Journal of the American Society for Mass Spectrometry最新文献

Ion mobility separates ion in the gas phase based on rotationally averaged cross section, a parameter often correlated with size, providing a versatile measurement strategy when integrated with mass spectrometry. The rapid growth in the field of ion mobility mass spectrometry has been catalyzed by numerous innovative advances in instrumentation that have improved resolution, sensitivity, and the ability to measure collision cross sections. These advances in ion mobility instrumentation and methods have been translated into many applications in the fields of metabolomics, lipidomics, proteomics, and structural biology. This Perspective focuses on developments in ion mobility instrumentation, spanning the impressive capabilities of commercial platforms to customized designs and modifications that establish new benchmarks at the frontiers of ion mobility mass spectrometry.

The effectiveness of collision-induced dissociation (CID) in ion trap mass spectrometry (ITMS) is limited by a low-mass cutoff and weak fragmentation yields. Theoretically, the q value is optimized to balance the fractional product ion mass range with adequate energy deposition to improve fragment ion detection in the CID process; however, many promising technologies still depend on the traditional sinusoidal waveform-driven IT. Additionally, traditional CID-based multistage mass spectrometry (MSⁿ) experiments on ITMS rely on complex and time-consuming “tuning” to optimize CID for a particular ion. The digital ion trap (DIT) has a very promising application field in MSⁿ analysis, because of its many unique features. Herein, we conducted a theoretical and experimental investigation of a developed synchronized reverse scan–CID (SRS-CID) using a digital linear ion trap. Specifically, (1) simulations and experiments demonstrated that in the SRS-CID, ions were sequentially scanned from high to low m/z value via the resonance excitation point (q_excitation), producing multiple fragment ions without the need to know the m/z value or complex radiofrequency (rf) tuning of each product ion. The simulations demonstrated that the heating rate in the SRS-CID could reach 0.022 eV/μs. The experiments demonstrated that the optimal reverse scan speed was −0.053 ns/step. (2) We preliminary increased the period by a fixed value (T_step) to control q_excitation to study the molecule fragmentation approach. Different mass spectra were obtained by controlling t_excitation with a fixed T_step. (3) This paper introduces the phase space method to study the motion trajectories of precursor ions and daughter ions. The calculations used and the entire program were uploaded to GitHub. (4) Changing the duty cycle to advantageously shift q_excitation improved the heating rate (0.033 eV/μs) in SRS-CID. Overall, we demonstrated the effectiveness of the developed SRS-CID technique in fragment ion analysis via theoretical derivation, simulation, and experimentation. Furthermore, DIT mass spectrometry was advantageous in tandem mass spectrometry analysis by facilitating modulation of the driving rf period.

Mass spectrometry imaging (MSI) can be used to survey numerous molecular species from a wide variety of surfaces, including biological tissue sections. Atmospheric-pressure (AP) infrared laser-ablation plasma postionization (IR-PPI) has recently been shown to allow matrix free analysis of small molecules from both fresh frozen and formalin fixed paraffin embedded (FFPE) tissue. Detected ion intensities in IR-PPI as well as other AP inlet modalities such as desorption electrospray ionization (DESI) show a strong dependence on the inlet capillary temperature. In this study, the relationship between detected ion intensity and inlet capillary temperature is evaluated, between room temperature and 650 °C, for analyte pipetted on various substrates, as well as fresh frozen and FFPE tissue, by IR-PPI. Temperature trends for exemplar ions of interest show a variety of dependencies with optimal temperatures observed throughout this temperature range. For example, detection of lactate [M-H]⁻ m/z 89.0244 is optimal at ∼100 °C, glutamine [M-H]⁻ m/z 145.0618 at ∼250 °C, arachidonic acid [M-H]⁻ m/z 303.2324 at ∼150 °C and PI(18:0/20:4) [M-H]⁻ m/z 885.5488 at ∼500 °C. Data reduction and clustering of these data by uniform manifold approximation and projection (UMAP) and k-means provides a summary of all temperature trends within the data and association of different ions with these trends are presented. Finally, the implications of different inlet capillary temperature settings in tissue MSI are demonstrated by comparing detected glucose and lactate ion intensities in response to different inlet temperatures in mouse brain. The choice and control of inlet temperature are shown to be critical variables for the interpretation of biological MSI data in AP modalities.

Metabolites are essential small molecules that are naturally occurring in biological processes as end or intermediate products of various pathways. Matrix-assisted laser desorption/ionization–trapped ion mobility separation–mass spectrometry imaging (MALDI-TIMS-MSI) is an emerging technique that can be used to identify the spatial localization of endogenous compounds on tissue. We evaluated the potential of ammonium fluoride (NH₄F) to enhance the ionization efficiency of metabolites in negative polarity mode when used as a comatrix additive in N-(1-naphthyl)ethylenediamine dihydrochloride (NEDC), 9-aminoacridine (9AA), and 1,5-diaminonaphthalene (DAN) matrices. An extensive list of 234 isotopically labeled metabolites (IROA-IS) was used to establish a quantitative ionization efficiency model with respect to the metabolite chemical structures. In addition, we extended our evaluation to endogenous compounds observed in brain samples collected from male mice. Overall, our study demonstrates that NH₄F improves the sensitivity and ionization efficiency of metabolites and lipids in MALDI-TIMS-MSI. This effect was found to vary depending on the matrix, with the ionization efficiency of the studied metabolites increasing in the order NEDC < 9AA < DAN. The quantitative structure–ionization efficiency relationship model can facilitate the appropriate selection of the matrix in MALDI prior to the analysis of analytes of interest.

Staphylococcal enterotoxins (SEs) make up a superfamily of virulence factors that make Staphylococcus aureus a major cause of food poisoning. The amount of SEs produced by a strain may correlate with its virulence; however, their accurate quantification remains a major challenge. This difficulty arises from two main factors: SEs exhibit emetic activity at nanogram levels, and they are secreted into complex biological matrices during bacterial growth, which typically requires immunoaffinity enrichment before multiplex mass spectrometry (MS) analysis. This study presents an innovative method combining laser-induced dissociation (LID) with mass spectrometry to detect and quantify low-abundance SEs without prior immunoenrichment. To enhance detection specificity based on optical properties, a 473 nm laser was used to selectively fragment chromophore-derivatized cysteine peptides from SEs via LID-MS/MS. The derivatization strategy was first validated on synthetic peptides from five major SEs. Sample preparation was then optimized using purified toxins spiked into biological matrices. The method linearity was assessed by spiking SE synthetic peptides into the matrix across a wide concentration range. Finally, the full analytical protocol was validated by the detection and quantification of endogenous SEs produced by S. aureus strains. This LID-MS/MS approach offers a promising alternative to antibody-based methods for the precise quantification of staphylococcal enterotoxins in complex samples.

Accurate characterization of RNAs and their chemical modifications is critical for understanding RNA biology and post-transcriptional regulation. Mass spectrometry using data-dependent acquisition (DDA) is a crucial tool for identifying oligonucleotides (OGN) in epitranscriptomics. In this study, the key DDA parameters on an Orbitrap Fusion Lumos mass spectrometer were optimized, and an iterative mass exclusion MS/MS acquisition method was developed to enhance the OGN identification. Optimal performance was achieved with full MS resolving power of 120,000 and an MS/MS resolving power of 15,000, top 15 MS/MS scans, and 30% normalized HCD collision energy. Applying these settings to analyze RNase T1 digested E. coli rRNA resulted in the identification of an average of 358 unique OGNs and 58% rRNA sequence coverage. Our findings highlight the importance of tailored DDA parameter optimization and establish a robust workflow for confident OGN identification in MS-based epitranscriptomics.

Nitazenes, a class of new psychoactive substances, have emerged as a significant public health and safety concern due to their widespread abuse. While various detection methods, particularly mass spectrometry, have been developed for these substances, there is limited information regarding their fragmentation pathways and isomeric identification. This knowledge is crucial for drug analysis and forensic toxicology. Among the mass spectrometry techniques, collision-induced dissociation (CID) is commonly used for analyzing the fragmentation of analytes; however, its fragmentation pattern may not be sufficient for complete characterization or differentiation of isomers. Electron-activated dissociation (EAD), a fragmentation technique, provides complementary data to CID by generating distinct fragment ions that aid in the identification and characterization of small molecules. This study aims to characterize and identify 17 kinds of nitazenes using CID and EAD in combination with liquid chromatography time-of-flight mass spectrometry (LC-TOF-MS). The chromatographic and mass spectrometric behaviors of these compounds were examined, and the fragments were analyzed, with a particular focus on the differences between isomers under CID and EAD modes. Notably, EAD generated more detailed fragmentation profiles than CID, revealing unique fragmentation pathways and characteristic fragment ions. In addition, the doubly charged ions of nitazenes were identified in the EAD spectra. Based on the CID and EAD fragmentation pathways of nitazenes, a novel substance was identified in a seized sample. These findings underscore the value of CID and EAD in enhancing forensic toxicology workflows by providing complementary fragmentation data that improve the identification and characterization of novel and unknown compounds.

Careful regulation of monovalent metal ions (M⁺) is necessary to maintain a functional cellular system. Of these ions, appropriate sodium (Na⁺) and potassium (K⁺) concentrations are particularly integral for electrochemical signaling, as well as the secondary transport of nutrients and waste. Dysregulation of M⁺ homeostasis can disrupt these mechanisms, potentially influencing the metabolism of downstream biomolecules such as lipids. Thus, the relationship between M⁺ abundances and related biomolecular distributions must be elucidated to better understand the physiology of healthy and disordered tissues. Traditional techniques for imaging biological metal distributions include SIMS, LA-ICP-MS, and XRF; however, these capabilities are limited to elemental analysis or the analysis of molecular fragments and must be paired with other modalities to visualize distributions of more complex biomolecules within the same or similar samples. Conversely, matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a powerful tool often used for mapping such biomolecular distributions, but current methods are unable to detect metals within tissue. This study illustrates a novel methodology that adds metal detection to the MALDI IMS repertoire through which the simultaneous detection of M⁺ metals and lipids is achievable. Using a robotic sprayer for homogeneous application, on-tissue deposition of the chelator deferiprone (DEF) enables subsequent detection of the ionizable metal-chelator complex by MALDI without hindering lipid detection. Our work provides proof-of-concept data for the simultaneous detection of K⁺, Na⁺, and intact lipids using MALDI IMS.

Perfluorooctanesulfonic acid (PFOS) exists in the environment as a mixture of linear (L-PFOS) and branched (Br-PFOS) isomers. Although it is one of the most frequently detected per- and polyfluoroalkyl substances (PFAS), it is often analyzed and reported as total PFOS with limited attention to the distribution of individual isomers. Here, we used cyclic ion mobility spectrometry (cIMS) with a tunable drift length as an added dimension of separation combined with ultra-high performance liquid chromatography-quadrupole time-of-flight-mass spectrometry (UHPLC-cIMS-qToF-MS) to separate six PFOS isomers. By applying six passes in cIMS, the differences in drift times (DT) and collision cross sections (CCS) allowed us to distinguish L-PFOS from five Br-PFOS isomers. The disubstituted PFOS isomers were not separated from each other because they had the same DTs. Calibration curves prepared with individual isomers revealed that all Br-PFOS had higher ionization efficiencies (more than 2 to 5 times higher) in electrospray MS than L-PFOS, emphasizing the need for isomer-specific analysis for accurate PFOS quantification. The optimized UHPLC-cIMS-qToF-MS method was then used to determine PFOS isomer patterns in influent and effluent wastewater samples, as well as in avian egg yolk samples. The results showed that Br-PFOS isomers dominate in wastewater (more than 50% of total PFOS), while L-PFOS is significantly enriched in egg yolk samples (over 88%). This study highlights the effectiveness of UHPLC-cIMS-qToF-MS for separating and quantifying PFOS isomers in complex matrices and underscores the importance of evaluating the isomer distribution of other PFAS compounds in environmental and biological samples.

Cyclic ion mobility spectrometry (cIMS) provides the potential for high resolution separations of small molecule isomers using multiple passes in a closed-loop geometry. Achieving this potential, however, is limited by the analyte stability in the instrument. Fragile ions are susceptible to dissociation when employing long analysis times required by multipass separations. Correctly identifying the causes of analyte ion loss is critical to facilitating high resolution ion mobility separations. Our previous work with dexamethasone and betamethasone demonstrated that the separation of these two epimers was partially limited by fragmentation over long multipass separations. Further investigations into the cause suggest that most analyte loss occurs because of accumulating many ions in the trap prior to cIMS injection rather than long exposure times to the cIMS region of the instrument. This observation aligns with previous observations of ion activation due to space charge effects in high density trapped ion populations. This work demonstrates the unique aspects of space charge induced fragmentation in cIMS directly resulting from variable pre-cIMS ion accumulation times due to multipath separations while reinforcing the importance of regulating ion accumulation prior to IMS.