bims-metlip 2022-07-03 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2022–07–03
nineteen papers selected by
Sofia Costa, Matterworks

Deep kernel learning improves molecular fingerprint prediction from tandem mass spectra.
Mitigating Matrix Effects in LC-ESI-MS/MS Analysis of a Urinary Biomarker of Xylenes Exposure.
A Pyridinium Derivatization Reagent for Highly Sensitive Detection of Poly(carboxylic acid)s Using Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry.
A quantitative metabolomics assay targeting 14 intracellular metabolites associated with the methionine transsulfuration pathway using LC-MS/MS in breast cancer cells.
TIMSCONVER: A workflow to convert trapped ion mobility data to open data formats.
High-Coverage Quantitative Metabolomics of Human Urine: Effects of Freeze-Thaw Cycles on the Urine Metabolome and Biomarker Discovery.
A green and rapid analytical method for determination of kitasamycin in animal feedstuffs by ultra-high performance liquid chromatography tandem mass spectrometry.
Approaches to Improve the Quantitation of Oxytocin in Human Serum by Mass Spectrometry.
High-throughput analysis for the simultaneous quantification of nine beta-lactam antibiotics in human plasma by UPC2-MS/MS: Method development, validation, and clinical application.
Development, validation, and clinical application of a rapid UPLC-MS/MS method for detection of colchicine in human whole blood and urine.
A dilute-and-shoot liquid chromatography-tandem mass spectrometry method for urinary 18-hydroxycortisol quantification and its application in establishing reference intervals.
Application of liquid chromatography mass spectrometry-based lipidomics to dairy products research: An emerging modulator of gut microbiota and human metabolic disease risk.
Optimizing accelerated solvent extraction combined with liquid chromatography-Orbitrap mass spectrometry for efficient lipid profile characterization of mozzarella cheese.
A Review of Extraction Methods and Analytical Techniques for Styrene and its Metabolites in Biological Matrices.
Characterization of LC-MS based urine metabolomics in healthy children and adults.
High-Throughput Nano-DESI Mass Spectrometry Imaging of Biological Tissues Using an Integrated Microfluidic Probe.
Rapid and sensitive UHPLC-MS/MS methods for dietary sample analysis of 43 mycotoxins in China total diet study.
Application of LEDA algorithm for the recognition of P-glycoprotein and Carbonic Anhydrase hybrid inhibitors and evaluation of their plasma stability by HPLC-MS/MS analysis.
Profiling of Urine Carbonyl Metabolic Fingerprints in Bladder Cancer Based on Ambient Ionization Mass Spectrometry.

Bioinformatics. 2022 06 24. 38(Suppl 1): i342-i349

Deep kernel learning improves molecular fingerprint prediction from tandem mass spectra.

Kai Dührkop.

MOTIVATION: Untargeted metabolomics experiments rely on spectral libraries for structure annotation, but these libraries are vastly incomplete; in silico methods search in structure databases, allowing us to overcome this limitation. The best-performing in silico methods use machine learning to predict a molecular fingerprint from tandem mass spectra, then use the predicted fingerprint to search in a molecular structure database. Predicted molecular fingerprints are also of great interest for compound class annotation, de novo structure elucidation, and other tasks. So far, kernel support vector machines are the best tool for fingerprint prediction. However, they cannot be trained on all publicly available reference spectra because their training time scales cubically with the number of training data.
RESULTS: We use the Nyström approximation to transform the kernel into a linear feature map. We evaluate two methods that use this feature map as input: a linear support vector machine and a deep neural network (DNN). For evaluation, we use a cross-validated dataset of 156 017 compounds and three independent datasets with 1734 compounds. We show that the combination of kernel method and DNN outperforms the kernel support vector machine, which is the current gold standard, as well as a DNN on tandem mass spectra on all evaluation datasets.
AVAILABILITY AND IMPLEMENTATION: The deep kernel learning method for fingerprint prediction is part of the SIRIUS software, available at https://bio.informatik.uni-jena.de/software/sirius.

DOI: https://doi.org/10.1093/bioinformatics/btac260
J Anal Toxicol. 2022 Jun 29. pii: bkac046. [Epub ahead of print]

Mitigating Matrix Effects in LC-ESI-MS/MS Analysis of a Urinary Biomarker of Xylenes Exposure.

Brett A Bowman, Elizabeth Ejzak, Christopher M Reese, Benjamin C Blount, Deepak Bhandari.

  Liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) with stable isotope labeled internal standards (SIL-IS) is the gold standard for quantitative analysis of drugs and metabolites in complex biological samples. Significant isotopic effects associated with deuterium labeling often causes the deuterated IS to elute at a different retention time from the target analyte, diminishing its capability to compensate for matrix effects. In this study, we systematically compared the analytical performance of deuterated (2H) SIL-IS to non-deuterated (13C and 15N) SIL-ISs for quantifying urinary 2-methylhippuric acid (2MHA) and 4-methylhippuric acid (4MHA), biomarkers of xylenes exposure, with an LC-ESI-MS/MS assay. Analytical method comparison between IS demonstrated a quantitative bias for urinary 2MHA results, with concentrations generated with 2MHA-[2H7] on average 59.2% lower than concentrations generated by 2MHA-[13C6]. Spike accuracy, measured by quantifying analyte-spiked urine matrix and comparing the result to the known spike concentration, determined that 2MHA-[2H7] generated negatively biased urinary results of -38.4% whereas no significant bias was observed for 2MHA-[13C6]. Post-column infusion demonstrated that ion suppression experienced by 2MHA and 2MHA-[13C6] was not equally experienced by 2MHA-[2H7], explaining the negatively biased 2MHA results. Quantitation of urinary 4MHA results between IS exhibited no significant quantitative bias. These results underscore the importance of careful selection of internal standards for targeted quantitative analysis in complex biological samples.

Keywords:  2-methylhippuric acid; 4-methylhippuric acid; accuracy; biomonitoring; deuterated; internal standard; liquid chromatography-mass spectrometry; matrix effects; xylenes

DOI:  https://doi.org/10.1093/jat/bkac046
J Am Soc Mass Spectrom. 2022 Jun 28.

A Pyridinium Derivatization Reagent for Highly Sensitive Detection of Poly(carboxylic acid)s Using Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry.

Shimba Kawasue, Yohei Sakaguchi, Reiko Koga, Hideyuki Yoshida, Hitoshi Nohta.

Short-chain fatty acids are difficult to analyze with high sensitivity using liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) owing to the high polarity of their carboxyl groups. Various derivatization methods have been developed; however, most are effective only for monocarboxylic acids and not for those having multiple carboxyl groups. Therefore, we successfully attempted to synthesize a derivatization reagent that could analyze both mono- and poly(carboxylic acid)s with high sensitivity. We optimized our derivatization reagent by modifying the structure of the reaction site, hydrophobicity of the derivatized compound, and linker structure connecting the reaction site to the permanently charged substructure. The reactivity toward carboxyl groups was improved by employing a piperidine moiety as the reaction site, and the ESI efficiency was improved by the highly hydrophobic and permanently charged triphenylpyridinium group. Furthermore, the incorporation of an alkyl linker enabled polylabeling. When the optimized reagent was applied to mono-, di-, tri-, and tetracarboxylic acids, the ESI efficiency increased with polylabeling; thus, our derivatization reagent outperforms existing derivatization methods and enables the analysis of poly(carboxylic acid)s with high sensitivity. Since this derivatization reagent can be applied to most carboxyl-containing compounds, it can be widely used for lipidomics, proteomics, and metabolomics.

DOI: https://doi.org/10.1021/jasms.2c00086
J Chromatogr B Analyt Technol Biomed Life Sci. 2022 Jun 02. pii: S1570-0232(22)00218-5. [Epub ahead of print]1205 123314

A quantitative metabolomics assay targeting 14 intracellular metabolites associated with the methionine transsulfuration pathway using LC-MS/MS in breast cancer cells.

Zixin Jiao, Zhenyao Lu, Ying Peng, Chen Xu, Yunge Lou, Guangji Wang, Jiye Aa, Yue Zhang.

  The methionine transsulfuration pathway plays an important role in some fundamental biological processes, such as redox and methylation reactions. However, quantitative analysis of the majority of intracellular metabolites is rather challenging. In this study, we developed a simple, fast and reliable method using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the simultaneous detection of 14 methionine-related metabolites, including methionine, S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), homocysteine (HCY), cystathionine (Cysta), cysteine (CYS), glutathione (GSH), dimethylglycine (DMG), betaine, serine, folic acid (FA), dihydrofolic acid (DHF), tetrahydrofolic acid (THF) and 5-methyltetrahydrofolic acid (5-MTHF), in MCF-7 and MDA-MB-231 breast cancer cells. By taking advantage of a surrogate matrix, the linearity, sensitivity, precision, accuracy, stability, matrix effect, recovery, dilution integrity and carryover of the established method were evaluated and validated. This method enabled the precise measurement of methionine-related metabolites both in cells and in the medium and was successfully applied to profile these metabolites involved in the methionine transsulfuration pathway. The data showed that cystine deprivation or excessive supplementation with cystine had a marked impact on methionine metabolism, in addition to its effects on intracellular CYS and GSH levels, indicating that the methionine transsulfuration pathway was dependent on intracellular cystine levels. The established method provides a reliable way to target metabolomics for the quantitative determination of intracellular metabolites in the methionine transsulfuration pathway, which can greatly facilitate the understanding of the mechanisms involved in methylation and redox homeostasis in cellular metabolomic studies.

Keywords:  Breast cancer cells; Cystine; LC–MS/MS; Methionine metabolism; Oxidative stress

DOI:  https://doi.org/10.1016/j.jchromb.2022.123314
Bioinformatics. 2022 Jun 27. pii: btac419. [Epub ahead of print]

TIMSCONVER: A workflow to convert trapped ion mobility data to open data formats.

Gordon T Luu, Michael A Freitas, Itzel Lizama-Chamu, Catherine S McCaughey, Laura M Sanchez, Mingxun Wang.

MOTIVATION: Advances in mass spectrometry have led to the development of mass spectrometers with ion mobility spectrometry (IMS) capabilities and dual source instrumentation, however the current software ecosystem lacks interoperability with downstream data analysis using open-source software and pipelines.
RESULTS: Here, we present TIMSCONVERT, a data conversion high-throughput workflow from timsTOF Pro/fleX mass spectrometer raw data files to mzML and imzML formats that incorporates ion mobility data while maintaining compatibility with data analysis tools. We showcase several examples using data acquired across different experiments and acquisition modalities on the timsTOF fleX MS.
AVAILABILITY: TIMSCONVERT and its documentation can be found at https://github.com/gtluu/timsconvert and is available as a standalone command line interface tool for Windows and Linux, NextFlow workflow, and online in the Global Natural Products Social (GNPS) platform.
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

DOI: https://doi.org/10.1093/bioinformatics/btac419
Anal Chem. 2022 Jun 27.

High-Coverage Quantitative Metabolomics of Human Urine: Effects of Freeze-Thaw Cycles on the Urine Metabolome and Biomarker Discovery.

Deying Chen, Wan Chan, Shuang Zhao, Lanjuan Li, Liang Li.

Urine sample storage after collection at ultra-low-temperature (e.g., -80 °C) is normally required for comparative metabolome analysis of many samples, and therefore, freeze-thaw cycles (FTCs) are unavoidable. However, the reported effects of FTCs on the urine metabolome are controversial. Moreover, there is no report on the study of how urine FTCs affect biomarker discovery. Herein, we present our study of the FTC effects on the urine metabolome and biomarker discovery using a high-coverage quantitative metabolomics platform. Our study involved two centers located in Hangzhou, China, and Edmonton, Canada, to perform metabolome analysis of two separate cohorts of urine samples. The same workflow of sample preparation and dansylation isotope labeling LC-MS was used for in-depth analysis of the amine/phenol submetabolome. The analysis of 320 samples from the Hangzhou cohort consisting of 80 healthy subjects with each urine being subjected to four FTCs resulted in relative quantification of 3682 metabolites with 3307 identified or mass-matched. The analysis of 176 samples from the Edmonton cohort of 44 subjects with four FTCs quantified 3516 metabolites with 3166 identified or mass-matched. Multivariate and univariate analyses indicated that significant variations (fold change ≥ 1.5 with q-value ≤ 0.05) from FTCs were only observed in a very small fraction of the metabolites (<0.3%). Moreover, various metabolites did not show a consistent pattern of concentration changes from one to four FTCs, allowing the use of two separate cohorts of samples to remove these randomly changed metabolites. Three metabolite biomarkers for separating males and females were discovered, and FTC did not influence their discovery.

DOI: https://doi.org/10.1021/acs.analchem.2c01816
J Chromatogr A. 2022 Jun 06. pii: S0021-9673(22)00396-X. [Epub ahead of print]1676 463203

A green and rapid analytical method for determination of kitasamycin in animal feedstuffs by ultra-high performance liquid chromatography tandem mass spectrometry.

Feng Xu, Jiayong Yu, Rongrong Zhang, Zeming Zhang, Aili Sun, Xizhi Shi, Yinliang Wu.

  A rapid, simple, highly efficiency analytical method for detecting kitasamycin A1, A4, A5, and A13 in different feedstuffs was successfully developed by combining enhanced matrix removal (EMR) lipid cartridge and ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). After extraction with acetonitrile, the sample supernatants were directly passed through the EMR lipid cartridge. Then, the cartridge was rinsed and eluted with acetonitrile and methanol, respectively, followed by UHPLC-MS/MS analysis with positive mode using multiple reaction monitoring. Optimized pretreatment procedure without solvent conversion, multiple nitrogen drying steps and activated cartridge before loading, and no significant interference were found during the analysis of different types of animal feedstuffs. Excellent sensitivity (Limit of quantification, LOQ) of kitasamycin A1, A4, A5, and A13 was 1.1-2.0 µg/kg. Satisfactory recoveries of kitasamycin A1, A4, A5, and A13 in different feedstuffs were from 74.0% to 98.8%, with the relative standard deviations (RSDs) below 10.4%, and good linear correlation coefficient (r)>0.9990 in the matrix matched standard curve range of 0.02-50.0 µg/L. Results demonstrated that the developed method exhibited excellent linearity, accuracy, precision, sensitivity, and the feasibility of using this method in kitasamycin determination of animal feedstuffs. The method was evaluated using the greenness analysis method through Eco-Scale assessment tool.

Keywords:  Feedstuff; Kitasamycin; Matrix removal lipid cartridge; Solid phase extraction; UHPLC-MS/MS

DOI:  https://doi.org/10.1016/j.chroma.2022.463203
Front Chem. 2022 ;10 889154

Approaches to Improve the Quantitation of Oxytocin in Human Serum by Mass Spectrometry.

Anke Hering, Beverly Jieu, Alun Jones, Markus Muttenthaler.

  The neuropeptide oxytocin (OT) regulates several peripheral and central functions and is a molecule of interest in psychiatric diseases such as autism spectrum disorder, schizophrenia, anxiety and depression. The study of OT in human serum samples is however hampered by inconsistent sample preparation and analysis as well as low endogenous blood concentration (1-10 pM). This results in varying reports on OT's blood levels and interpretation of OT's role in different (patho)physiological states. Quantitative mass spectrometry (MS) is a highly promising technology to address this problem but still requires large sample volumes to achieve adequate sensitivity and reliability for the quantitation of compounds at low concentrations. We therefore systematically evaluated sample preparation methods for MS to achieve a reliable sample preparation protocol with good peptide recovery, minimal matrix effects and good overall method efficiency in line with FDA guidelines for bioanalytic method development and validation. Additionally, we investigated a strategy to improve the ionization efficiency of OT by adding charged and/or hydrophobic moieties to OT to improve the lower limit of quantitation. Optimized sample preparation in combination with OT modification with a quaternary pyridinium ion improved the sensitivity of OT by ∼40-fold on a tandem triple quadrupole mass spectrometer (API4000 QTRAP), resulting in a lower limit of quantitation of 5 pM in water (linear range 5 pM - 1 mM) and 2 nM in human serum (linear range 2 nM - 1 mM) compared to 200 pM in water and 86 nM in serum with unmodified OT. This approach and protocol provide a solid foundation towards method development for OT quantitation using MS, which should be of high value for fundamental research as well as clinical monitoring of OT upon drug treatments.

Keywords:  LC-MS; analytical method development; neuropeptide; oxytocin; sample preparation

DOI:  https://doi.org/10.3389/fchem.2022.889154
J Pharm Biomed Anal. 2022 Jun 21. pii: S0731-7085(22)00325-9. [Epub ahead of print]219 114904

High-throughput analysis for the simultaneous quantification of nine beta-lactam antibiotics in human plasma by UPC2-MS/MS: Method development, validation, and clinical application.

Soma Bahmany, Alan Abdulla, Tim M J Ewoldt, Philip L Oehlers, Brenda C M de Winter, Birgit C P Koch.

  Quantification of beta-lactam antibiotics can be performed by using liquid chromatography in combination with tandem mass spectrometry (MS/MS) or ultraviolet (UV) detection. Since beta-lactam antibiotics are known as highly polar analytes, using standard reversed phase chromatography will result in very early elution, which is often not desirable. Some retention is preferred to reduce matrix effects, because a high amount of non-retained molecular matrix species elute early from the column. For highly polar analytes, ultra-performance convergence chromatography (UPC2) may be a suitable alternative. This method is based on supercritical fluid chromatography. To our knowledge, we developed the first UPC2-MS/MS method for the determination of amoxicillin, benzylpenicillin, flucloxacillin, piperacillin, cefotaxime, cefuroxime, ceftazidime, imipenem, meropenem, and the free fraction of cefuroxime and flucloxacillin in human plasma. The method was validated according to the Food and Drug Administration guidelines. The method was found linear (r2 >0.990) for all analytes. The inaccuracies and imprecisions were < 15% for all analytes. The matrix effect and recovery were nearly all consistent with coefficient of variation of less than 15% and no significant carryover effect was observed. Furthermore, this method was found to be suitable for daily routine analysis in hospital settings, requiring only 50 µL of plasma. This novel, sensitive, and specific UPC2-MS/MS method demonstrated its value in the analysis of a more than 800 human plasma samples in a clinical trial using simple and fast sample preparation and short analysis run time of only 5 min.

Keywords:  Beta-lactam antibiotics; Tandem mass spectrometry; Therapeutic drug monitoring; Ultra‐performance convergence chromatography

DOI:  https://doi.org/10.1016/j.jpba.2022.114904
Biomed Chromatogr. 2022 Jun 29. e5437

Development, validation, and clinical application of a rapid UPLC-MS/MS method for detection of colchicine in human whole blood and urine.

Chongwen Bi, Ying Gao, Chunlin Li, Yuyang Gao, Kaiwen Wang, Yifan He, Zhengxiang Li, Hengjie Yuan.

  A rapid, simple and economical method has been developed to determine the colchicine in both human whole blood and urine by ultra-performance liquid chromatography-tandem mass spectrometry. Colchicine and isotope-labeled internal standard were extracted from the matrix by liquid-liquid extraction with saturated borax and ethyl acetate, and separated by a reversed-phase chromatography C18 column. Gradient elution was carried out using acetonitrile and water spiked with 0.01% formic acid. Multiple reaction monitoring was performed at positive ion mode. The quantitative transitions were m/z 400.27→310.28 for colchicine and m/z 406.16→313.18 for colchicine-D6. The method has good linearity in the range of 0.5-200 ng/mL for blood and 2-2000 ng/mL for urine. The sensitivity, accuracy and matrix effect were all in line with the guidelines of FDA and EMA. The extraction recovery was above 63.94%. The samples were stable under various storage conditions. Six deuterium-substituted isotopic internal standard was used to demonstrate a different mode of colchicine cleavage from the existing literature. This method has been successfully used in the diagnosis of patients with colchicine poisoning. Blood is recommended as the optimal sample compared with urine.

Keywords:  UPLC-MS/MS; blood and urine; clinical poisoning detection; colchicine; isotopic internal standard

DOI:  https://doi.org/10.1002/bmc.5437
J Clin Lab Anal. 2022 Jul 02. e24580

A dilute-and-shoot liquid chromatography-tandem mass spectrometry method for urinary 18-hydroxycortisol quantification and its application in establishing reference intervals.

Beibei Zhao, Jin Bian, Menghua Rao, Xuhui She, Ying Lou, Jun Cai, Wenjun Ma.

   BACKGROUND: Eighteen-hydroxycortisol (18-OHF) is a potential biomarker for differential diagnosis of the two major primary aldosteronism subtypes, aldosterone-producing adenoma, and idiopathic hyperaldosteronism.
METHODS: Urine samples were processed, and the 18-OHF in urine samples were successfully quantified by in-house established dilute-and-shoot liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. Separation was accomplished on a Sigma Ascentis Express C18 column with a gradient mixture of phase (A) 0.2% formic acid in water and phase (B) 0.2% formic acid in methanol at a flow rate of 0.4 ml/min. Mass spectrometric detection was performed in positive electrospray ionization mode via a mass spectrometer.
RESULTS: The linearity of urinary 18-OHF ranged from 4.28 to 8.77 × 103 nmol/L, with a lower limit of quantification at 4.28 nmol/L. The intra- and inter-precision were both below 3%. The range of analytical recovery was 97.8%-109.2%. The validated dilute-and-shoot LC-MS/MS method was compared with the SPE LC-MS/MS method modified from the one reported in 2013. The results by Passing-Bablok regression analysis and Bland-Altman plotting demonstrated a good agreement between the two methods. The presented method was then applied to establish sex-specific reference intervals from 62 males and 62 females, respectively. The calculated 2.5%-97.5% reference intervals for 24-h urinary 18-OHF were 113-703 nmol/day for males and 71.2-450 nmol/day for females.
CONCLUSION: The presented dilute-and-shoot LC-MS/MS method for 18-OHF quantification showed a good performance in the clinical application. Furthermore, the sex-specific reference intervals for 24-h urinary 18-OHF were first established and quite important for its application in primary aldosteronism subtyping.

Keywords:  18-Hydroxycortisol; dilute-and-shoot LC-MS/MS; primary Aldosteronism; reference interval

DOI:  https://doi.org/10.1002/jcla.24580
Food Res Int. 2022 Jul;pii: S0963-9969(22)00263-0. [Epub ahead of print]157 111206

Application of liquid chromatography mass spectrometry-based lipidomics to dairy products research: An emerging modulator of gut microbiota and human metabolic disease risk.

Wei Jia, Chenna Di, Rong Zhang, Lin Shi.

  Lipids in dairy products, as the crucial components in the most essential biological processes, were among the many nutrients delivered to the infants and adults. The structures of lipids are not only intricate but also varied, which makes the holistic and systematic analyses challenging. In recent years, lipidomics, as the new promoter of lipomics in the omics era, has become one of the fastest growing scientific disciplines in food science research. In addition, lipidomics is also one of the major methods to explore the dynamic changes and chemical compositions of lipids in dairy products in recent years. It is a relatively new frontier of intermolecular interactions research, using advanced liquid chromatography mass spectrometry technology to explore lipids. In this paper, the latest progress of liquid chromatography mass spectrometry based lipidomics was reviewed, with emphasis on the application of lipidomics and metabolic disease risk in dairy research. This includes a detailed workflow for routine lipidomics analysis, as well as examples of applications devoted dairy foods components, quality, safety, and revealing the health benefits of dairy products lipids. Advanced and effective methods of MS promote the in-depth study of gut microbiota and human metabolic disease risk and provide tangible solutions for further research in this field.

Keywords:  Dairy products; Dairy safety; Lipid molecules; Lipidomics; Mass spectrometry

DOI:  https://doi.org/10.1016/j.foodres.2022.111206
Food Chem. 2022 Jun 21. pii: S0308-8146(22)01504-7. [Epub ahead of print]394 133542

Optimizing accelerated solvent extraction combined with liquid chromatography-Orbitrap mass spectrometry for efficient lipid profile characterization of mozzarella cheese.

Annalisa Mentana, Rosalia Zianni, Maria Campaniello, Michele Tomaiuolo, Andrea Chiappinelli, Marco Iammarino, Valeria Nardelli.

  In this study, a novel Accelerated Solvent Extraction (ASE) procedure combined with UHPLC-Q-Orbitrap-MS was developed for detailed untargeted lipid profile of mozzarella cheese. Response Surface Methodology and Pareto front, using a Central Composite Design (CCD), were employed to define the optimised combination of extraction temperature, number of extraction cycles and mix of solvents. LipidSearch™ software was used for a reliable and accurate lipid identification. A total of 13 subclasses, including ceramides, diacylglycerols, triacylglycerols, lysophosphatidylcholines, lysophosphatidylethanolamines, phosphatidylcholines, phosphatidylethanolamines, phosphatidylinositols, sphingomyelins, bismethyl phosphatidic acids, cholesterol ester, zymosterol ester, hexosyl ceramides were measured. The elaboration of the CCD showed that the solvents ratio was the main factor affecting the extraction efficiency. The optimised ASE method, together with the Folch extraction, synergistically contributed to a complete characterization of lipid profile of mozzarella cheese, confirming ASE technique, associated with high resolution mass spectrometry detection, as an efficient tool for Lipidomics in food science.

Keywords:  ASE; Central composite design; Lipidomics; Mozzarella cheese; UHPLC-Q-Orbitrap-MS

DOI:  https://doi.org/10.1016/j.foodchem.2022.133542
Biomed Chromatogr. 2022 Jul 02. e5440

A Review of Extraction Methods and Analytical Techniques for Styrene and its Metabolites in Biological Matrices.

Fatemeh Rahimian, Esmaeel Soleimani.

  We reviewed the toxicokinetics of styrene to introduce reliable surrogates for biological monitoring of styrene workers. Also, extraction techniques and analytical methods for styrene and its metabolites have been discussed. Sample preparation is the main bottleneck of the analytical techniques for styrene and its metabolites. While some microextraction methods have been developed to overcome such drawbacks, some still have limitations such as long extraction time, fiber swelling and breakage, and the cost and the limited lifetime of the fiber. Among all, microextraction by packed sorbents coupled with high performance liquid chromatography with ultraviolet detection (MEPS-HPLC-UV) can be the method of choice for determining styrene metabolites. Few studies investigated unchanged styrene in breath samples. Chemical determination in exhaled breath provides new insights into organ toxicity in workers with inhalation exposures and can be considered as a fascinating tool in risk assessment strategies. Taking blood samples is invasive and less accepted by workers than other samples. In contrast, breath analysis is the most attractive method for workers because breath samples are easy to collect and non-invasive, and does not require worker transfer to health facilities. Therefore, developing selective and sensitive methods for determining styrene in breath samples is recommended for future studies.

Keywords:  breath; mandelic acid; phenylglyoxylic acid; styrene; urine

DOI:  https://doi.org/10.1002/bmc.5440
PeerJ. 2022 ;10 e13545

Characterization of LC-MS based urine metabolomics in healthy children and adults.

Xiaoyan Liu, Xiaoyi Tian, Shi Qinghong, Haidan Sun, Li Jing, Xiaoyue Tang, Zhengguang Guo, Ying Liu, Yan Wang, Jie Ma, Ren Na, Chengyan He, Wenqi Song, Wei Sun.

  Previous studies reported that sex and age could influence urine metabolomics, which should be considered in biomarker discovery. As a consequence, for the baseline of urine metabolomics characteristics, it becomes critical to avoid confounding effects in clinical cohort studies. In this study, we provided a comprehensive lifespan characterization of urine metabolomics in a cohort of 348 healthy children and 315 adults, aged 1 to 78 years, using liquid chromatography coupled with high resolution mass spectrometry. Our results suggest that sex-dependent urine metabolites are much greater in adults than in children. The pantothenate and CoA biosynthesis and alanine metabolism pathways were enriched in early life. Androgen and estrogen metabolism showed high activity during adolescence and youth stages. Pyrimidine metabolism was enriched in the geriatric stage. Based on the above analysis, metabolomic characteristics of each age stage were provided. This work could help us understand the baseline of urine metabolism characteristics and contribute to further studies of clinical disease biomarker discovery.

Keywords:  Adults; Characterization; Children; Metabolomics; Urine

DOI:  https://doi.org/10.7717/peerj.13545
Anal Chem. 2022 Jun 30.

High-Throughput Nano-DESI Mass Spectrometry Imaging of Biological Tissues Using an Integrated Microfluidic Probe.

Xiangtang Li, Hang Hu, Ruichuan Yin, Yingju Li, Xiaofei Sun, Sudhansu K Dey, Julia Laskin.

Nanospray desorption electrospray mass spectrometry imaging (nano-DESI MSI) enables quantitative mapping of hundreds of molecules in biological samples with minimal sample pretreatment. We have recently developed an integrated microfluidic probe (iMFP) for nano-DESI MSI. Herein, we describe an improved design of the iMFP for the high-throughput imaging of tissue sections. We increased the dimensions of the primary and spray channels and optimized the spray voltage and solvent flow rate to obtain a stable operation of the iMFP at both low and high scan rates. We observe that the sensitivity, molecular coverage, and spatial resolution obtained using the iMFP do not change to a significant extent as the scan rate increases. Using a scan rate of 0.4 mm/s, we obtained high-quality images of mouse uterine tissue sections (scan area: 3.2 mm × 2.3 mm) in only 9.5 min and of mouse brain tissue (scan area: 7.0 mm × 5.4 mm) in 21.7 min, which corresponds to a 10-15-fold improvement in the experimental throughput. We have also developed a quantitative metric for evaluating the quality of ion images obtained at different scan rates. Using this metric, we demonstrate that the quality of nano-DESI MSI data does not degrade substantially with an increase in the scan rate. The ability to image biological tissues with high throughput using iMFP-based nano-DESI MSI will substantially speed up tissue mapping efforts.

DOI: https://doi.org/10.1021/acs.analchem.2c01093
J Adv Res. 2022 Jul;pii: S2090-1232(21)00204-6. [Epub ahead of print]39 15-47

Rapid and sensitive UHPLC-MS/MS methods for dietary sample analysis of 43 mycotoxins in China total diet study.

Nannan Qiu, Danlei Sun, Shuang Zhou, Jingguang Li, Yunfeng Zhao, Yongning Wu.

   INTRODUCTION: Mycotoxins are toxic metabolites produced by fungi that commonly contaminate foods. As recommended by the World Health Organization, total diet study (TDS) is the most efficient and effective way to estimate the dietary intakes of certain chemical substances for general populations. It requires sensitive and reliable analytical methods applicable to a wide range of complex food matrices and ready-to-eat dishes.
OBJECTIVES: A novel strategy with high selectivity and sensitivity, incorporating three methods based on ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS), was designed for measuring 43 mycotoxins in dietary samples in a China TDS.
METHODS: The 43 mycotoxins were divided into 3 groups for analysis to achieve better performance. For each group, an UHPLC-MS/MS method was developed to determine the target compounds after clean-up by solid phase extraction. A total of 21 isotope internal standards were employed for accurate quantitation. Method validation in terms of linearity, selectivity, sensitivity, accuracy, and precision was performed for all the 43 mycotoxins in 12 complex food matrices.
RESULTS: The limits of detection (LODs) and limits of quantitation (LOQs) were 0.002-1 ng mL-1 and 0.006-3 ng mL-1, respectively. The method recoveries of the 43 mycotoxins spiked in 12 food categories were in the range of 60.3%-175.9% after internal standard correction, with relative standard deviations (RSDs) below 13.9%. For practical application, this method was utilized for 72 dietary samples collected from 6 provinces in the 6th China TDS. More than 80% of the samples were found contaminated by mycotoxins. DON, SMC, FB1, ZEN, BEA, ENNB1, and ENNB were most detected.
CONCLUSIONS: The proposed methods with high sensitivity, accuracy, and robustness provide powerful tools for multi-mycotoxin monitoring and dietary exposure assessment, allowing 43 mycotoxins, including some emerging mycotoxins, to be accurately investigated in a total diet study for the first time.

Keywords:  Complex food matrices; Determination; Mycotoxins; Total diet study; UHPLC-MS/MS

DOI:  https://doi.org/10.1016/j.jare.2021.10.008
J Pharm Biomed Anal. 2022 Jun 10. pii: S0731-7085(22)00308-9. [Epub ahead of print]219 114887

Application of LEDA algorithm for the recognition of P-glycoprotein and Carbonic Anhydrase hybrid inhibitors and evaluation of their plasma stability by HPLC-MS/MS analysis.

Marco Pallecchi, Marta Menicatti, Laura Braconi, Claudiu T Supuran, Silvia Dei, Elisabetta Teodori, Gianluca Bartolucci.

  Design and synthesis of new candidate drugs produces a large number of compounds that must be qualified and tested to evaluate their characteristics and potential applications. Therefore, many studies will be scheduled and, consequently, it will be necessary to arrange specific, reliable, fast and relatively cheap analytic methods to support this research. The manuscript proposes a new approach in the HPLC-MS/MS analysis by using a sole chromatographic set up, tuned to minimize the run time, without requiring high efficiency or resolution between the analytes. The chromatographic column was used only to avoid or limit the interference of sample matrix towards the analyte ionization process (matrix-effects). Then, the MS/MS properties were explored to solve the signal assignment, by performing a series of energy resolved experiments to optimize the parameters and applying an interesting post-processing data elaboration tool (LEDA). The reliability of the new approach was evaluated in a chemical stability study in PBS and human plasma samples of a series of isomeric compounds P-glycoprotein/Carbonic Anhydrase (P-gp/CA) hybrid inhibitors. The obtained results demonstrated the effectiveness (reliability 97%-100%) of the LEDA algorithm to recognize and to separate the possible isomers present in the samples. The obtained matrix-effects values (ME 96%-106%) established that the chromatographic set up (short column and fast elution gradient) was proper to avoid the matrix interferences, while recovery values (RE 88%-108%) indicate a suitable sample preparation, despite only a protein precipitation was carried out. The quantitative performances of proposed HPLC-MS/MS methods showed an accuracy ranging between 92% and 108% and a precision lower than 13% that allows to be confident on the determination of new P-gp/CA hybrid inhibitors in the degradation study. Therefore, the general procedure proposed was found adequate to study a series of isomeric compounds without their chromatographic separation but only by applying and developing the MS/MS features.

Keywords:  Drug plasma stability; ERMS; Isomer separation; MS/MS speciation; Post-run reprocessing; Tandem mass spectrometry

DOI:  https://doi.org/10.1016/j.jpba.2022.114887
Anal Chem. 2022 Jun 28.

Profiling of Urine Carbonyl Metabolic Fingerprints in Bladder Cancer Based on Ambient Ionization Mass Spectrometry.

Yuze Li, Lixia Jiang, Zhenpeng Wang, Yiran Wang, Xiaohua Cao, Lingwei Meng, Jinghan Fan, Caiqiao Xiong, Zongxiu Nie.

The diagnosis of bladder cancer (BC) is currently based on cystoscopy, which is invasive and expensive. Here, we describe a noninvasive profiling method for carbonyl metabolic fingerprints in BC, which is based on a desorption, separation, and ionization mass spectrometry (DSI-MS) platform with N,N-dimethylethylenediamine (DMED) as a differential labeling reagent. The DSI-MS platform avoids the interferences from intra- and/or intersamples. Additionally, the DMED derivatization increases detection sensitivity and distinguishes carboxyl, aldehyde, and ketone groups in untreated urine samples. Carbonyl metabolic fingerprints of urine from 41 BC patients and 41 controls were portrayed and 9 potential biomarkers were identified. The mechanisms of the regulations of these biomarkers have been tentatively discussed. A logistic regression (LR) machine learning algorithm was applied to discriminate BC from controls, and an accuracy of 85% was achieved. We believe that the method proposed here may pave the way toward the point-of-care diagnosis of BC in a patient-friendly manner.

DOI: https://doi.org/10.1021/acs.analchem.2c01890