Analytical and Bioanalytical Chemistry最新文献

Chlorinated paraffins (CP) are complex molecular mixtures occurring in a wide range of isomers and homologs of environmental hazards, whose analytical complexity demand advanced mass spectrometry (MS) methods for their characterization. The reported formation of chlorinated olefins (COs) and other transformation products during CP biotransformation and degradation can alter the MS analysis, increasing the high resolution required to distinguish CPs from their degradation products. An advanced setup hyphenating a plasma ionization source and an external high-performance data acquisition and processing system to the legacy hybrid LTQ Orbitrap XL mass spectrometer is reported. First, the study demonstrated the versatility of a liquid sampling atmospheric pressure glow discharge, as a soft ionization technique, for CP analysis. Second, enhanced resolution and sensitivity provided by the absorption mode Fourier transform spectral representation on this legacy mass spectrometer are shown. The developed Orbitrap-based platform allowed the detection of new isotopic clusters and CPs and COs to be distinguished at medium resolution (setting 30,000 at m/z 400, ~ 400 ms transients), and even chlorinated di-olefins (CdiOs) at higher resolution (setting 100,000 at m/z 400, ~ 1500 ms transients). Overall, such proof-of-principle instrumental improvements are promising for environmental and analytical research in the field of CP analysis.

Mechanotransduction is the essential process that cells convert mechanical force into biochemical responses, and electrochemical sensor stands out from existing techniques by providing quantitative and real-time information about the biochemical signals during cellular mechanotransduction. However, the intracellular biochemical response evoked by mechanical force has been poorly monitored. In this paper, we report a method to apply local stretch on single cell and simultaneously monitor the ensuing intracellular biochemical signals. Specifically, a ferromagnetic micropipette was fabricated to locally stretch a single cell labeled with Fe₃O₄ nanoparticles under the external magnetic field, and the SiC@Pt nanowire electrode (SiC@Pt NWE) was inserted into the cell to monitor the intracellular hydrogen peroxide (H₂O₂) production induced by the local stretch. As a proof of concept, this work quantitatively investigated the elevated amount of H₂O₂ levels in single endothelial cell under different stretching amplitudes. This work puts forward a new research modality to manipulate and monitor the mechanotransduction at the single-cell level.

Exhaled breath volatilomics is a powerful non-invasive tool for biomarker discovery in medical applications, but compound annotation is essential for pathophysiological insights and technology transfer. This study was aimed at investigating the interest of a hybrid approach combining real-time proton transfer reaction-time-of-flight mass spectrometry (PTR-TOF-MS) with comprehensive thermal desorption-two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (TD-GCxGC-TOF-MS) to enhance the analysis and characterization of VOCs in clinical research, using COVID-19 as a use case. VOC biomarker candidates were selected from clinical research using PTR-TOF-MS fingerprinting in patients with COVID-19 and matched to the Human Breathomic Database. Corresponding analytical standards were analysed using both a liquid calibration unit coupled to PTR-TOF-MS and TD-GCxGC-TOF-MS, together with confirmation on new clinical samples with TD-GCxGC-TOF-MS. From 26 potential VOC biomarkers, 23 were successfully detected with PTR-TOF-MS. All VOCs were successfully detected using TD-GCxGC-TOF-MS, providing effective separation of highly chemically related compounds, including isomers, and enabling high-confidence annotation based on two-dimensional chromatographic separation and mass spectra. Four VOCs were identified with a level 1 annotation in the clinical samples. For future applications, the combination of real-time PTR-TOF-MS and comprehensive TD-GCxGC-TOF-MS, at least on a subset of samples from a whole study, would enhance the performance of VOC annotation, offering potential advancements in biomarker discovery for clinical research.

The soft ionization mechanism of helium-based plasma seems to be understood while it still remains challenging in argon-based plasma, although many studies have used argon plasmas as a soft ionization source with good ionization efficiencies. In this study, helium, argon, krypton, and xenon were fed into the same discharge geometry, a flexible micro-tube plasma (FµTP), to determine the ionization mechanisms. The FµTPs operated with the named noble gases obtained comparable ionization efficiencies by MS measurements. The optical emission results showed that N₂⁺ were the dominant ions within the helium-FµTP and noble gas ions were dominant for the other plasmas. These ions support the development of excitation and eventually stop at the end of the capillary. Therefore, Penning ionization and charge transfer between plasma and ambient air/analytes in the open atmosphere have been proven not to be the primary soft ionization mechanism. Furthermore, it was found that photoionization played a minor role in soft ionization. Using helium as a diagnosis gas in front of the discharge capillary nozzle of the FµTP, where the sample is usually positioned, shows that helium can be ignited by all of these FµTPs. This demonstrates that the excitation of a diagnosis gas as well as the ionization of analytes is independent of the type of the discharge gas. An alternative mechanism that a transient potential created by the ions is responsible for the soft ionization is subsequently proposed.

The increasing recognition of the health impacts from human exposure to per- and polyfluorinated alkyl substances (PFAS) has surged the need for sophisticated analytical techniques and advanced data analyses, especially for assessing exposure by food of animal origin. Despite the existence of nearly 15,000 PFAS listed in the CompTox chemicals dashboard by the US Environmental Protection Agency, conventional monitoring and suspect screening methods often fall short, covering only a fraction of these substances. This study introduces an innovative automated data processing workflow, named PFlow, for identifying PFAS in environmental samples using direct infusion Fourier transform ion cyclotron resonance mass spectrometry (DI-FT-ICR MS). PFlow's validation on a bream liver sample, representative of low-concentration biota, involves data pre-processing, annotation of PFAS based on their precursor masses, and verification through isotopologues. Notably, PFlow annotated 17 PFAS absent in the comprehensive targeted approach and tentatively identified an additional 53 compounds, thereby demonstrating its efficiency in enhancing PFAS detection coverage. From an initial dataset of 30,332 distinct m/z values, PFlow thoroughly narrowed down the candidates to 84 potential PFAS compounds, utilizing precise mass measurements and chemical logic criteria, underscoring its potential in advancing our understanding of PFAS prevalence and of human exposure.

In atmospheric pressure chemical ionization mass spectrometry (APCI-MS), [M-3H+H₂O]⁺ ions can deliver analyte-specific signals that enable direct analysis of volatile n-alkane mixtures. The underlying ionization mechanisms have been the subject of open debate, and in particular the role of water is insufficiently clarified to allow for reliable process analytics when the humidity level changes over time. This can be a problem, particularly in online monitoring, where analyte accumulation in the ion source can also occur. Here, we investigated the role of water during APCI-MS of volatile n-alkanes by changing the carrier gas for sample injection from a dry to a wetted state as well as by using ¹⁸O-labeled water. This allowed for a distinction between gaseous and surface-adsorbed water molecules. While adsorbed water seems to be responsible for the desired [M-3H+H₂O]⁺ signals through surface reactions with the analyte molecules, gaseous water was found to promote the formation of C_nH_2n+1O⁺ of different (and analyte-independent) hydrocarbons, revealing a reaction with hydrocarbon species which accumulated in the ion source during continuous operation. At the same time, gaseous water competed with analyte molecules for ionization and thus suppressed the formation of alkyl (C_nH_2n+1⁺) and alkenyl (C_nH_2n-1⁺) ions. The results reveal a memory effect due to hydrocarbon adsorption, which may cause severe interpretation difficulties when the ionization chamber undergoes sudden humidity changes. The use of [M-3H+H₂O]⁺ for n-alkane analysis in alkane/water mixtures can be facilitated by constantly maintaining high humidity and hence stabilizing the ionization conditions.

Nanocatalytic medicine, which aims to accurately target and effectively treat tumors through intratumoral in situ catalytic reactions triggered by tumor-specific environments or markers, is an emerging technology. However, the relative lack of catalytic activity of nanoenzymes in the tumor microenvironment (TME) has hampered their use in biomedical applications. Therefore, it is crucial to develop a highly sensitive probe that specifically responds to the TME or disease markers in the TME for precision diagnosis and treatment of diseases. In this work, a chiral photoacoustic (PA) nanoprobe (D/L-Ce@MoO₃) based on the H₂O₂-catalyzed TME activation reaction was constructed in a one-step method using D-cysteine (D-Cys) or L-cysteine (L-Cys), polymolybdate, and cerium nitrate as raw materials. The designed and synthesized D/L-Ce@MoO₃ chiral nanoprobe can perform in situ, non-invasive, and precise imaging of pharmacological acute liver injury. In vivo and in vitro experiments have shown that the D/L-Ce@MoO₃ probe had chiral properties, the CD signal decreased upon reaction with H₂O₂, and the absorption and PA signals increased with increasing H₂O₂ concentration. This is because of the catalytic reaction between Ce ions doped in the nanoenzyme and the high expression of H₂O₂ caused by drug-induced liver injury to produce ·OH, which has a strong oxidizing property to kill tumor cells and destroy the Mo-S bond in the probe, thus converting the chiral probe into an achiral polyoxometalate (POM) with PA signal.

Reverse transcription-digital PCR (RT-dPCR) is attracting attention as a method that enables SI-traceable RNA quantification without calibration, but its accuracy and bias have not been thoroughly studied. In this study, the accurate quantification of RNA by the RT-dPCR method was investigated using NMIJ CRM 6204-b, an RNA certified reference material whose certified value was assigned by orthogonal chemical measurement methods. Moreover, a two-step RT-dPCR method was adopted to examine in detail the conditions for the RT reaction process, which was expected to be the major uncertainty component in the RT-dPCR measurement. Optimization experiments revealed that the type of reverse transcriptase, the concentration of template RNA, and the type and concentration of primers in the RT reaction affected the value quantified by RT-dPCR. Under the optimal conditions, the value quantified by RT-dPCR, 76.4 ng/μL ± 6.7 ng/μL (the quantified value ± expanded uncertainty (k = 2)), was consistent with the certified value, 68.2 ng/μL ± 5.8 ng/μL, of NMIJ CRM 6204-b RNA 1000-A within the expanded uncertainty. From the results of the uncertainty evaluation, the relative combined uncertainty of the RT-dPCR method was 4.42%, and the major uncertainty components in the RT-dPCR method were the preparation of RT solution (3.68%), the inter-day difference (1.80%), and the RT reaction (1.30%). Together, the results suggested that the contribution of the RT reaction process to the total uncertainty was greater than that of the dPCR process.

The highly blistering sulfur mustard analogue agent T (bis(2-chloroethylthioethyl) ether), also known as O-mustard or oxy-mustard, is a common impurity in military grade sulfur mustard (SM) and a component of mixtures such as "HT" that are still found in old munitions. Together with sesquimustard (Q), it is the most important SM analogue and tightly regulated as a Schedule 1 chemical under the Chemical Weapons Convention. We report the adducts of T with nucleophilic Cys³⁴ and other residues in human serum albumin (HSA) formed in vitro. A micro liquid chromatography electrospray ionization high-resolution tandem-mass spectrometry method (µLC-ESI MS/HR MS) was developed for the detection and identification of biomarker peptides alkylated by a T-derived hydroxyethylthioethyloxyethylthioethyl (HETEOETE)-moiety (as indicated by an asterisk below). Following proteolysis of T-exposed human plasma with pronase, the dipeptide Cys³⁴*Pro and the single amino acid residue His* were produced. The use of proteinase K yielded Cys³⁴*ProPhe and the use of pepsin generated ValThrGlu⁴⁸*Phe, AlaGlu²³⁰*ValSerLysLeu, and LeuGlyMet³²⁹*Phe. Corresponding peptide-adducts of SM and Q were detected in a common workflow that in principle allowed the estimation of the mustard or mustard composition encountered during exposure. Novel adducts of Q at the Glu²³⁰ and Met²³⁹ residues were detected and are reported accordingly. Based on molecular dynamics simulations, we identified regular interactions of the Cys³⁴(-HETEOETE)-moiety with several glutamic acid residues in HSA including Glu⁸⁶, which is not an obvious interaction partner by visual inspection of the HSA crystal structure. The existence of this and other intramolecular cross-links was experimentally proven for the first time.