]+•, and [M+3H]+ peaks. EI was optimal for smaller derivatives, as molecular ion peaks were predominant. For larger analytes, molecular ion intensities weakened, favoring hydrogen adduct peaks. EI and DART-MS failed to detect bis-cholesterol TEMPO derivative 4. Under APCI and APPI-MS, N–O bond cleavage was observed. MALDI-MS detected only hydrogen adduct peaks. FAB, ESI, APCI, and APPI-MS detections varied with scan numbers, unlike EI, DART, and MALDI-MS.

No abstract is available for this article.

Our healthcare system provides reactive sick-care, treating patients after symptoms have appeared by prescription of generic and often suboptimal therapy. This strategy brings along high costs and high pressure which is not sustainable. Alternatively, P5 healthcare is proposed focusing on five key elements: prevention, personalization, prediction, participation, psychocognition, however, changes in current clinical care pathways are required, for which antithrombin deficiency is a prime example. Hereditary antithrombin deficiency (ATD) is a genetic disorder, for which screening is instigated after a thrombotic episode. Current diagnostic tests for ATD lack sensitivity and refinement to correctly classify patients, and generic treatments are prescribed. A molecular understanding of ATD through a molecular diagnostic test that analyzes all clinically relevant features of antithrombin is required. Here, clinically relevant molecular characteristics of antithrombin, the diversity of antithrombin (deficiency) in heath and disease, and the strengths and weaknesses of antithrombin tests are reviewed. A mass spectrometry test that molecularly characterizes a patients antithrombin proteoforms harbors the highest potential to improve the clinical pathway for ATD. Application of this MS-based test in a future enhanced clinical pathway will improve patient management and outcome through molecular characterization of antithrombin and enables the promise of P5 healthcare for ATD.

Liquid chromatography-mass spectrometry (LC-MS) has become an indispensable tool for elucidating molecular structures and quantifying diverse compounds within complex mixtures. Despite its versatility, it faces various challenges such as ion suppression, low sensitivity, analyte instability, and matrix effects, which are being overcome by different kinds of offline and online derivatization techniques to improve specificity and reduce potential interferences. In this context, considerable advancements have been made in reviewing and critically evaluating a wide range of developed methods and techniques; however, little attention has been given to post-column derivatization (PCD) in LC-MS. Therefore, this comprehensive review highlights state-of-the-art advancements in LC-MS with a specific focus on various types of chemical and physical PCD, and in-source derivatization. It also examines the latest instrumentation developments, highlights methods and influencing factors, and explores applications in food, proteomics, biology, pharmaceuticals, and environmental analysis from the past four decades. Besides, this review critically examines the role of PCD in LC-MS along with outlining its advantages and disadvantages. Furthermore, special emphasis is also made on prospects and insights for developing more versatile LC-PCD-MS techniques and in-source methodologies, to address ongoing challenges and aim to open new research avenues for analysts.

Gas chromatography-mass spectrometry (GC-MS) with Cold electron ionization (EI) is based on interfacing the GC and MS with a supersonic molecular beam (SMB) along with electron ionization of vibrationally cold sample compounds in the SMB in a contact-free fly-through ion source (hence the name Cold EI). Cold EI improves all the central performance aspects of GC-MS, including: a significantly extended range of compounds that are amenable for analysis, enhanced molecular ions, highly improved sample identification, faster analysis (much faster), uniform response to all analytes, greater selectivity and higher signal to noise ratios and lower limits of detection, particularly for compounds that are difficult to analyze. GC-MS with Cold EI executes the analysis of the full range of standard EI applications and most with major improvements of various metrics. Furthermore, it significantly extends the range of compounds and applications amenable for GC-MS analysis. Accordingly, it is a highly superior replacement ion source. In this review article, we describe Cold EI and its main features, discuss its benefits, and demonstrate several of its unique applications including cannabinoids analysis, synthetic organic compounds analysis, whole blood analysis for medical diagnostics, isomer distribution analysis for improved fuels and oils, and explosives analysis.

Diabetes mellitus (DM) is a prevalent metabolic disorder attributed to insulin secretion and action defects, affecting a growing adult population with hyperglycemia expected to reach 578 million by 2030. This study explores the antidiabetic potential of 25% aqueous methanol extract of Cupressus torulosa needles, utilizing in vitro and in vivo assays. In the α-glucosidase inhibition assay, the extract exhibited significant in vitro antidiabetic activity with an IC50, 123.45 ± 1.8 μg/mL, comparable to the standard drug acarbose (IC50, 58.21 ± 3.1 μg/mL). Toxicity assessment indicated non-toxic nature of the extract at 2000 mg/kg b.w. In STZ-induced diabetic mice, it displayed dose-dependent antihyperglycemic effects, evident at the fourth hour and 14th day, paralleling the positive control glibenclamide. In the chemical profiling of the extract using UPLC-QTOF-MS, the mobile phases consisted of 0.1% formic acid in water (Solvent A) and 100% methanol (Solvent B). The gradient elution started with 5% B (0 to 2 min) and gradually increased to 95% B by 25 min, followed by a post-run time of 2 min. Preliminarily, 50 constituents were identified, predominantly phenolics, with hypoglycemic effects attributed to flavonoids like (−)-epicatechin, amentoflavone, and cupressuflavone, as well as iridoid O-glycoside, exemplified by haprpagoside. Further studies are needed to assess the long-term efficacy, safety, and molecular mechanisms of the extract of C. torulosa needles in diabetes management.

Monosaccharides play a central role in metabolic networks and in the biosynthesis of glycomolecules, which perform essential functions across all domains of life. Thus, identifying and quantifying these building blocks is crucial in both research and industry. Routine methods have been established to facilitate the analysis of common monosaccharides. However, despite the presence of common metabolites, most organisms utilize distinct sets of monosaccharides and derivatives. These molecules therefore display a large diversity, potentially numbering in the hundreds or thousands, with many still unknown. This complexity presents significant challenges in the study of glycomolecules, particularly in microbes, including pathogens and those with the potential to serve as novel model organisms. This review discusses mass spectrometric techniques for the isomer-sensitive analysis of monosaccharides, their derivatives, and activated forms. Although mass spectrometry allows for untargeted analysis and sensitive detection in complex matrices, the presence of stereoisomers and extensive modifications necessitates the integration of advanced chromatographic, electrophoretic, ion mobility, or ion spectroscopic methods. Furthermore, stable-isotope incorporation studies are critical in elucidating biosynthetic routes in novel organisms.

Proteomics is nowadays increasingly becoming part of the routine clinical practice of diagnostic laboratories, especially due to the advent of advanced mass spectrometry techniques. This review focuses on the application of proteomic analysis in the identification of pathological conditions in a hospital setting, with a particular focus on the analysis of protein biomarkers. In particular, the main purpose of the review is to highlight the challenges associated with the identification of specific disease-causing proteins, given their complex nature and the variety of posttranslational modifications (PTMs) they can undergo. PTMs, such as phosphorylation and glycosylation, play critical roles in protein function but can also lead to diseases if dysregulated. Proteomics plays an important role especially in various medical fields ranging from cardiology, internal medicine to hemato-oncology emphasizing the interdisciplinary nature of this field. Traditional methods such as electrophoretic or immunochemical methods have been mainstay in protein detection; however, these techniques are limited in terms of specificity and sensitivity. Examples include the diagnosis of multiple myeloma and the detection of its specific protein or amyloidosis, which relies heavily on these conventional methods, which sometimes lead to false positives or inadequate disease monitoring. Mass spectrometry in this respect emerges as a superior alternative, providing high sensitivity and specificity in the detection and quantification of specific protein sequences. This technique is particularly beneficial for monitoring minimal residual disease (MRD) in the diagnosis of multiple myeloma where traditional methods fall short. Furthermore mass spectrometry can provide precise typing of amyloid proteins, which is crucial for the appropriate treatment of amyloidosis. This review summarizes the opportunities for proteomic determination using mass spectrometry between 2012 and 2024, highlighting the transformative potential of mass spectrometry in clinical proteomics and encouraging its wider use in diagnostic laboratories.

The development of LA-ICPMS and LA-MC-ICPMS resulted in analytical methods for zircon geochronology using a 20- to 60-μm laser spot size. A high amount of complexly zoned zircons promotes the requirement of U–Pb dating at smaller spot sizes. When spot size reduces, downhole fractionation (DHF) increases, increasing the DHF discrepancy between zircon grains and the primary reference zircon standard and resulting in inaccurate results. With MC-ICPMS's high sensitivity and multi-collector capabilities, this work attempts to accurately determine age with spatial resolutions below 20 μm. Three well-characterized zircon standards (91500, GJ-1, and Plešovice) were tested at spot sizes of 35, 20, 15, and 10 μm. Laser and mass spectrometry tuning, laser shot count, and ablation time have been optimized to reduce the DHF effect on measurement accuracy. Static ablation of 35- to 15-μm spots with 150 laser shot counts (30 s) yielded precision of less than 1.5% and age offset of less than 2%. The DHF differs significantly from the reference standard and two test zircon samples used for validation, with an age offset of 4% at 10-μm spot sizes. Masking shot counts from the end enhanced accuracy, notably for lower laser shot counts and shorter ablation times. At 75 laser shot counts, precision reached 1.4% and age offsets reduced to 1.6% for 206Pb/238U age. This method minimizes laser shot counts to avoid sampling two age groups. Higher zircon DHF variability at smaller spot sizes may influence a well-calibrated, sensitive LA-MC-ICP-MS analytical figure of merit.

MS analysis of cyclic peptides in negative ion mode has been a challenge, in particular if the peptide does not contain acidic functional groups. In this paper, we present a way to easily produce negative ions from anionic peptide adducts, utilising collision-induced dissociation (CID)-mediated elimination. Using two different mass spectrometers, we have generated series of adducts of three cyclic and one linear peptide with various anions. They were then isolated and subjected to CID with a range of collision energies. The deprotonation percentage was then calculated from the resultant spectrum, and compared between the spectrometers, as well as with an external reference—proton affinity values. The susceptibility to deprotonate by detaching a HX moiety is proportional to the proton affinity of the X− species. Also, the linear peptide deprotonated more readily than the cyclic ones. On the other hand, lack of amino or acidic groups resulted in higher collision voltage (CV) necessary for the formation of deprotonated species. Moreover, the exact propensity for neutral loss depends on the ion temperature, which differs between mass spectrometers. We have developed a facile method for generating peptide anions for MS analysis of cyclic peptides, which works even if the peptide in question does not have easily ionisable groups. The deprotonated species generated in this way can be fragmented again in order to identify the peptide.