Mass spectrometry最新文献

Isotope dilution mass spectrometry is a widely used method for measuring intracellular metabolite concentrations, relying on the ratio of peak areas between the target compound and its stable isotope-labeled internal standard. For metabolome analysis of microorganisms, comprehensive concentration measurements have been achieved through the preparation of stable isotope-labeled internal standard extracts (SILIS). Methods have been developed to prepare SILIS by extracting crude metabolites from fully ¹³C-labeled bacteria Escherichia coli and yeasts Saccharomyces cerevisiae and Pichia pastoris (Komagataella phaffii). For cost-effective preparation of SILIS, ideal characteristics of host yeasts include rapid cell growth, high biomass production, and significant metabolite accumulation. In this study, suitable yeast species for SILIS production were investigated from diverse candidates. Batch cultures of 15 yeast species from 12 genera were performed in synthetic defined medium, with cells harvested at different growth phases and metabolites extracted using the methanol/chloroform/water method. Metabolomic analysis by liquid chromatography-tandem mass spectrometry revealed the relative concentrations of 65 metabolites. The results demonstrated that S. cerevisiae and Kluyveromyces marxianus in the stationary phase were the most effective for SILIS production of central metabolic intermediates. SILIS production using S. cerevisiae and K. marxianus can be widely applied in standard laboratories because these species are safe, the media are commercially available, and the extraction methods are easily implementable.

Artificial intelligence (AI) has provided viable methods for retrieving, organizing, and analyzing mass spectrometry (MS) data in various applications. However, several challenges remain as this technique is still in its early, preliminary stages. Critical limitations include the need for more effective methods for identification, quantification, and interpretation to ensure rapid and accurate results. Recently, high-throughput MS data have been leveraged to advance machine learning (ML) techniques, particularly in matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS and MS imaging (MSI). The accuracy of AI models is intricately linked to the sampling techniques used in MALDI and MALDI imaging measurements. With the help of artificial neural networks, traditional barriers are being overcome, accelerating data acquisition for different applications. AI-driven analysis of chemical specificity and spatial mapping in two-dimensional datasets has gained significant attention, highlighting its potential impact. This review focuses on recent AI applications, particularly supervised ML in MALDI-TOF MS and MALDI-MSI data analysis. Additionally, this review provides an overview of sample preparation methods and sampling techniques essential for ensuring high-quality data in deep learning-based models.

Amphibians, as one of the leaders of immune resistance, have lived on Earth for hundreds of millions of years. Their dorsal glands produce a cocktail of biologically active peptides that successfully fight microorganisms and even predators. Since this mechanism prevents the development of pathogen resistance, antimicrobial peptides are very promising pharmaceuticals for future generations. Mass spectrometry is the most powerful tool for sequencing peptides/proteins. For over 30 years of studies in this field, mass spectrometry has resolved all the problems associated with the de novo sequencing of amphibian peptides. This review covers the modern de novo sequencing algorithms that enable achieving complete sequence coverage of all frog peptides, including long ones (up to 46 amino acids). Accurate mass measurements have reliably solved the problem of isobaric amino acids. Moreover, there is no longer any need to carry out any preliminary derivatization procedures such as breaking disulfide bonds or N-terminal acetylation. EThcD and ExD tools with manual spectra interpretation provide an efficient approach for reliable differentiation between isomeric leucine and isoleucine residues in the chain, using secondary w- and d-ions, and they resolve the problems of sequencing inside the intact S-S cycles.

[This corrects the article DOI: 10.5702/massspectrometry.A0145.].

In matrix-assisted laser desorption ionization mass spectrometry, a suitable matrix is often selected for the analyte. Herein, we first developed a novel matrix, alkylated hydroxychalcone (AHC), which has properties similar to alkylated trihydroxyacetophenone (ATHAP) (Anal. Chem., 85: 9444-9448, 2013) developed as a matrix for hydrophobic peptides. However, the sample-to-sample reproducibility was low because of the poor crystallinity of AHC. The crystalline morphology of AHC changed when AHC/2,5-dihydroxybenzoic acid (DHB) was used as a binary matrix. As a result, the use of AHC/DHB improved sample-to-sample reproducibility and increased sensitivity for hydrophobic peptides. Mass imaging indicated that these results were due to an increased number of sweet spots wherein the analytes were detected as ion peaks, in a matrix/analyte crystal spot.

The appearance of the characteristic peak of the hydride-eliminated molecule [M-H]⁺ under a positive ion mode (positive) fast atom bombardment (FAB) ionization condition and liquid-assisted secondary ion mass spectrometry (LSIMS) conditions is known for some compounds and the mechanism of its formation has been investigated. In this study, we investigated the formation mechanism of the hydride-eliminated molecule [M-H]⁺ from 4-substituted-1-(methoxymethyl)benzene under a positive FAB ionization condition. The mass spectra of 4-methoxy-1-(methoxymethyl)benzene (1), 4-methoxy-1-(methoxymethyl-d ₂-)benzene (1-d ₂), and 4-methoxy-1-(methoxymethyl-d ₃)benzene (1-d ₃) were measured under the positive FAB conditions. [M-H]⁺ was observed for 1 and 1-d ₃, and [M-D]⁺ for 1-d ₂, indicating that the site of hydride elimination was the methylene of the 1-(methoxymethyl) moiety. Since [M-H]⁺ was hardly observed under the conditions of positive electron ionization and positive chemical ionization in the gas phase, the hydride elimination is a reaction specific to positive FAB ionization. To examine the contribution of the 4-substituent to the hydride elimination reaction, the mass spectra of (methoxymethyl)benzene (2) and 4-nitro-1-(methoxymethyl)benzene (3) were measured using the positive FAB. The ordering of the relative peak intensity of [M-H]⁺ for [M+H]⁺ in the FAB mass spectra was 1 > 2 > 3, and the results suggest that the electron-donating power of the substituents is an important factor in the formation of [M-H]⁺.

Scorpion venoms contain a variety of peptides that exhibit toxicity toward insects or mammals by acting on ion channels. We previously isolated four insecticidal peptides (Bl-1, Bl-2, Bl-3, and Bl-4) from the venom of Buthacus leptochelys. Among these, the complete amino acid sequence of Bl-1 was determined, whereas only N-terminal partial sequences were obtained for the others. In the present study, we determined the complete sequence of Bl-3 through de novo sequencing of enzymatically digested fragments. The discrimination between Leu and Ile was achieved based on side-chain fragmentation observed under high-energy collision-induced dissociation conditions. Bl-3 was identified as a 65-residue peptide containing four disulfide bonds. During the sequencing analysis, deamidation of the Asn residue at position 30 was observed, which is likely to have occurred after the purification step. Sequence comparison revealed that Bl-3 shares high similarity with α-toxins that act on sodium channels and exhibit nonselective toxicity toward both insects and mammals. These findings suggest that Bl-3 is likely to exert nonselective toxicity through a mechanism similar to that of α-toxins.

Chlorogenic acids, esters of hydroxycinnamic acids with quinic acid, are abundant plant metabolites with over 400 known derivatives. Due to the limited availability of commercial standards, mass spectrometry fragmentation data are essential for structural identification. We acquired fragmentation spectra of six chlorogenic acid homologs in both positive- and negative-ion modes using direct infusion mass spectrometry. In positive-ion mode, sodiated molecules provided additional structural information in addition to that from protonated molecules, although the difference in substitution positions had minimal effects on fragmentation patterns. In negative-ion mode, fragmentation differed significantly depending on the acyl group substitution position on the quinic acid moiety, enabling isomer differentiation. This positional selectivity in negative-ion fragmentation parallels previous observations with anhydrous monosaccharides and oligosaccharides. Comparative analysis with maltotriose and β-glucan trisaccharides demonstrated that negative-ion mode fragmentation yields more diagnostic ring cleavage information for structural characterization. This study also emphasizes that the adoption of unambiguous IUPAC (International Union of Pure and Applied Chemistry)-based nomenclature is fundamental to ensuring the reliability of mass spectra databases.

Pesticide residues in water contamination represent a significant public and political issue due to their harmful effects on the environment, biodiversity, and human health, even at low concentrations. Pesticides are chemically heterogeneous, covering a wide range of LogK _o/w values. Therefore, developing sensitive methods to detect a broad spectrum of hazardous chemicals in aqueous matrices is challenging. Gas and liquid chromatography/high-performance liquid chromatography-mass spectrometry (GC/HPLC-MS) are established tools but typically require pre-concentration steps. Stir bar sorptive extraction (SBSE) is a green, simple, automatable, and HPLC-compatible technique. This study presents a multi-residue method for determining 131 pesticides in mineral water using SBSE followed by HPLC-tandem MS. The selected pesticides, from various chemical classes, were evaluated in fortified ultra-pure and mineral water samples. The method demonstrated excellent sensitivity, with lower limits of quantification ranging from 20 to 50 ng/L for all analytes, enabling detection at trace levels. Selectivity was high (SEL% <20%), and reproducibility was confirmed with RSD% values below 20%. Intra- and interday precision tests revealed RSD% values from 0.23% to 19.81%. Trueness was validated with BIAS% below 20% at all concentrations. Uncertainty values were acceptable, with U% ranging from 1.44% to 49.24%. This SBSE-HPLC-tandem MS method is a robust, efficient, and reliable alternative to traditional approaches for routine monitoring of pesticide residues in drinking water, with quantification limits below regulatory requirements. It offers a suitable tool for public health applications, ensuring reliable pesticide detection in complex water matrices.

Temperature effects on differentiating d-amino acids using the molecular recognition ability of l-tryptophan were investigated by ultraviolet photodissociation spectroscopy in the gas phase. Temperature-dependent ultraviolet photodissociation spectra of hydrogen-bonded protonated clusters of l-tryptophan with arginine, lysine, asparagine, and glutamine enantiomers, generated via electrospray ionization, were obtained using a tandem mass spectrometer containing a variable-temperature ion trap. The spectra at 8 K differed between the amino acids and their enantiomers, indicating that l-tryptophan recognized amino acids and their enantiomers through its hydrogen bonding and electronic structure. The spectral differences observed at 100 K were significantly smaller than those at 8 K. Hot bands and entropic effects at liquid nitrogen cooling temperature prevented the differentiation of d-amino acids. To avoid these contributions in the spectra, cooling of the hydrogen-bonded clusters using a cryogenic refrigerator was necessary to distinguish amino acids and their enantiomers based on the molecular recognition of l-tryptophan.