Rapid Communications in Mass Spectrometry最新文献

Rationale

Direct analysis in real time (DART) and electrospray ionization (ESI) are commonly used in forensic science and are combined with different activation techniques, which impact the degree and type of fragmentation. Therefore, this study compares collision-induced dissociation (CID), in-source CID (IS-CID), and all ions fragmentation (AIF) activation for nitazene analog identification using the NIST/NIJ Data Interpretation Tool (DIT).

Methods

DART and ESI were used to analyze 18 nitazene analogs on a quadrupole time-of-flight (Q-TOF) mass spectrometer. CID, IS-CID, and AIF were explored with DART, and CID was used for ESI analysis. All activation techniques included the use of low, medium, and high activation energies. The resulting data were compared with the NIST DART-MS Forensics Database using the NIST/NIJ DIT.

Results

The CID spectra were generally less noisy and had fewer ions than the IS-CID or the AIF spectra. The IS-CID spectra generated higher mass-to-charge (m/z) product ions and had fewer characteristic product ions than either CID or AIF. The NIST/NIJ DIT correctly identified 14/18 nitazene analogs when using AIF compared with 13/18 correct identifications using both CID and IS-CID. The ESI-CID data produced 12/18 correctly identified samples.

Conclusions

All three activation techniques performed similarly, based on the NIST/NIJ DIT results. However, IS-CID produced significantly less fragmentation than CID or AIF, indicating that AIF is a suitable screening technique for this instrumental setup. The NIST/NIJ DIT can also be utilized for ESI-CID spectra, although slightly fewer nitazene analogs were correctly identified with ESI-CID than with DART-CID.

Rationale

In January 2024, the IAEA experts meeting endorsed both δ¹³C scales that are currently used within the scientific community: the Vienna Peedee belemnite (VPDB) δ¹³C scale defined by NBS 19 with a value of +1.95‰ exactly, and the VPDB-LSVEC scale defined by NBS 19 and the lithium carbonate LSVEC with a value of −46.60‰ exactly. Following the discovery of the instability of the LSVEC material, several expert laboratories independently proposed replacement reference materials (RMs). This study compares these calcium carbonate RMs IAEA-610, -611, -612, and USGS44 at the highest level of the metrological traceability chain and recommends values that enable users to realize the VPDB and VPDB-LSVEC scales.

Methods

The phosphoric acid reaction that is required to evolve CO₂ from calcium carbonate RMs for isotope analyses is scrutinized by comparing the results of the different apparatuses used in the three participating laboratories. All three laboratories use high-precision dual-inlet isotope ratio mass spectrometers and assess the individual instrument offsets in terms of their effects on interlaboratory comparability of samples.

Results

The reported values for IAEA-610, -611, -612, and USGS44 on the δ¹³C_VPDB scale are −9.114 ± 0.011‰, −30.815 ± 0.011‰, −36.739 ± 0.020‰, and −42.073 ± 0.015‰, respectively. Within their measurement uncertainty they are identical to previously published values. Finally, we provide values on the δ¹³C_VPDB and δ¹³C_VPDB-LSVEC scales for some RMs that are routinely used in elemental analysis–isotope ratio mass spectrometry.

Rationale

Diuretics are banned in sports in- and out-of-competition without applicable threshold or reporting levels, and are classified as S5 Diuretics & Masking Agents by the World Anti-Doping Agency. In 2024, two cases reported by the Beijing Anti-Doping Laboratory involved the simultaneous detection of buthiazide and its degradation product, 4-amino-6-chloro-1,3-benzenedisulphonamide (ACB), in athlete urine samples. Subsequent investigations revealed that certain weight-loss medications contained undeclared buthiazide. While instability of thiazide diuretics such as hydrochlorothiazide and chlorothiazide in urine has been previously reported, the degradation behavior of structurally related compounds, buthiazide, mebutizide, and bemethizide, remains largely unexplored.

Methods

This study systematically evaluated the short-term stability of these three thiazide diuretics in human and artificial urine under various pH conditions (2.6, 5.0, 7.0, and 8.8), temperatures (−20°C, 25°C, 40°C, and 60°C), and light exposures (daylight and UV light). The degradation rate was evaluated using a single-point quantitative method by testing samples at predetermined time points in artificial urine (pH 5.0 and 7.0) at concentrations of 200, 1000, and 2000 ng/mL, and the results were subjected to statistical analysis. Long-term stability was evaluated after 1 week, 1 month, 3 months, and 6 months of storage at −20°C.

Results

Three thiazide diuretics and ACB were detected across all tested conditions, with higher degradation rates observed at higher pH values and temperatures. Single-point quantitative analysis corroborated that degradation was most rapid at 60°C, with mebutizide and bemethizide degrading faster than buthiazide. Long-term storage tests showed that buthiazide remained stable for 1 month at −20°C but partially degraded by 3 months, whereas mebutizide and bemethizide remained stable only for 1 week at low pH. Additionally, repeated freeze–thaw cycles promoted degradation in some samples.

Conclusions

These findings underscore the necessity of carefully controlled storage conditions in routine anti-doping analyses, especially when confirmatory testing of B samples is delayed.

Rationale

Saccharides are often analyzed by matrix-assisted laser desorption/ionization mass spectrometry and electrospray ionization mass spectrometry. In this study, electrospray droplet impact/secondary ion mass spectrometry (EDI/SIMS) was applied to the analysis of saccharides and PEG600 with and without using matrices of alkali halides AX (A: alkali, X: halide).

Methods

1 μL aqueous solutions of 2 × 10⁻³ mol·L⁻¹ analytes M (glucose, sucrose, raffinose or PEG 600) and 2 × 10⁻³ mol·L⁻¹ alkali halide mixture of [Li, Na, K, Rb, Cs]X for X = F, Cl, Br, and I were deposited on the substrate. After drying in vacuum, EDI mass spectra of analytes were measured with and without the AX matrices. The charged water droplets were generated by electrospraying 10⁻² mol⋅L⁻¹ aqueous solution of CF₃COOH.

Results

By the addition of four sets of AX matrices, relative intensities of [M + H]⁺ and [M + A]⁺ in the positive mode, and [M − H]⁻, [M + CF₃COO]⁻, and [M + X]⁻ in the negative mode showed characteristic behavior depending on the combination of A and X. The matrices which gave the strongest ion signals were LiI for [M + Li]⁺, and LiF for [saccharide + CF₃COO]⁻ and [saccharide − H]⁻. CsF gave a relatively strong ion signal for [PEG600 + Cs]⁺.

Conclusion

The high detection sensitivities obtained by LiI for [M + Li]⁺ may be due to the largest bond energies of Li⁺⋅⋅⋅M and the weak lattice energy of LiI. LiF gave good results for the detection of [saccharide − H]⁻ and [saccharide + CF₃COO]⁻ due to the segregation of analytes on the LiF matrix.

Rationale

In mass spectrometry measurements, mass shifts may be inadvertently introduced due to instrumental drift and calibration inaccuracies, potentially compromising the accuracy of subsequent data analysis. This work presents a novel, label-free algorithm to improve relative mass alignment between mass spectra. The warping function is modeled as a natural cubic spline, a suitable model for gradual, nonlinear mass shifts.

Methods

The algorithm is validated on a dataset generated from human glioblastoma multiforme samples using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI) and rapid evaporative ionization mass spectrometry (REIMS), and public datasets generated from MALDI-TOF and desorption electrospray ionization (DESI) Orbitrap instruments.

Results

The algorithm considerably reduces the mass dispersion of the dataset and improves the similarity to a reference mass spectrum for MALDI-TOF, REIMS and DESI-Orbitrap data. It is demonstrated that the proposed method reliably corrects for severe mass shifts. The algorithm is competitive in speed and mass dispersion reduction compared to MSIWarp, a common method to correct for mass misalignment.

Conclusions

This paper presents a novel algorithm to reduce relative mass misalignment for mass spectrometry, validated on an extensive dataset. Thanks to the use of profile data, the peak shapes contribute to the computation of the warping function, and the warping function is approximated in a robust manner. An open-source Python implementation of the proposed methodology will be made accessible on GitHub: https://github.com/VanhemelThomas/psalign.

Rationale

Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba (R. crenulata) is widely acknowledged as a traditional medicine in both Asia and Europe, renowned for its exceptional health-promoting properties. Currently, galloylated polyphenols are distinguished by their high structural similarity and notable isomerism, presenting considerable challenges in their annotation and evaluation. However, there is currently a lack of detailed information on the galloylated polyphenols of R. crenulata.

Methods

We optimized the extraction conditions for galloylated polyphenols based on single-factor and response surface experiments. The galloylated polyphenol components were enriched and isolated via HP-20 macroporous resin column chromatography. To this end, we proposed a comprehensive annotation and evaluation of the galloylated polyphenols in R. crenulata by utilizing an improved precursor ion list (PIL)–data dependent acquisition (DDA)/mass spectrometry (MS) combined with the biogenetic building block strategy, along with the preparation and evaluation of the refined fractions.

Results

An acetone concentration of 65% and a solid–liquid ratio of 1:29 were employed to extract galloylated polyphenols at 50°C for 46 min, which facilitated the maximum extraction of these compounds. The galloylated polyphenols were characteristically enriched in a 30% ethanol elution using an HP-20 macroporous resin column chromatography. Subsequently, we annotated 56 galloylated polyphenols from R. crenulata, including 24 galloyl catechins, 17 gallotannins, 7 gallic acid esters, 4 galloyl salidrosides, 3 gallic acids, and 1 ellagitannin. These galloylated polyphenols and their fine fractions all showed significant nitric oxide inhibitory activity and exhibited potential anti-inflammatory effects.

Conclusion

Based on this study, galloylated polyphenols are demonstrated to be a class of abundant and bioactive constituents in R. crenulata, thereby providing a foundation for further elucidating the material basis of its medicinal effects.

Rationale

Sitravatinib is a receptor tyrosine kinase inhibitor that has been developed for the treatment of advanced nonsmall-cell lung cancer (NSCLC) and urothelial carcinoma. This study aimed at developing an integrated LC–MS/MS and LC–Orbitrap–HRMS platform for evaluating the metabolic stability and profiling the metabolites in rat and human liver microsomes.

Method

Separation was achieved using a Waters ACQUITY BEH C₁₈ column with a gradient of 0.1% formic acid in water and acetonitrile. Detection utilized positive electrospray ionization and multiple reaction monitoring of transitions m/z 630.2 → 555.2 for sitravatinib and m/z 502.5 → 323.2 for the internal standard. Metabolite identification was performed using LC–Orbitrap–HRMS through full-scan MS/dd-MS² and parallel reaction monitoring. The structures of the metabolites were characterized via accurate mass measurement and MS² fragmentation interpretation.

Results

The LC–MS/MS method showed excellent linearity over a concentration range of 1.0–2000 nM, suitable for high-throughput in vitro assays. Sitravatinib demonstrated poor metabolic stability (t_1/2 = 17.47 ± 2.57 min) in rat liver microsomes, with seven metabolites identified. Among these, M3 and M5 were the major metabolites. In contrast, sitravatinib exhibited high metabolic stability in human liver microsomes (t_1/2 = 96.06 ± 12.17 min), with only seven minor metabolites detected.

Conclusions

Key metabolic pathways included O-demethylation, amide formation, N-dealkylation, and oxidative deamination. This study establishes the first integrated LC–MS/MS and HRMS strategy for in vitro metabolic profiling of sitravatinib.

Rationale

Agrochemicals are vital for modern agriculture, ensuring high yields and effective pest control. For high-demand products, impurity profiling is crucial, as even trace contaminants can pose serious health and environmental risks. This study analyzes fipronil impurities using advanced techniques to enhance product safety and minimize environmental contamination.

Methods

Impurities in commercial fipronil were separated using high-performance liquid chromatography with a diode array detector (HPLC-DAD, Agilent 1260) at 220 nm on a Phenomenex Luna C18 column (250 × 4.6 mm, 5 μm). Structural elucidation of impurities was performed using liquid chromatography–high resolution mass spectrometry (LC-HRMS) and tandem mass spectrometry (MS/MS) on an Agilent 6545 Q-TOF system with electrospray ionization. Further, impurities were isolated by preparative HPLC (Gemini-NX-5 C18, 250 × 10 mm, 5 μm) and the structures were confirmed using nuclear magnetic resonance (NMR) spectroscopy.

Results

Three impurities were separated by HPLC-DAD, each impurity exhibiting an area percentage greater than 0.1%. LC-HRMS Negative electrospray ionization (ESI) mass spectra of fipronil and its impurities showed dominant [M-H]⁻ ions, allowing precise determination of their exact masses, elemental compositions, and isotopic patterns. Subsequent LC–MS/MS analysis provided detailed fragmentation data to elucidate the structural features of these impurities. Finally, the proposed structures for the studied compounds were confirmed by NMR spectroscopy.

Conclusion

Comprehensive impurity profiling of the widely used insecticide fipronil is studied which is essential for ensuring safety, efficacy, and regulatory compliance. In this, three impurities were identified and structurally characterized using HPLC, LC-HRMS, LC–MS/MS, and NMR. This analytical strategy effectively confirms product purity and supports quality control and risk assessment in agrochemical formulations.

Trial Registration: CSIR-IICT Communication Number: IICT/Pubs./2025/199