Search

Modern mass spectrometry technology allows for extensive sequencing of the ~ 25 kDa subunits of monoclonal antibodies (mAbs) produced by IdeS proteolysis followed by disulfide bond reduction, an approach known as middle-down mass spectrometry (MD MS). However, the spectral congestion of tandem mass spectra of large polypeptides dramatically complicates fragment ion assignment. Here, we report the development and benchmark of an MD MS strategy based on the combination of different ion fragmentation techniques with proton transfer charge reduction (PTCR) to simplify the gas-phase sequencing of mAb subunits. Applied on the liquid chromatography time scale using an Orbitrap Tribrid mass spectrometer, PTCR produces easy-to-interpret mass spectra with limited ion signal overlap. We demonstrate that the accurate estimation of the number of charges submitted to the Orbitrap mass analyzer after PTCR allows for the detection of charge-reduced product ions over a wide mass-over-charge (m/z) window with low parts per million m/z accuracy. Therefore, PTCR-based MD MS analysis increases not only sequence coverage, number of uniquely identified fragments, and number of assigned complementary ion pairs, but also the general confidence in the assignment of subunit fragments. This data acquisition method can be readily applied to any class of mAbs without an apparent need for optimization, and benefits from the high resolving power of the Orbitrap mass analyzer to return sequence coverage of individual subunits exceeding 80% in a single run, and > 90% when just two experiments are combined. [ABSTRACT FROM AUTHOR]

The consequences of climate change along with diverse food regulations and agricultural practices worldwide are complexifying the occurrence and management of chemical contaminants in food. In this context, we present an ultra-high-performance liquid chromatography high-resolution mass spectrometry (LC-HRMS) approach for the simultaneous identification and quantitation of over 1100 pesticide residues, mycotoxins, and plant toxins in cereals and fruits and vegetables. Analytical conditions were optimized to maximize the scope of the targeted molecules, the reliability of compound identification, and quantification performance within a single method. The method was further transferred and validated in another laboratory to assess its ruggedness. Validation according to the SANTE 11312/2021v2 guidelines showed that 92% and 98% of the molecules fulfill the quantification criteria at the lowest validated level in the cereals and fruits and vegetables groups, respectively. Analysis of fifteen certified reference materials led to a 96% satisfactory rate of z-scores confirming method's competitiveness. Furthermore, the occurrence of these contaminants was studied in 205 cereals and grains samples collected worldwide. The low µg/kg quantification limits make this LC-HRMS method a valuable tool to ensure compliance toward regulations and to screen for non-regulated substances for which occurrence data are crucial for an appropriate risk evaluation. [ABSTRACT FROM AUTHOR]

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. A simple liquid chromatography (LC) method was developed, where significant resolution between Dalbavancin, its homologs, and the generated degradation products was achieved. Seven degradation products were identified under acidic, basic, and oxidative stress conditions. Using liquid chromatography and high‐resolution mass spectrometry (LC‐HRMS), MS/MS studies, the generated degradation products were identified and characterized. Formation of isomeric degradation products was identified especially upon exposure to basic stress conditions. The mechanistic fragmentation pathway for the seven degradation products was established, and the chemical structure for the identified degradation products was elucidated. The results strongly suggest that Dalbavancin is highly susceptible to degradation under oxidative and hydrolytic stress conditions. This study provides insights into the hydrolytic and oxidative stability of Dalbavancin, which can be employed during drug development and discovery in synthesizing relatively stable analogs. [ABSTRACT FROM AUTHOR]

The widespread use of per- and polyfluoroalkyl substances (PFASs) with different physico–chemical properties poses a great threat to the environment and human health. Simultaneous detection of different classes of PFASs is a difficult task, especially for rapid analysis of polluted water samples in environmental forensic cases. In this study, a simple sample preparation ultrahigh-performance liquid chromatography coupled with quadrupole Orbitrap high-resolution mass spectrometry was established for the detection of PFASs in a wide range of water matrices. By optimizing the conditions of pretreatment and the parameters of the instrument, the developed method provided good linearity of calibration standards (R2 > 0.99), and demonstrated excellent MLOQ (0.008–1.2 µg/L), with spiked recoveries ranging from 57.7% to 151% for 47 targets in surface water samples, and from 45.7 to 165% for 46 targets in ground and waste water samples, respectively. This method required an injection volume of 3 µL and an analysis time of only 18 min per sample. The validation method was successfully applied to the analysis of 20 environmental water samples, in which 15 target substances with different concentrations were detected, with total concentrations of 0.082 to 262.455 μg/L. The method is simple and exclusive, and can rapidly confirm the occurrence of PFASs in different water samples, providing a convenient and fast high-throughput analysis, which is especially suitable for the application in the environmental forensic investigation of PFASs pollution. [ABSTRACT FROM AUTHOR]

Immature citruses are an important resource for the pharmaceutical industry due to their high levels of metabolites with health benefits. In this study, we used untargeted liquid chromatography–mass spectrometry (LC-MS) metabolomics to investigate the changes associated with fruit size in immature citrus fruits in the first weeks of growth. Three orange cultivars (Citrus sinensis 'Navel', Citrus sinensis 'Valencia', and Citrus sinensis 'Valencia Late') and a mandarin (Citrus reticulata Blanco 'Fremont') were separated into eight fruit sizes, extracted, and analyzed. Statistical analyses revealed a distinct separation between the mandarin and the oranges based on 56 metabolites, with an additional separation between the 'Navel' orange and the 'Valencia' and 'Valencia Late' oranges based on 21 metabolites. Then, metabolites that evolved significantly with fruit size growth were identified, including 40 up-regulated and 31 down-regulated metabolites. This study provides new insights into the metabolite modifications of immature Citrus sinensis and Citrus reticulata in the first weeks of growth and emphasizes the significance of including early sampled fruits in citrus maturation studies. [ABSTRACT FROM AUTHOR]

We investigated the pharmacokinetic pathway of berberine and its metabolites in vitro, in Caco-2 cells, and in human participants following the administration of dihydroberberine (DHB) and micellar berberine (LipoMicel®, LMB) formulations. A pilot trial involving nine healthy volunteers was conducted over a 24 h period; blood samples were collected and subjected to Ultra High-Performance Liquid Chromatography–High Resolution Mass Spectrometry (UHPLC-HRMS) analyses to quantify the concentrations of berberine and its metabolites. Pharmacokinetic correlations indicated that berberrubine and thalifendine follow distinct metabolic pathways. Additionally, jatrorrhizine sulfate appeared to undergo metabolism differently compared to the other sulfated metabolites. Moreover, berberrubine glucuronide likely has a unique metabolic pathway distinct from other glucuronides. The human trial revealed significantly higher blood concentrations of berberine metabolites in participants of the DHB treatment group compared to the LMB treatment group—except for berberrubine glucuronide, which was only detected in the LMB treatment group. Similarly, results from in vitro investigations showed significant differences in berberine metabolite profiles between DHB and LMB. Dihydroberberine, dihydroxy-berberrubine/thalifendine and jatrorrhizine sulfate were detected in LMB-treated cells, but not in DHB-treated cells; thalifendine and jatrorrhizine-glucuronide were detected in DHB-treated cells only. While DHB treatment provided higher blood concentrations of berberine and most berberine metabolites, both in vitro (Caco-2 cells) and in vivo human studies showed that treatment with LMB resulted in a higher proportion of unmetabolized berberine compared to DHB. These findings suggest potential clinical implications that merit further investigation in future large-scale trials. [ABSTRACT FROM AUTHOR]

Peptide pools consist of short amino acid sequences and have proven to be versatile tools in various research areas in immunology and clinical applications. They are commercially available in many different compositions and variants. However, unlike other reagents that consist of only one or a few compounds, peptide pools are highly complex products which makes their quality control a major challenge. Quantitative peptide analysis usually requires sophisticated methods, in most cases isotope-labeled standards and reference materials. Usually, this would be prohibitively laborious and expensive. Therefore, an approach is needed to provide a practical and feasible method for quality control of peptide pools. With insufficient quality control, the use of such products could lead to incorrect experimental results, worsening the well-known reproducibility crisis in the biomedical sciences. Here we propose the use of ultra-high performance liquid chromatography (UHPLC) with two detectors, a standard UV detector at 214 nm for quantitative analysis and a high-resolution mass spectrometer (HRMS) for identity confirmation. To be cost-efficient and fast, quantification and identification are performed in one chromatographic run. An optimized protocol is shown, and different peak integration methods are compared and discussed. This work was performed using a peptide pool known as CEF advanced, which consists of 32 peptides derived from cytomegalovirus (CMV), Epstein–Barr virus (EBV) and influenza virus, ranging from 8 to 12 amino acids in length. [ABSTRACT FROM AUTHOR]

Orbitrap mass spectrometry in full scan mode enables the simultaneous detection of hundreds of metabolites and their isotope-labeled forms. Yet, sensitivity remains limiting for many metabolites, including low-concentration species, poor ionizers, and low-fractional-abundance isotope-labeled forms in isotope-tracing studies. Here, we explore selected ion monitoring (SIM) as a means of sensitivity enhancement. The analytes of interest are enriched in the orbitrap analyzer by using the quadrupole as a mass filter to select particular ions. In tissue extracts, SIM significantly enhances the detection of ions of low intensity, as indicated by improved signal-to-noise (S/N) ratios and measurement precision. In addition, SIM improves the accuracy of isotope-ratio measurements. SIM, however, must be deployed with care, as excessive accumulation in the orbitrap of similar m/z ions can lead, via space-charge effects, to decreased performance (signal loss, mass shift, and ion coalescence). Ion accumulation can be controlled by adjusting settings including injection time and target ion quantity. Overall, we suggest using a full scan to ensure broad metabolic coverage, in tandem with SIM, for the accurate quantitation of targeted low-intensity ions, and provide methods deploying this approach to enhance metabolome coverage. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Chinese Mass Spectrometry Society is the property of Journal of Chinese Mass Spectrometry Society and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

This manuscript presents a comprehensive review of high‐resolution mass spectrometry in the field of food analysis and metabolomics. We have followed the historical evolution of metabolomics, its associated techniques and technologies, and its increasing role in food science and research. The review provides a critical comparison and synthesis of tentative identification guidelines proposed for over 15 years, offering a condensed resource for researchers in the field. We have also examined a wide range of recent metabolomics studies, showcasing various methodologies and highlighting key findings as a testimony of the versatility of the field and the possibilities it offers. In doing so, we have also carefully provided a compilation of the software tools that may be employed in this type of studies. The manuscript also explores the prospects of high‐resolution mass spectrometry and metabolomics in food science. By covering the history, guidelines, applications, and tools of metabolomics, this review attempts to become a comprehensive guide for researchers in a rapidly evolving field. [ABSTRACT FROM AUTHOR]

Purslane (Portulaca oleracea L., Portulacaceae) is a widespread edible plant with significant ethnobotanical and ethnopharmacological importance. The plant is characteristic for the presence of a class of indoline amide glucoside alkaloids, called cyclo-dopa amides, or oleraceins. Additionally, a new, structurally similar to oleraceins, class of indole amides have been discovered recently, called oleraindoles. These compounds have been evaluated to possess antiinflammatory and anticholinesterase activities. Herein, utilizing UHPLC-Orbitrap-MS with MS² filtering by diagnostic ion filtering (DIF), and diagnostic difference filtering (DDF) using different data analysis tools, eight compounds with oleraindole structure were tentatively identified. [ABSTRACT FROM AUTHOR]

Food supplements should not contain substances considered unsafe or pose a health risk to consumers. In recent years illegal adulterants have been found in various functional foods without notification of their presence or amount in the labelling. In this study, a validated method was developed and applied as a screening method to detect 124 forbidden substances belonging to 13 classes of compounds in food supplements. Liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) and a simple and rapid extraction protocol was applied to 110 food supplements collected from the internet market or during official controls in Italy. The percentage of non-compliant samples was 4.5%, relatively high compared with the official control results for these substances usually obtained on other food matrices. The results suggested the need to strengthen controls in this field to detect food supplement adulteration, which represents a potential health risk for the consumer. [ABSTRACT FROM AUTHOR]

Targeted biomonitoring studies quantifying the concentration of zeranols in biological matrices have focused on liquid chromatography interfaced to mass spectrometry (LC–MS). The MS platform for measurement, quadrupole, time-of-flight (ToF), ion trap, etc., is often chosen based on either sensitivity or selectivity. An instrument performance comparison of the benefits and limitations using matrix-matched standards containing 6 zeranols on 4 MS instruments, 2 low-resolution (linear ion traps), and 2 high-resolution (Orbitrap and ToF) was undertaken to identify the best measurement platform for multiple biomonitoring projects characterizing the endocrine disruptive properties of zeranols. Analytical figures of merit were calculated for each analyte to compare instrument performance across platforms. The calibration curves had correlation coefficients r = 0.989 ± 0.012 for all analytes and LODs and LOQs were ranked for sensitivity: Orbitrap > LTQ > LTQXL > G1 (V mode) > G1 (W mode). The Orbitrap had the smallest measured variation (lowest %CV), while the G1 had the highest. Instrumental selectivity was calculated using full width at half maximum (FWHM) and as expected, the low-resolution instruments had the broadest spectrometric peaks, concealing coeluting peaks under the same mass window as the analyte. Multiple peaks from concomitant ions, unresolved at low resolution (within a unit mass window), were present but did not match the exact mass predicted for the analyte. For example, the high-resolution platforms were able to differentiate between a concomitant peak at 319.1915 from the analyte at 319.1551, included in low-resolution quantitative analyses demonstrating the need to consider coeluting interfering ions in biomonitoring studies. Finally, a validated method using the Orbitrap was applied to human urine samples from a pilot cohort study. [ABSTRACT FROM AUTHOR]

The vast pool of structurally and functionally distinct secondary metabolites (i.e., natural products (NPs)) is constantly being expanded, a process also driven by the rapid progress in the development of analytical techniques. Such NPs often show potent biological activities and are therefore prime candidates for drug development and medical applications. The ethyl acetate extract of the tuber of Citrullus naudinianus (C. naudinianus), an African melon with edible fruits and seeds, shows in vitro immunomodulatory activity presumably elicited by cucurbitacins that are known major constituents of this plant. Further potentially immunomodulatory cucurbitacins or cucurbitacin derivatives were assumed to be in the tuber. Given the typically high content of cucurbitacins with similar physicochemical features but often distinct bioactivities, an efficient and reliable separation process is a prerequisite for their detailed characterization and assessment in terms of bioactivity. We therefore developed a detection method to screen and differentiate cucurbitacins via high-performance liquid chromatography/quadrupole-time-of-flight tandem mass spectrometry (HPLC-QTOF-MS/MS). In order to confirm the identification, the fragmentation patterns of two cucurbitacins and one 23,24-dihydrocucurbitacin were also investigated. Six characteristic fragments were identified and three of them were employed for the identification of cucurbitacins and 23,24-dihydrocucurbitacins in the extract. As a result, in addition to eight previously reported cucurbitacins from this plant four distinct 23,24-dihydrocucurbitacins (B, D, E, and I) were putatively identified and newly found in the ethyl acetate extract of the tuber of C. naudinianus. The established methodology enables rapid and efficient LC-MS-based analysis and identification of cucurbitacins and 23,24-dihydrocucurbitacins in plant extracts. [ABSTRACT FROM AUTHOR]

Synovial fluid (SF) from human knee joints with osteoarthritis (OA) has elevated levels of lysophosphatidylcholine (LPC) species, but their functional role is not well understood. This in vitro study was designed to test the hypothesis that various LPCs found elevated in OA SF and their metabolites, lysophosphatidic acids (LPAs), modulate the abundance of proteins and phospholipids (PLs) in human fibroblast-like synoviocytes (FLSs), with even minute chemical variations in lysophospholipids determining the extent of regulation. Cultured FLSs (n = 5–7) were treated with one of the LPC species, LPA species, IL-1β, or a vehicle. Tandem mass tag peptide labeling coupled with LC-MS/MS/MS was performed to quantify proteins. The expression of mRNA from regulated proteins was analyzed using RT-PCR. PL synthesis was determined via ESI-MS/MS, and the release of radiolabeled PLs was determined by means of liquid scintillation counting. In total, 3960 proteins were quantified using multiplexed MS, of which 119, 8, and 3 were significantly and reproducibly regulated by IL-1β, LPC 16:0, and LPC 18:0, respectively. LPC 16:0 significantly inhibited the release of PLs and the synthesis of phosphatidylcholine, LPC, and sphingomyelin. Neither LPC metabolite—LPA 16:0 nor LPA 18:0—had any reproducible effect on the levels of each protein. In conclusion, small chemical variations in LPC species can result in the significantly altered expression and secretion of proteins and PLs from FLSs. IL-1β influenced all proteins that were reproducibly regulated by LPC 16:0. LPC species are likely to modulate FLS protein expression only in more advanced OA stages with low IL-1β levels. None of the eight proteins being significantly regulated by LPC 16:0 have been previously reported in OA. However, our in vitro findings show that the CD81 antigen, calumenin, and B4E2C1 are promising candidates for further study, focusing in particular on their potential ability to modulate inflammatory and catabolic mechanisms. [ABSTRACT FROM AUTHOR]

Summary: Mepiquat (MQ) and chlormequat (CQ) are two main food contaminants that could be naturally present in various animal and plant foods due to heat treatments. Cocoa and its derivative products seem to be a matrix inclined, for both chemical characteristics and production technologies, to the formation of MQ and CQ. In this study, exploiting the selectivity and sensitivity of IC‐HRMS, a new reliable analytical method was developed and validated for MQ and CQ quantification. The LOD of the method was set at 1.9 μg kg−1 for both CQ and MQ, and the linearity was defined from 6.25 to 625 μg kg−1. Average recoveries, at 2 μg L−1, were 96.5% and 97.5% for CQ and MQ, respectively, while precision as repeatability (RSDr%) was 2.3% and 5.1% respectively. Subsequently, a laboratory‐scale experiment was conducted to assess, for the first time, the actual risk of MQ and CQ formation, confirming in cocoa products the presence of MQ at detectable concentration after 55 min at 180 °C, approximately 8 μg kg−1 for cocoa and 7 μg kg−1 for chips. However, the investigation of a wide variety of commercial cocoa and nuts products has fortunately ruled out the presence of such contaminants at detectable concentrations. [ABSTRACT FROM AUTHOR]

A new laboratory OrbitrapTM cell-based mass spectrometer, OLYMPIA (Orbitrap anaLYseur MultiPle IonisAtion), without a C-trap module, has been developed and constructed. The first operation of the OrbitrapTM cell-based device with the continuous ion source and without the C-trap module is reported. OLYMPIA is being developed and used as a workbench platform to test and develop technologies for the next generation of spaceborne mass spectrometers and as a laboratory instrument to perform high-resolution studies of space-relevant chemical processes. This instrument has been used to measure the quantitative composition of CO/N2/C2H4 mixtures of the same nominal mass using an electron ionization ion source. The relative abundance of ions has been measured using a short acquisition time (up to 250 ms) with a precision of better than 10% (for most abundant ions) and a mass resolution of 30,000–50,000 (full width at half maximum) over the mass range of m/z 28–86. The achieved mass accuracy of measurements is better than 20 ppm. This performance level is sufficient to resolve and identify the CO/N2/C2H4 components of the mixtures. The dynamic range and relative ion abundance measurements have been evaluated using a reference normal isotopic distribution of krypton gas. The measurement accuracy is about 10% for the 4 most abundant isotopes; 6 isotopes are detectable. [ABSTRACT FROM AUTHOR]

Pharmaceutical analysis refers to an area of analytical chemistry that deals with active compounds either by themselves (drug substance) or when formulated with excipients (drug product). In a less simplistic way, it can be defined as a complex science involving various disciplines, e.g., drug development, pharmacokinetics, drug metabolism, tissue distribution studies, and environmental contamination analyses. As such, the pharmaceutical analysis covers drug development to its impact on health and the environment. Moreover, due to the need for safe and effective medications, the pharmaceutical industry is one of the most heavily regulated sectors of the global economy. For this reason, powerful analytical instrumentation and efficient methods are required. In the last decades, mass spectrometry has been increasingly used in pharmaceutical analysis both for research aims and routine quality controls. Among different instrumental setups, ultra-high-resolution mass spectrometry with Fourier transform instruments, i.e., Fourier transform ion cyclotron resonance (FTICR) and Orbitrap, gives access to valuable molecular information for pharmaceutical analysis. In fact, thanks to their high resolving power, mass accuracy, and dynamic range, reliable molecular formula assignments or trace analysis in complex mixtures can be obtained. This review summarizes the principles of the two main types of Fourier transform mass spectrometers, and it highlights applications, developments, and future perspectives in pharmaceutical analysis. [ABSTRACT FROM AUTHOR]

Herein, a chemophenetic significance, based on the phenolic metabolite profiling of three Senecio (S. hercynicus, S. ovatus, and S. rupestris) and two Jacobaea species (J. pancicii and J. maritima), coupled to morphometric data, is presented. A set of twelve morphometric characters were recorded from each plant species and used as predictor variables in a linear discriminant analysis (LDA) model. From a total 75 observations (15 from each of the five species), the model correctly assumed their species' membership, except for 2 observations. Among the studied species, S. hercynicus and S. ovatus presented the greatest morphological similarity. A phytochemical profiling of phenolic specialized metabolites by UHPLC-Orbitrap-MS revealed 46 hydroxybenzoic, hydroxycinnamic, and acylquinic acids and their derivatives, 1 coumarin and 21 flavonoids. Hierarchical and PCA clustering applied to the phytochemical data corroborated the similarity of S. hercynicus and S. ovatus, observed in the morphometric analysis. This study contributes to the phylogenetic relationships between the tribe Senecioneae taxa and highlights the chemophenetic similarity/dissimilarity of the studied species belonging to Senecio and Jacobaea genera. [ABSTRACT FROM AUTHOR]

(1) Background: In toxicological laboratories, various screening methods can be used to identify compounds involved in intoxication. High-resolution mass spectrometry has been increasingly used in this context for the last years, because of its sensitivity and reliability. Here, we present the development and validation of a screening method that uses liquid chromatography coupled with a high-resolution mass spectrometer. (2) Methods: This method required only 100 µL of whole blood or plasma sample. Pretreatment consisted of a rapid and simple deproteinisation with methanol/acetonitrile and zinc sulphate. This new assay was validated according to international guidelines. (3) Results: To perform the method validation, 53 compounds were selected. The selection criteria were as follows: various chemical structures and therapeutic families (>15), large m/z distribution, positive or negative ionisation mode, and various elution times. The assays showed high selectivity and specificity, with optimal process efficiency. The identification limits, determined using predefined criteria, were established at sub-therapeutic or therapeutic concentrations. Applicability was evaluated using spiked plasma controls and external quality controls. (4) Conclusions: The new method was then successfully applied to routine clinical and forensic samples. [ABSTRACT FROM AUTHOR]

Modern mass spectrometry is the most demanded method of analysis of biopolymers, first of all, proteins and peptides. The novel methods of ionization of biomolecules developed at the end of the 20th century allowed increasing the efficiency of mass spectrometry in life sciences, first of all, proteomics. The methods of qualitative and quantitative analysis of these compounds are permanently improving. They become faster, more reliable, informative, sensitive, and selective. Significant role plays automation based on the original algorithms of the machine data processing and usage of spectral databases. One of the challenging questions of mass spectrometry involves analysis of polypeptides of various living organisms, whose genomes remain unknown so far. Amphibians represent a typical example of that species. In these cases, de novo sequencing of novel peptides is required. Development of an automated method of the de novo sequencing exclusively by mass spectrometry is of high scientific priority. It deals with top-down analysis, when an array of tandem mass spectrometry tools allows establishing amino acid sequence of each peptide using their polyprotonated molecules as precursor ions and eliminating assembling based on the hydrolysates of these compounds. A number of nuances still require time-consuming manual spectral elucidation. The present review throws light upon certain problems of mass spectrometry for the de novo sequencing of natural peptides of ranid and hylid frogs as typical representatives of amphibians. It tackles the questions related to post translational modifications, differentiation of isobaric and isomeric amino acid residues, cyclization of short peptides, and application of complementary methods of tandem mass spectrometry to trigger fragmentation. The benefits of mass spectrometry in taxonomy studies, e.g., for the reliable differentiation of closely related ranid and hylid frog species and even populations, are also discussed. [ABSTRACT FROM AUTHOR]

The present study aimed to assess the effects of epileptic treatment of the aerial part of Bupleurum chinense DC. in kainic acid (KA)-induced epilepsy rats and LPS-induced BV2 cells, as well as to identify the active chemical constituents. The in vivo and vitro results showed that 20% ethanol elution fractions of the aerial part of B. chinense DC. (BCE-20) and 70% ethanol elution fractions of the aerial part of B. chinense DC. (BCE-70) could improve the epileptic state of the rats and status epilepticus (SE%). Moreover, ultra-high-performance liquid chromatography (UPLC)-Orbitrap- mass spectrometry (MS) analysis identified BCE-20 and 70 as flavonoids and phenylpropanoids, respectively. The mechanistic analysis also showed that BCE-20 and 70 could regulate neurotransmitter abnormalities and suppresses the expression and secretion of pro-inflammatory cytokines. Notably, BCE-20 and 70 could regulate the Triggering receptor expressed on myeloid cells 2 (TREM2)/nuclear factor-k-gene binding (NF-κB)/inhibitor of NF-κB α (IκBα) pathway to inhibit the neuroinflammation. Our findings support the ethnopharmacological use of the constituent polyphenols and flavonoids from the aerial part of B. chinense DC., as the strong anti-epileptic agents. [ABSTRACT FROM AUTHOR]

The water-based extract of broad bean hulls contains several bioactive molecules, including polyphenols well-known to exert antioxidant activity, which could justify its use in nutraceutical formulations. Hence, the current investigation aimed to establish the polyphenolic profile of water-based extracts from broad bean hulls through UHPLC–Q-Orbitrap HRMS analysis. The findings highlighted that p-coumaric acid, chlorogenic acid, and epicatechin were the most common compounds found in the tested extracts, being quantified at a mean concentration of 42.1, 32.6, and 31.2 mg/100 g, respectively. Moreover, broad bean hull extracts were encapsulated into a nutraceutical formulation, after which the antioxidant properties and the bioaccessibility of phenolic compounds during the simulated gastrointestinal (GI) process were investigated and compared with the digested non-encapsulated extract. The data highlighted that following the GI process, the capsules were able to preserve active compounds from the adverse effects of digestion, resulting in a greater antioxidant capacity and polyphenol bioaccessibility in the duodenal and colonic phases, compared with the non-encapsulated extract. Our results showed that the water extract from broad bean hulls may be considered a valuable source of natural polyphenolic compounds; in addition, the use of a gastric-resistant capsule could be a suitable alternative to transport these bioactive compounds to the target tissues. [ABSTRACT FROM AUTHOR]

Litchi fruit is consumed across the globe for its high nutritional value and taste. The qualitative profiling of litchi fruit has been carried out by using ultra-high-performance liquid chromatography with QExactive high-resolution accurate mass spectrometry. Acidified water: methanol: acetonitrile (1:1:1) extracts from individual parts (skin, pulp, and seed) of matured litchi, were subjected to LC–MS analysis with electrospray ionization in full MS-ddMS2 mode as a non-target approach. The data was processed through compound discoverer software by the use of mzCloud and ChemSpider databases, for compound identification. We identified 77 compounds with protonated or deprotonated forms based on the polarity and their characteristic fragments are within ± 4 ppm mass error and retention time ± 0.1 min for parent and fragments. Hypoglycin B is the first time reported in litchi fruit along with hypoglycin A. Further, we verified the distribution of the identified components and differentiation of three different parts of litchi through principal component analysis. [ABSTRACT FROM AUTHOR]

Cultivated soybean (Glycine max (L.)), the world's most important legume crop, has high-to-moderate salt sensitivity. Being the frontier for sensing and controlling solute transport, membrane proteins could be involved in cell signaling, osmoregulation, and stress-sensing mechanisms, but their roles in abiotic stresses are still largely unknown. By analyzing salt-induced membrane proteomic changes in the roots and leaves of salt-sensitive soybean cultivar (C08) seedlings germinated under NaCl, we detected 972 membrane proteins, with those present in both leaves and roots annotated as receptor kinases, calcium-sensing proteins, abscisic acid receptors, cation and anion channel proteins, proton pumps, amide and peptide transporters, and vesicle transport-related proteins etc. Endocytosis, linoleic acid metabolism, and fatty acid biosynthesis pathway-related proteins were enriched in roots whereas phagosome, spliceosome and soluble NSF attachment protein receptor (SNARE) interaction-related proteins were enriched in leaves. Using label-free quantitation, 129 differentially expressed membrane proteins were found in both tissues upon NaCl treatment. Additionally, the 140 NaCl-induced proteins identified in roots and 57 in leaves are vesicle-, mitochondrial-, and chloroplast-associated membrane proteins and those with functions related to ion transport, protein transport, ATP hydrolysis, protein folding, and receptor kinases, etc. Our proteomic results were verified against corresponding gene expression patterns from published C08 RNA-seq data, demonstrating the importance of solute transport and sensing in salt stress responses. [ABSTRACT FROM AUTHOR]

Copyright of Evaluation & Analysis of Drug-Use in Hospitals of China is the property of Evaluation & Analysis of Drug-Use in Hospitals of China Editorial Board and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Producers of drinking water (DW) occasionally require chemical identification of new or unexpected contamination, e.g., caused by an incident. The state-of-the-art technique for the identification of organic compounds is High-Resolution Mass Spectrometry (HRMS). The ability to discover unexpected compounds at low concentrations in any sample by HRMS is facilitated by comparison to non-contaminated reference samples. Samples of raw and drinking water were collected regularly over one whole year from six Swedish drinking water treatment plants (DWTPs). The samples were analyzed by LC-HRMS together with spiked samples mimicking an incident. This setup enabled evaluation of the significance of having access to a collection of matrix-matched reference samples. The main variation in the organic compound profile in the data set was explained by the uniqueness of the raw water and purification steps in the individual DWTPs. Seasonal variations were also significant but subordinate. This subject was further explored by the analysis of drinking water sampled once from 90 of the 1750 Swedish DWTPs, where a similarity between DW originating from rock aquifers was observed. No other significant correlation between samples was observed—e.g., other types of raw water, which types of purification steps were involved, or which additives or process chemicals were added—which could aid in the selection of relevant reference samples. The conclusion from the study is that it would be imperative for the DWTPs to have access to their individual reference samples for use in the investigation of an incident. A library of such reference samples, e.g., collected monthly and covering the last 12 months, could be stored and used together with a fresh "suspected sample" for non-target HRMS investigations as described. [ABSTRACT FROM AUTHOR]

A method for the simultaneous screening of 258 pesticide residues in silage using modified QuEChERS combined with liquid chromatography (LC)- and gas chromatography (GC)- quadrupole-Orbitrap mass spectrometry (Q-Orbitrap/MS) has been developed. After hydration, the silage was homogenized with a 1% acetic acid–acetonitrile solution, and the extract was purified using C18, PSA, and anhydrous magnesium sulfate. Finally, the sample was detected using LC/GC-Q-Orbitrap/MS, and quantified using an external standard method. The results showed that 258 pesticides had an excellent linear relationship in the range of 0.1–50 μg L−1, and that the coefficients of determination (R2) were more than 0.99. The screening detection limit (SDL) of silage was in the range of 0.5–50 μg kg−1, and the limit of quantitation (LOQ) was in the range of 1–50 μg kg−1. The accuracy and precision of the method were verified at the spiked levels of 1-, 2- and 10-times LOQ, and the recovery of 258 pesticides was in the range of 66.5–119.8%, with relative standard deviations (RSDs) of less than 20% (n = 6). This method was simple, rapid, and reliable, and could be applied to screen and quantify multi-pesticide residues in silage. [ABSTRACT FROM AUTHOR]

The principal purpose of this study was to determine antioxidant and ACE-inhibitory (angiotensin converting enzyme) activities of 5 ripened Turkish cheese varieties (Kashar, Erzincan Tulum, Izmir Tulum Gruyere, Mihalic). Among the water-soluble extracts with a molecular weight smaller than 3 kDa Gruyere and Erzincan Tulum exhibited the highest free radical scavenger activity in FRAP (Ferric Reducing Ability of Plasma) and ABTS [2,20-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid)] assays, respectively. For the inhibition of ACE, Mihalic was more effective than others. The water-soluble extracts of five cheese were analyzed with liquid chromatography-tandem mass spectrometry to identify the peptide sequences. Totally 394 peptides were sequenced; of these, 134 from αs1-casein, 44 from αs2-casein, 198 from β-casein, and 18 from κ-casein. Among these sequences, 48 peptides were reported as bioactive with the activities including ACE-inhibitory, antioxidant, antimicrobial, DPP-IV inhibitory, antidiabetic, immunomodulating, antithrombotic, anti-inflammatory, and inhibition of cholesterol solubility. These results showed that Turkish cheeses are good sources of bioactive peptides. [ABSTRACT FROM AUTHOR]

A novel method for detecting pesticide multi‐residue in grass forage (alfalfa and oat) was established based on the one‐step automatic extraction and purification technology of quick, easy, cheap, effective, rugged, and safe combined with ultrahigh‐performance liquid chromatography quadrupole Orbitrap high‐resolution mass spectrometry. The crushed sample was extracted with acetonitrile with 1% acetate, followed by a cleanup step with a primary‐secondary amine, octadecylsilane, and graphitized carbon black. The extraction and purification were carried out using the one‐step automatic pretreatment equipment. The target pesticides were acquired in positive ion electrospray ionization mode and full scan/data dependent secondary scan mode. The calibration curve shows good linearity over the corresponding concentration range, with the coefficient of determination greater than 0.99. The screening detection limits were 0.5–50 μg/kg, and the limit of quantification for the 206 pesticides was set at 1–50 μg/kg. At the spiking levels of one, two, and 10 times of limit of quantification, more than 95% of pesticides had recovery between 70–120%, with a relative standard deviation ≤20%. The method was proved to be simple, rapid, high‐sensitivity, and could be routinely used for the high throughput screening and quantitative analysis of pesticide residues in alfalfa and oat. [ABSTRACT FROM AUTHOR]

The removal of contaminants of emerging concern (CECs) occurring in wastewater effluents, such as pharmaceutically active substances (PhACs) and personal care products, pose a big research challenge since they can be a major source of pollution for water bodies and a danger to public health. The objective of this work was to perform a comprehensive monitoring of a broad set of PhACs (>130) in a wastewater treatment plant (WWTP) close to Thessaloniki (Greece), as well as to evaluate the potential of heterogeneous catalytic ozonation for the removal of CECs from wastewater through a continuous flow system. The high-resolution mass spectrometry analysis revealed the highest average concentrations for irbesartan (1817 ng/L). Antihypertensives along with antibiotics, psychiatrics, and β-blockers were found to aggravate the effluents. Removal efficiency after conventional treatment was >30%. The results from catalytic ozonation unit operation indicate that the introduction of a proper solid material that acts as catalyst can enhance the removal of CECs. A preliminary risk assessment using the risk quotient (RQ) revealed that irbesartan and telmisartan entail high acute risk. The overall results underline the urgent need to incessantly monitor PhACs and expand the toxicological studies to establish the sublethal and chronic effects on aquatic organisms. [ABSTRACT FROM AUTHOR]

Ranibizumab is an FDA-approved drug used to treat wet age-related macular degeneration (AMD), diabetic retinopathy, macular edema, and myopic choroidal neovascularization. Bevacizumab is another drug often used off-label to treat wet AMD. In order to reduce unwanted angiogenesis, ranibizumab and bevacizumab target circulating VEGF-A in the eye. Concentration levels in human vitreous and aqueous humor can be used to provide valuable efficacy information. However, vitreous and aqueous humor's aqueous environment, and vitreous humor's viscosity, as well as the stickiness of the analytes can provide bioanalytical challenges. In this manuscript, we describe the development, optimization, and fit-for-purpose validation of an LC-HRMS method designed for intact quantitative bioanalysis of ranibizumab and bevacizumab in human vitreous and aqueous humor following intravitreal administration. In order to fully develop this method, evaluations were conducted to optimize the conditions, including the data processing model (extracted ion chromatograms (XICs) vs deconvolution), carryover mitigation, sample preparation scheme optimization for surrogate and primary matrices, use of internal standard/immunocapture/deglycosylation, and optimization of the extraction and dilution procedure, as well as optimization of the liquid chromatography and mass spectrometry conditions. Once the method was fully optimized, a fit-for-purpose validation was conducted, including matrix parallelism, with a linear calibration range of 10 to 200 µg/mL. The development of this intact quantitative method using LC-HRMS provides a proof-of-concept template for challenging, but valuable new and exciting bioanalytical techniques. [ABSTRACT FROM AUTHOR]

The main objectives of this study were to determine antioxidant, angiotensin-converting enzyme-inhibitory (ACE-I), and antibacterial activities of traditional Turkish white cheeses. The bioactivities of the cheeses tested in the low-molecular-weight fractions of water-soluble extracts. The effect of in vitro digestion on the functional potential of white cheeses was also considered. The antioxidant activity of water-soluble extracts of white cheeses was tested with FRAP (Ferric Reducing Ability of Plasma) and ABTS [2,20-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid)] assays. Cow cheese and goat cheese showed better antioxidant activities in the FRAP and ABTS assays, respectively. However, after in vitro digestion, goat cheese displayed higher radical scavenging activity than its counterparts. Goat cheese also exhibited better ACE-I activity, although ACE-I activity was inhibited by the in vitro digestion, probably due to the further proteolysis of ACE-I peptides. All samples showed an antibacterial effect on Salmonella typhimurium ATCC 14028 and Staphylococcus aureus ATCC 25923. The fractions were analyzed with orbitrap liquid chromatography–tandem mass spectrometry to determine the sequences of peptides. White cheeses contained 42 sequences previously reported as bioactive peptides with the functions of ACE-I, antioxidant, antimicrobial, inhibition of cholesterol solubility, osteoanabolic, and antidiabetic. White cheese is a rich source of bioactive peptides with various properties. [ABSTRACT FROM AUTHOR]

We present a novel, scan-centric method for characterizing peaks from direct injection multi-scan Fourier transform mass spectra of complex samples that utilizes frequency values derived directly from the spacing of raw m/z points in spectral scans. Our peak characterization method utilizes intensity-independent noise removal and normalization of scan-level data to provide a much better fit of relative intensity to natural abundance probabilities for low abundance isotopologues that are not present in all of the acquired scans. Moreover, our method calculates both peak- and scan-specific statistics incorporated within a series of quality control steps that are designed to robustly derive peak centers, intensities, and intensity ratios with their scan-level variances. These cross-scan characterized peaks are suitable for use in our previously published peak assignment methodology, Small Molecule Isotope Resolved Formula Enumeration (SMIRFE). [ABSTRACT FROM AUTHOR]

Food safety and quality issues are becoming increasingly important and attract much attention, requiring the development of better analytical platforms. For example, high-resolution (especially Orbitrap) mass spectrometry simultaneously offers versatile functions such as targeted/non-targeted screening while providing qualitative and quantitative information on an almost unlimited number of analytes to facilitate routine analysis and even allows for official surveillance in the food field. This review covers the current state of Orbitrap mass spectrometry (OMS) usage in food analysis based on research reported in 2012–2019, particularly highlighting the technical aspects of OMS application and the achievement of OMS-based screening and quantitative analysis in the food field. The gained insights enhance our understanding of state-of-the-art high-resolution mass spectrometry and highlight the challenges and directions of future research. [ABSTRACT FROM AUTHOR]

Natural biomass magnetic porous carbon was successfully prepared via a cost‐effective and green route using mangosteen shells as raw material. The prepared magnetic porous carbon was used as a magnetic solid‐phase extraction adsorbent for bisphenols enrichment from beverages followed by high‐performance liquid chromatography‐quadrupole‐Orbitrap high‐resolution mass spectrometry. Parameters affecting extraction efficiency including sample solution pH, adsorbent amount, extraction time, eluent type, and volume were optimized. Results showed that biomass magnetic porous carbon had excellent adsorption properties for bisphenols due to its large specific surface area and abundant functional groups, which could form hydrogen bonding and π‐π stacking with bisphenols. The enrichment factor of 3 bisphenols was in the range of 15–19. Under optimum conditions, favorable linearity for all analytes was obtained with correlation coefficients higher than 0.998. Recoveries of spiked samples were in the range of 88.5–105.1% with a relative standard deviation of 3.4–5.5%. These results demonstrated that magnetic porous carbon may be a promising adsorbent for the enrichment of aromatic compounds. [ABSTRACT FROM AUTHOR]

Endothelial cells (ECs) comprise the lumenal lining of all blood vessels and are critical for the functioning of the cardiovascular system. Their phenotypes can be modulated by alternative splicing of RNA to produce distinct protein isoforms. To characterize the RNA and protein isoform landscape within ECs, we applied a long read proteogenomics approach to analyse human umbilical vein endothelial cells (HUVECs). Transcripts delineated from PacBio sequencing serve as the basis for a sample-specific protein database used for downstream mass-spectrometry (MS) analysis to infer protein isoform expression. We detected 53,863 transcript isoforms from 10,426 genes, with 22,195 of those transcripts being novel. Furthermore, the predominant isoform in HUVECs does not correspond with the accepted "reference isoform" 25% of the time, with vascular pathway-related genes among this group. We found 2,597 protein isoforms supported through unique peptides, with an additional 2,280 isoforms nominated upon incorporation of long-read transcript evidence. We characterized a novel alternative acceptor for endothelial-related gene CDH5, suggesting potential changes in its associated signalling pathways. Finally, we identified novel protein isoforms arising from a diversity of RNA splicing mechanisms supported by uniquely mapped novel peptides. Our results represent a high-resolution atlas of known and novel isoforms of potential relevance to endothelial phenotypes and function. [ABSTRACT FROM AUTHOR]

Skin secretion represents the only means of defense for the majority of frog species. That phenomenon is based on the fact that the main components of the secretion are peptides demonstrating greatly varying types of bioactivity. They fulfill regulatory functions, fight microorganisms and may be even helpful against predators. These peptides are considered to be rather promising pharmaceuticals of future generation as according to the present knowledge microorganisms are unlikely to develop resistance to them. Mass spectrometry sequencing of these peptides is the most efficient first step of their study providing reliably their primary structures, i.e., amino acids sequence and S-S bond motif. Besides discovering new bioactive peptides, mass spectrometry appears to be an efficient tool of taxonomy studies, allowing for distinguishing not only between closely related species, but also between populations of the same species. Application of several tandem mass spectrometry tools (CID, HCD, ETD, EThcD) available with Orbitrap mass analyzer allowed us to obtain full sequence of about 60 peptides in the secretion of Slovenian population of brown ranid frog Rana temporaria. The problem of sequence inside C-terminal cycle formed by two Cys and differentiation of isomeric Leu and Ile residues was done in top-down mode without any derivatization steps. Besides general biomarkers of Rana temporaria species, Central Slovenian population of Rana temporaria demonstrates six novel temporins and one brevinin 1, which may be treated as biomarkers of that population. [ABSTRACT FROM AUTHOR]

β-agonists have been illegally used for growth promoting purposes in animal husbandry, leading to residue concentrations capable of inducing acute toxic reactions among consumers of animal-based food. There is not only a need for detecting β-agonist residues at low concentrations, but also to increase the number of compounds to be monitored. It was therefore the aim of this paper to develop a unified method capable of detecting a wide range of different β-agonists (20 analytes including some metabolites) in a variety of matrices (muscle, liver, plasma, milk and urine). The developed procedure permits the quick processing of samples with limited labour input and consumption of consumables. The method has been validated according to the Commission Decision 98/536/EC. Detection is based on ultrahigh-performance chromatography coupled to high-resolution mass spectrometry. Validation was performed on two different instruments (Orbitrap and time of flight). The obtained limit of quantification (0.05 to 0.5 μg/kg and the average recovery of 78% in the most complex matrix (liver) satisfies current regulatory requirements. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Forensic Medicine / Fayixue Zazhi is the property of Journal of Forensic Medicine Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Escherichia coli normally cannot grow in the environment. One environmental stress that prevents E. coli growth may be the competition for growth substrates with co‐existing micro‐organisms. In this study, the growth substrates of E. coli were screened by high‐resolution mass spectrometry and compared with those of indigenous bacteria in river water. In an incubation experiment, E. coli multiplied in sterilized river water, but did not multiply when indigenous micro‐organisms were present in the water. By analysing dissolved organic matter in the river water before and after E. coli growth, 35 compounds were identified as putative growth substrates of E. coli. Among them, 33 compounds were also identified as putative growth substrates of indigenous bacteria. These results indicate that E. coli and indigenous bacteria compete for organic substrates in river water, which could suppress the growth of E. coli. [ABSTRACT FROM AUTHOR]

Food can contain unwanted compounds and need to be analyzed for compounds like carcinogenic polycyclic aromatic hydrocarbons to ensure consumer safety. The analytes need to be extracted from the sample matrix and cleaned‐up to enable detection. However, established methods for clean‐up are labor intensive and have a high expenditure on organic solvents. Here, we show a newly developed micro‐solid‐phase extraction cartridge method to automate the clean‐up for analysis of polycyclic aromatic hydrocarbons in sunflower oil using gas chromatography with quadrupole‐orbitrap mass spectrometry with a TriPlus autosampler. This automated micro‐solid‐phase extraction cartridge method needs only 4 μL of vegetable oil sample and requires only 360 μL acetonitrile for elution, and, therefore, it needs only small amounts of organic solvent. Two different micro‐solid‐phase extraction cartridge methods were developed, one using two commercially available cartridges with florisil and octadecylsilane/Z‐Sep/CarbonX, and the other method using one commercially available cartridge with florisil followed by one self‐made cartridge with octadecylsilane/Z‐Sep. The latter method showed successful lipid removal and was further validated for 22 of 24 polycyclic aromatic hydrocarbon compounds in sunflower oil at a spiked level of 1090 μg/kg with recoveries ranging from 53 to 118% and relative standard deviation below 22%. This method shows promising short‐time clean‐up with low expenditure of solvents. [ABSTRACT FROM AUTHOR]

Quadrupole time‐of‐flight (QTof) collision‐induced dissociation (CID) and Orbitrap higher‐energy collisional dissociation (HCD) are the most commonly used fragmentation techniques in mass spectrometry‐based proteomics workflows. The information content of the MS/MS spectra is first and foremost determined by the applied collision energy. How can we set up the two instrument types to achieve maximum transferability? To answer this question, we compared MS/MS spectra obtained on a Bruker QTof CID and a Thermo Q‐Exactive Focus Orbitrap HCD instrument as a function of collision energy using the similarity index. Results show that with a few eV lower collision energy setting on HCD (Orbitrap‐specific CID) than on QTof CID, nearly identical MS/MS spectra can be obtained for leucine enkephalin pentapeptide standard, for selected +2 and +3 enolase tryptic peptides and for a large number of peptides in a HeLa protein digest. The Bruker QTof was able to produce colder ions, which may be significant to study inherently labile compounds. Further, we examined energy dependence of peptide identification confidence, as characterized by Mascot scores, on the HeLa peptides. In line with earlier QTof results, this dependence shows one or two maxima (unimodal or bimodal behavior) on Orbitrap. The fraction of bimodal peptides is lower on Orbitrap. Optimal energies as a function of m/z show a similar linear trend on both instruments, which suggests that with appropriate collision energy adjustment, matching conditions for proteomics can be achieved. Data have been deposited in the MassIVE repository (MSV000086434). [ABSTRACT FROM AUTHOR]

Purpose: Validated biomarkers of food intake (BFIs) have recently been suggested as a useful tool to assess adherence to dietary guidelines or compliance in human dietary interventions. Although many new candidate biomarkers have emerged in the last decades for different foods from metabolic profiling studies, the number of comprehensively validated biomarkers of food intake is limited. Apples are among the most frequently consumed fruits and a rich source of polyphenols and fibers, an important mediator for their health-protective properties. Methods: Using an untargeted metabolomics approach, we aimed to identify biomarkers of long-term apple intake and explore how apples impact on the human plasma and urine metabolite profiles. Forty mildly hypercholesterolemic volunteers consumed two whole apples or a sugar and energy-matched control beverage, daily for 8 weeks in a randomized, controlled, crossover intervention study. The metabolome in plasma and urine samples was analyzed via untargeted metabolomics. Results: We found 61 urine and 9 plasma metabolites being statistically significant after the whole apple intake compared to the control beverage, including several polyphenol metabolites that could be used as BFIs. Furthermore, we identified several endogenous indole and phenylacetyl-glutamine microbial metabolites significantly increasing in urine after apple consumption. The multiomic dataset allowed exploration of the correlations between metabolites modulated significantly by the dietary intervention and fecal microbiota species at genus level, showing interesting interactions between Granulicatella genus and phenyl-acetic acid metabolites. Phloretin glucuronide and phloretin glucuronide sulfate appeared promising biomarkers of apple intake; however, robustness, reliability and stability data are needed for full BFI validation. Conclusion: The identified apple BFIs can be used in future studies to assess compliance and to explore their health effects after apple intake. Moreover, the identification of polyphenol microbial metabolites suggests that apple consumption mediates significant gut microbial metabolic activity which should be further explored. [ABSTRACT FROM AUTHOR]

In an effort to characterize differently aged bourbons and to determine whether bourbons could be "fingerprinted" by their chemical compositions, we used Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) to characterize 2 distinct sets of bourbon samples. The first set of bourbons were prepared using the same mash bill but were aged differently (unaged (0 years), 2 years, 4 years, and 6 years) in oak barrels. The results showed an increase in the number of chemical compounds present as the bourbon ages. Most of the large changes in chemical composition occur in the first 2 years of aging. We also analyzed single barrel bourbons, which were produced identically but maturated in different, adjacent barrels, and the results suggested that significant differences exist among these samples. These results suggest that "fingerprinting" of different bourbons for authentication purposes may be complicated and that careful analyses, coupled with more comprehensive identification of chemical compounds in bourbons, are needed. [ABSTRACT FROM AUTHOR]

Matrix‐assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) allows for highly multiplexed, untargeted detection of many hundreds of analytes from tissue. Recently, laser postionization (MALDI‐2) has been developed for increased ion yield and sensitivity for lipid IMS. However, the dependence of MALDI‐2 performance on the various lipid classes is largely unknown. To understand the effect of the applied matrix on MALDI‐2 analysis of lipids, samples including an equimolar lipid standard mixture, various tissue homogenates, and intact rat kidney tissue sections were analyzed using the following matrices: α‐cyano‐4‐hydroxycinnamic acid, 2′,5′‐dihydroxyacetophenone, 2′,5′‐dihydroxybenzoic acid (DHB), and norharmane (NOR). Lipid signal enhancement of protonated species using MALDI‐2 technology varied based on the matrix used. Although signal improvements were observed for all matrices, the most dramatic effects using MALDI‐2 were observed using NOR and DHB. For lipid standards analyzed by MALDI‐2, NOR provided the broadest coverage, enabling the detection of all 13 protonated standards, including nonpolar lipids, whereas DHB gave less coverage but gave the highest signal increase for those lipids recorded. With respect to tissue homogenates and rat kidney tissue, mass spectra were compared and showed that the number and intensity of neutral lipids tentatively identified with MALDI‐2 using NOR increased significantly (e.g., fivefold intensity increase for triacylglycerol). In the cases of DHB with MALDI‐2, the number of protonated lipids identified from tissue homogenates doubled with 152 on average compared with 76 with MALDI alone. High spatial resolution imaging (~20 μm) of rat kidney tissue showed similar results using DHB with 125 lipids tentatively identified from MALDI‐2 spectra versus just 72 using standard MALDI. From the four matrices tested, NOR provided the greatest increase in sensitivity for neutral lipids (triacylglycerol, diacylglycerol, monoacylglycerol, and cholesterol ester), and DHB provided the highest overall number of lipids detected using MALDI‐2 technology. [ABSTRACT FROM AUTHOR]

To better understand cell-to-cell heterogeneity, advanced analytical tools are in a growing demand for elucidating chemical compositions of each cell within a population. However, the progress of single-cell chemical analysis has been restrained by the limitations of small cell volumes and minute cellular concentrations. Here, we present a rapid and sensitive method for investigating the lipid profiles of isolated single cells using infrared matrix-assisted laser desorption electrospray ionization mass spectrometry (IR-MALDESI-MS). In this work, HeLa cells were dispersed onto a glass slide, and the cellular contents were ionized by IR-MALDESI and measured using a Q-Exactive HF-X mass spectrometer. Importantly, this approach does not require extraction and/or enrichment of analytes prior to MS analysis. Using this approach, 45 distinct lipid species, predominantly phospholipids, were detected and putatively annotated from the single HeLa cells. The proof-of-concept study demonstrates the feasibility and efficacy of IR-MALDESI-MS for rapid lipidomic profiling of single cells, which provides an important basis for future work on differentiation between normal and diseased cells at various developmental states, which can offer new insights into cellular metabolic pathways and pathological processes. Although not yet accomplished, we believe this approach can be readily used as an assessment tool to compare the number of identified species during source evolution and method optimization (intra-laboratory), and also disclose the complementary nature of different direct analytical approaches for the coverage of different types of endogenous analytes (inter-laboratory). Graphical abstract [ABSTRACT FROM AUTHOR]

Hepatotoxic and genotoxic pyrrolizidine alkaloids have been involved in the acute poisoning of animals and humans. Crotalaria (Fabaceae) species contain these alkaloids. In this work, the diversity and distribution of pyrrolizidine alkaloids in roots, leaves, flowers, and seeds of Crotalaria pallida, Crotalaria maypurensis, Crotalaria retusa, Crotalaria spectabilis, Crotalaria incana, and Crotalaria nitens were studied. Matrix solid‐phase dispersion and ultra‐high‐performance liquid chromatography coupled with Orbitrap mass spectrometry were successfully employed in pyrrolizidine alkaloids extraction and analysis, respectively. Forty‐five pyrrolizidine alkaloids were detected and their identification was based on the mass spectrometry accurate mass measurement and fragmentation pattern analysis. The cyclic retronecine‐type diesters monocrotaline, crotaleschenine, integerrimine, usaramine, and their N‐oxides were predominantly present. Five novel alkaloids were identified for the first time in Crotalaria species, namely 14‐hydroxymonocrotaline, 12‐acetylcrotaleschenine, 12‐acetylmonocrotaline, 12‐acetylintegerrimine, and dihydrointegerrimine. Due to a lack of commercially available standards, the response factor of monocrotaline was used for quantification of pyrrolizidine alkaloids and their N‐oxides. Seeds and flowers possessed higher pyrrolizidine alkaloids amounts than roots and leaves. Due to their 1,2‐unsaturated pyrrolizidine alkaloids content, the ingestion of Crotalaria plant seeds or other parts through herbal products, infusions, or natural remedies is a serious health threat to humans and livestock. [ABSTRACT FROM AUTHOR]

A collaborative study was conducted to understand the correlation between pigments purity profile (primary aromatic amine content of the pigments) and the behaviour of these PAAs during cold water extraction (CWE) tests according to EN 645. From a selection of organic pigments based on seven colour indexes (PR122, PR184, PO13, PY74, PY111, PY138 and PY155), the pigment purity profile was established according to European Resolution AP (89) 1, then mono-pigmented inks were prepared and napkins printed with these inks. In a second step, cold water extraction and PAA determination were performed by two independent laboratories. In one laboratory, an analytical method based on LC-MS/MS was used, whereas in the other laboratory a method based on LC-HRMS using Orbitrap technology was developed for the simultaneous analysis of 35 PAAs. Good qualitative results were obtained if we consider that at significant levels the PAAs were positively detected in both laboratories, except for 3-amino-4-methoxybenzanilide and 8-amino-2-methyl-quinoline, for which inter-laboratory differences were observed. It was also shown that no contamination from unexpected PAAs was detected. The comparison between pigment analysis and CWE results shows that if the pigment purity profile is of major importance, other parameters such as pigment surface treatment, ink grinding process or ink formulation could have an important influence on the CWE results. For such sensitive applications, for example napkins or other Food Contact Materials (FCM), it is therefore recommended not only to select a pigment with a good purity profile but also to test the pigment in the final application. Finally, this work highlights the difficulty of validating a product on a single analysis and shows the importance of a multilevel global assessment on worst case application. [ABSTRACT FROM AUTHOR]