Development of a NiFe2O4 covalent organic framework based magnetic solid-phase extraction approach for specific capture of quinolones in animal innards prior to UHPLC-Q-Orbitrap HRMS detection.

This study aimed to present a selective and effective method for analyzing quinolones (QNs) in food matrix. Herein, a NiFe2O4-based magnetic sodium disulfonate covalent organic framework (NiFe2O4/COF) was prepared using a simple solvent-free grinding method, and was adopted as a selective adsorbent for magnetic solid phase extraction of QNs. Coupled with UHPLC-Q-Orbitrap HRMS, an efficient method for simultaneous detection of 18 kinds of QNs was established. The method exhibited good linearity (0.01-100 ng g-1), high sensitivity (LODs ranging from 0.0011 to 0.0652 ng g-1) and precision (RSDs below 9.5%). Successful extraction of QNs from liver and kidney samples was achieved with satisfactory recoveries ranging from 82.2% to 108.4%. The abundant sulfonate functional groups on NiFe2O4/COF facilitated strong affinity to QNs through electrostatic and hydrogen bonding interactions. The proposed method provides a new idea for the extraction of contaminants with target selectivity.

[Functional peptidomics analysis of Saccharomyces pastorianus protein hydrolysates based on different enzyme treatments].

The aim of this study is to explore differences in the peptidomics of Saccharomyces pastorianus protein hydrolysates treated with different enzymes. Briefly, differences in the peptide fingerprints and active peptides of neutral protease/papain-hydrolyzed S. pastorianus were analyzed using ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) combined with PEAKS Online 1.7 analysis software, Peptide Ranker, and the BIOPEP database. Compared to traditional databases, the PEAKS Online uses de novo sequencing for analysis to obtain oligopeptides smaller than pentapeptides. It provides more comprehensive data of the peptide sample. In this study, enzymatic hydrolysates of S. pastorianus protein were prepared under the optimum conditions of neutral protease and papain respectively. In total, 7221 and 7062 polypeptides were identified in the hydrolysates of neutral protease and papain, respectively; among these polypeptides, 980 were common to the two enzymes. The 6241 and 6082 unique peptides found in the hydrolysates of neutral protease and papain, respectively, indicated that the peptide fingerprints of the two hydrolysates are quite different. Peptide Ranker predicted that 3013 (41.73%) and 3095 (43.83%) peptides were potentially bioactive in the hydrolysates of neutral protease and papain, respectively. According to the BIOPEP database, neutral protease and papain contained 295 and 357 active peptides, respectively; these peptides were mainly composed of angiotensin converting enzyme (ACE) inhibitors and dipeptidyl peptidase IV inhibitors and antioxidant peptides. The number of active peptides in the hydrolysate of papain was higher than that in the hydrolysate of neutral protease, but the total ion intensity of active peptides in the former was lower than that in the latter. This study revealed the influence of protease type on the composition of enzymatic hydrolysates from S. pastorianus protein. The above results provide a reference for the development of functional products of S. pastorianus protein peptides and the high-value utilization of yeast resources.

Ultra-high Performance Liquid Chromatography-Ion Mobility-High-Resolution Mass Spectrometry to Evaluate the Metabolomic Response of Durum Wheat to Sustainable Treatments.

Sustainable agriculture aims at achieving a healthy food production while reducing the use of fertilizers and greenhouse gas emissions using biostimulants and soil amendments. Untargeted metabolomics by ultra-high performance liquid chromatography-ion mobility-high-resolution mass spectrometry, operating in a high-definition MSE mode, was applied to investigate the metabolome of durum wheat in response to sustainable treatments, i.e., the addition of biochar, commercial plant growth promoting microbes, and their combination. Partial least squares-discriminant analysis provided a good discrimination among treatments with sensitivity, specificity, and a non-error rate close to 1. A total of 88 and 45 discriminant compounds having biological, nutritional, and technological implications were tentatively identified in samples grown in 2020 and 2021. The addition of biochar-biostimulants produced the highest up-regulation of lipids and flavonoids, with the glycolipid desaturation being the most impacted pathway, whereas carbohydrates were mostly down-regulated. The findings achieved suggest the safe use of the combined biochar-biostimulant treatment for sustainable wheat cultivation.

A Proteomic Study of the Bioactivity of Annona muricata Leaf Extracts in HT-1080 Fibrosarcoma Cells.

Graviola (Annona muricata) is a tropical plant with many traditional ethnobotanic uses and pharmacologic applications. A metabolomic study of both aqueous and DMSO extracts from Annona muricata leaves recently allowed us to identify dozens of bioactive compounds. In the present study, we use a proteomic approach to detect altered patterns in proteins on both conditioned media and extracts of HT-1080 fibrosarcoma cells under treatment conditions, revealing new potential bioactivities of Annona muricata extracts. Our results reveal the complete sets of deregulated proteins after treatment with aqueous and DMSO extracts from Annona muricata leaves. Functional enrichment analysis of proteomic data suggests deregulation of cell cycle and iron metabolism, which are experimentally validated in vitro. Additional experimental data reveal that DMSO extracts protect HT-1080 fibrosarcoma cells and HMEC-1 endothelial cells from ferroptosis. Data from our proteomic study are available via ProteomeXchange with identifier PXD042354.

[Determination of free amino acids in Eriocheir sinensis by ultra-high performance liquid chromatography-high resolution mass spectrometry].

Eriocheir sinensis is a unique freshwater crab found in China, which is well known for its rich nutrition and sweet and delicious taste. Free amino acids in Eriocheir sinensis are not only important nutrients but also are closely related to their unique taste and aroma. Therefore, the determination of the free amino acid contents in Eriocheir sinensis is of great significance for product quality evaluation, flavor research, authenticity, and origin identification. Herein we proposed an ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS)-based method for the determination of 17 free amino acids in Eriocheir sinensis. First, 5 g of the Eriocheir sinensis sample was weighed into a 50-mL polypropylene centrifuge tube. Then, 10 mL of extraction solvents was added to the centrifuge tube, and the resultant solution was mixed well using a vortex mixer. We compared a variety of solvents and finally selected 5%(v/v) perchloric acid aqueous solution as the optimum extraction solvent. The supernatant was transferred to another polypropylene centrifuge tube after centrifuging at 8000 r/min for 5 min. The extraction procedure was repeated according to the above-mentioned steps, and the extraction solution was combined with the supernatant. The extracts were then adjusted to pH 6.5 with 1 mol/L potassium hydroxide solution, and were diluted to 50 mL with water. After filtering by both qualitative filter paper and a 0.45-µm polyether sulfone syringe filter, the extracts were determined by UHPLC-HRMS. We compared three types of mobile phases and chose 0.1%(v/v) formic acid aqueous solution mixed with acetonitrile as the optimum one. Precise parent ion and ion source parameters were also optimized. The 17 analytes, viz. aspartic acid, threonine, serine, glutamic acid, proline, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, arginine, glycine, alanine, and histidine, were separated on an XDB-C18 column (100 mm×4.6 mm, 1.7 µm) with gradient elution. The amino acids were then detected by HRMS in electrospray ionization and selected ion monitoring modes, and the analytes were quantified using external standards. The instrumental detection limit (IDL) and the instrumental quantification limit (IQL) were 0.3 mg/L and 1.0 mg/L, respectively. The linear correlation coefficients were all above 0.9990 in the concentration range of 10.0-200.0 mg/kg. Three levels of free amino acid standards were spiked into the edible parts of Eriocheir sinensis. The recoveries of the amino acids were between 78.4% and 105.3%. The intra-sample, intra-day, and inter-day repeatabilities were below 4.2%, 5.2%, and 11.4%, respectively, which were within reasonable ranges. Twenty samples of Eriocheir sinensis were tested using the proposed method. Thus, in this study, we developed an alternative method for the determination of free amino acids in Eriocheir sinensis with simple pretreatment, good selectivity, and high accuracy.

Secretory Profile of Adipose-Tissue-Derived Mesenchymal Stem Cells from Cats with Calicivirus-Positive Severe Chronic Gingivostomatitis.

The feline calicivirus (FCV) causes infections in cats all over the world and seems to be related to a broad variety of clinical presentations, such as feline chronic gingivostomatitis (FCGS), a severe oral pathology in cats. Although its etiopathogeny is largely unknown, FCV infection is likely to be a main predisposing factor for developing this pathology. During recent years, new strategies for treating FCGS have been proposed, based on the use of mesenchymal stem cells (MSC) and their regenerative and immunomodulatory properties. The main mechanism of action of MSC seems to be paracrine, due to the secretion of many biomolecules with different biological functions (secretome). Currently, several pathologies in humans have been shown to be related to functional alterations of the patient's MSCs. However, the possible roles that altered MSCs might have in different diseases, including virus-mediated diseases, remain unknown. We have recently demonstrated that the exosomes produced by the adipose-tissue-derived MSCs (fAd-MSCs) from cats suffering from FCV-positive severe and refractory FCGS showed altered protein contents. Based on these findings, the goal of this work was to analyze the proteomic profile of the secretome produced by feline adipose-tissue-derived MSCs (fAd-MSCs) from FCV-positive patients with FCGS, in order to identify differences between them and to increase our knowledge of the etiopathogenesis of this disease. We used high-resolution mass spectrometry and functional enrichment analysis with Gene Ontology to compare the secretomes produced by the fAd-MSCs of healthy and calicivirus-positive FCGS cats. We found that the fAd-MSCs from cats with FCGS had an increased expression of pro-inflammatory cytokines and an altered proteomic profile compared to the secretome produced by cells from healthy cats. These findings help us gain insight on the roles of MSCs and their possible relation to FCGS, and may be useful for selecting specific biomarkers and for identifying new therapeutic targets.

Use of ultra high performance liquid chromatography with high resolution mass spectrometry to analyze urinary metabolome alterations following acute kidney injury in post-cardiac surgery patients.

BACKGROUND: Cardiac surgery-associated acute kidney injury (AKI) can increase the mortality and morbidity, and the incidence of chronic kidney disease, in critically ill survivors. The purpose of this research was to investigate possible links between urinary metabolic changes and cardiac surgery-associated AKI. METHODS: Using ultra-high-performance liquid chromatography coupled with Q-Exactive Orbitrap mass spectrometry, non-targeted metabolomics was performed on urinary samples collected from groups of patients with cardiac surgery-associated AKI at different time points, including Before_AKI (uninjured kidney), AKI_Day1 (injured kidney) and AKI_Day14 (recovered kidney) groups. The data among the three groups were analyzed by combining multivariate and univariate statistical methods, and urine metabolites related to AKI in patients after cardiac surgery were screened. Altered metabolic pathways associated with cardiac surgery-induced AKI were identified by examining the Kyoto Encyclopedia of Genes and Genomes database. RESULTS: The secreted urinary metabolome of the injured kidney can be well separated from the urine metabolomes of uninjured or recovered patients using multivariate and univariate statistical analyses. However, urine samples from the AKI_Day14 and Before_AKI groups cannot be distinguished using either of the two statistical analyses. Nearly 4000 urinary metabolites were identified through bioinformatics methods at Annotation Levels 1-4. Several of these differential metabolites may also perform essential biological functions. Differential analysis of the urinary metabolome among groups was also performed to provide potential prognostic indicators and changes in signalling pathways. Compared with the uninjured kidney group, the patients with cardiac surgery-associated AKI displayed dramatic changes in renal metabolism, including sulphur metabolism and amino acid metabolism. CONCLUSIONS: Urinary metabolite disorder was observed in patients with cardiac surgery-associated AKI due to ischaemia and medical treatment, and the recovered patients' kidneys were able to return to normal. This work provides data on urine metabolite markers and essential resources for further research on AKI.

Non-targeted metabolomics characterization of Annona muricata leaf extracts with anti-angiogenic activity.

The tropical plant Annona muricata has been widely used for traditional ethnobotanic and pharmacologic applications. Extracts from different parts of this plant have been shown to have a wide range of biological activities. In the present study, we carry out a metabolomic study of both aqueous and DMSO extracts from Annona muricata leaves that has allowed us to identify 33 bioactive compounds. Furthermore, we have shown that aqueous extracts are able to inhibit endothelial cell migration and both aqueous and DMSO extracts inhibit the formation of tubule-like structures by endothelial cells cultured on Matrigel. We conclude that extracts of Annona muricata leaves have great potential as anti-angiogenic natural combinations of bioactive compounds.

Altered Proteomic Profile of Adipose Tissue-Derived Mesenchymal Stem Cell Exosomes from Cats with Severe Chronic Gingivostomatitis.

Feline chronic gingivostomatitis (FCGS) is a pathology with a complicated therapeutic approach and with a prevalence between 0.7 and 12%. Although the etiology of the disease is diverse, feline calicivirus infection is known to be a predisposing factor. To date, the available treatment helps in controlling the disease, but cannot always provide a cure, which leads to a high percentage of refractory animals. Mesenchymal stem cells (MSCs) play a pivotal role in the homeostasis and reparation of different tissues and have the ability to modulate the immune system responses. This ability is, in part, due to the capacity of exosomes to play a part in intercellular cell communication. However, the precise role of MSC-derived exosomes and their alterations in immunocompromised pathologies remains unknown, especially in veterinary patients. The goal of this work was to analyze the proteomic profile of feline adipose tissue-derived MSCs (fAd-MSCs) from calicivirus-positive FCGS patients, and to detect possible modifications of the exosomal cargo, to gain better knowledge of the disease's etiopathogenesis. Using high-resolution mass spectrometry and functional enrichment analysis with Gene Ontology, exosomes isolated from the fAd-MSCs of five healthy cats and five calicivirus-positive FCGS patients, were pooled and compared. The results showed that the fAd-MSCs from cats suffering from FCGS not only had a higher exosome production, but also their exosomes showed significant alterations in their proteomic profile. Eight proteins were exclusively found in the exosomes from the FCGS group, and five proteins could only be found in the exosomes from the healthy cats. When comparing the exosomal cargo between the two groups, significant upregulation of 17 and downregulation of 13 proteins were detected in the FCGS group compared to the control group. These findings shed light on new perspectives on the roles of MSCs and their relation to this disease, which may help in identifying new therapeutic targets and selecting specific biomarkers.

Metabolic Fingerprinting of Feces from Calves, Subjected to Gram-Negative Bacterial Endotoxin.

Gram-negative bacteria have a well-known impact on the disease state of neonatal calves and their mortality. This study was the first to implement untargeted metabolomics on calves' fecal samples to unravel the effect of Gram-negative bacterial endotoxin lipopolysaccharide (LPS). In this context, calves were challenged with LPS and administered with fish oil, nanocurcumin, or dexamethasone to evaluate treatment effects. Ultra-high-performance liquid-chromatography high-resolution mass spectrometry (UHPLC-HRMS) was employed to map fecal metabolic fingerprints from the various groups before and after LPS challenge. Based on the generated fingerprints, including 9650 unique feature ions, significant separation according to LPS group was achieved through orthogonal partial least squares discriminant analysis (Q2 of 0.57 and p-value of 0.022), which allowed the selection of 37 metabolites as bacterial endotoxin markers. Tentative identification of these markers suggested that the majority belonged to the subclass of the carboxylic acid derivatives-amino acids, peptides, and analogs-and fatty amides, with these subclasses playing a role in the metabolism of steroids, histidine, glutamate, and folate. Biological interpretations supported the revealed markers' potential to aid in disease diagnosis, whereas beneficial effects were observed following dexamethasone, fish oil, and nanocurcumin treatment.

Analysis of Five Mushroom Toxins in Blood by UPLC-HRMS. / 血液中5ç§è˜‘è‡æ¯’ç´ çš„UPLC-HRMS分析.

OBJECTIVES: To develop a method for the simultaneous and rapid detection of five mushroom toxins (α-amanitin, phallacidin, muscimol, muscarine and psilocin) in blood by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS). METHODS: The blood samples were precipitated with acetonitrile-water solution(Vacetonitril∶Vwater=3∶1) and PAX powder, then separated on ACQUITY Premier C18 column, eluted gradient. Five kinds of mushroom toxins were monitored by FullMS-ddMS2/positive ion scanning mode, and qualitative and quantitative analysis was conducted according to the accurate mass numbers of primary and secondary fragment ions. RESULTS: All the five mushroom toxins had good linearity in their linear range, with a determination coefficient (R2)≥0.99. The detection limit was 0.2-20 ng/mL. The ration limit was 0.5-50 ng/mL. The recoveries of low, medium and high additive levels were 89.6%-101.4%, the relative standard deviation was 1.7%-6.7%, the accuracy was 90.4%-101.3%, the intra-day precision was 0.6%-9.0%, the daytime precision was 1.7%-6.3%, and the matrix effect was 42.2%-129.8%. CONCLUSIONS: The method is simple, rapid, high recovery rate, and could be used for rapid and accurate qualitative screening and quantitative analysis of various mushroom toxins in biological samples at the same time, so as to provide basis for the identification of mushroom poisoning events.

UHPLC-HRMS-Based Untargeted Lipidomics Reveal Mechanism of Antifungal Activity of Carvacrol against Aspergillus flavus.

Aspergillus flavus is a common contaminant in grain, oil and their products. Its metabolite aflatoxin B1 (AFB1) has been proved to be highly carcinogenic. Therefore, it is of great importance to find possible antifungal substances to inhibit the growth and toxin production of Aspergillus flavus. Carvacrol (CV) was reported as a potent antifungal monoterpene derived from plants. In this paper, the antifungal effects and mechanism of CV on Aspergillus flavus were investigated. CV was shown good inhibition on the growth of Aspergillus flavus and the production of AFB1. CV used in concentrations ranging from 0, 50, 100 and 200 µg/mL inhibited the germination of spores, mycelia growth and AFB1 production dose-dependently. To explore the antifungal mechanism of CV on Aspergillus flavus, we also detected the ergosterol content of Aspergillus flavus mycelia, employed Scanning Electron Microscopy (SEM) to observe mycelia morphology and utilized Ultra-High-Performance Liquid Chromatography-High-Resolution Mass Spectrometry (UHPLC-HRMS) to explore the lipidome profiles of Aspergillus flavus. The results showed that the production of ergosterol of mycelia was reduced as the CV treatment concentration increased. SEM photographs demonstrated a rough surface and a reduction in the thickness of hyphae in Aspergillus flavus treated with CV (200 µg/mL). In positive ion mode, 21 lipids of Aspergillus flavus mycelium were downregulated, and 11 lipids were upregulated after treatment with 200-µg/mL CV. In negative ion mode, nine lipids of Aspergillus flavus mycelium were downregulated, and seven lipids upregulated after treatment with 200-µg/mL CV. In addition, the analysis of different lipid metabolic pathways between the control and 200-µg/mL CV-treated groups demonstrated that glycerophospholipid metabolism was the most enriched pathway related to CV treatment.

Analysis of Five Mushroom Toxins in Blood by UPLC-HRMS / 法医学杂志

OBJECTIVES@#To develop a method for the simultaneous and rapid detection of five mushroom toxins (α-amanitin, phallacidin, muscimol, muscarine and psilocin) in blood by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS).@*METHODS@#The blood samples were precipitated with acetonitrile-water solution(Vacetonitril∶Vwater=3∶1) and PAX powder, then separated on ACQUITY Premier C18 column, eluted gradient. Five kinds of mushroom toxins were monitored by FullMS-ddMS2/positive ion scanning mode, and qualitative and quantitative analysis was conducted according to the accurate mass numbers of primary and secondary fragment ions.@*RESULTS@#All the five mushroom toxins had good linearity in their linear range, with a determination coefficient (R2)≥0.99. The detection limit was 0.2-20 ng/mL. The ration limit was 0.5-50 ng/mL. The recoveries of low, medium and high additive levels were 89.6%-101.4%, the relative standard deviation was 1.7%-6.7%, the accuracy was 90.4%-101.3%, the intra-day precision was 0.6%-9.0%, the daytime precision was 1.7%-6.3%, and the matrix effect was 42.2%-129.8%.@*CONCLUSIONS@#The method is simple, rapid, high recovery rate, and could be used for rapid and accurate qualitative screening and quantitative analysis of various mushroom toxins in biological samples at the same time, so as to provide basis for the identification of mushroom poisoning events.

Rapid LA-REIMS and comprehensive UHPLC-HRMS for metabolic phenotyping of feces.

Ambient ionization-based techniques hold great potential for rapid point-of-care applicable metabolic fingerprinting of tissue and fluids. Hereby, feces represents a unique biospecimen as it integrates the complex interactions between the diet, gut microbiome and host, and is therefore ideally suited to study the involvement of the diet-gut microbiome axis in metabolic diseases and their treatments at a molecular level. We present a new method for rapid (<10 s) metabolic fingerprinting of feces, i.e. laser-assisted rapid evaporative ionization mass spectrometry (LA-REIMS) with an Nd:YAG laser (2940 nm) and quadrupole Time-of-Flight mass spectrometer as main components. The LA-REIMS method was implemented on mimicked crude feces samples from individuals that were assigned a state of type 2 diabetes or euglycaemia. Based on the generated fingerprints, enclosing 4923 feature ions, significant segregation according to disease classification was achieved through orthogonal partial least squares discriminant analysis (Q2(Y) of 0.734 and p-value of 1.93e-17) and endorsed by a general classification accuracy of 90.5%. A comparison between the discriminative performance of the novel LA-REIMS and our established ultra-high performance liquid-chromatography high-resolution MS (UHPLC-HRMS) metabolomics and lipidomics methodologies for fingerprinting of stool was performed. Based on the supervised modelling results upon UHPLC-HRMS (Q2(Y) ≥ 0.655 and p-value ≤ 4.11 e-5), equivalent or better discriminative performance of LA-REIMS fingerprinting was concluded. Eventually, comprehensive UHPLC-HRMS was employed to assess metabolic alterations as observed for the defined classes, whereby metformin treatment of the type 2 diabetes patients was considered a relevant study factor to acquire new mechanistic insights. More specifically, ten metabolization products of metformin were identified, with (hydroxylated) triazepinone and metformin-cholesterol reported for the first time in vivo.In conclusion, LA-REIMS was established as an expedient strategy for rapid metabolic fingerprinting of feces, whereby potential implementations may relate, but are not limited to differential diagnosis and treatment efficacy evaluation of metabolic diseases. Yet, LC-HRMS remains essential for in-depth biological interpretation.

Pigment profiles of Spanish extra virgin olive oils by ultra-high-performance liquid chromatography coupled to high-resolution mass spectrometry.

This work studies the natural pigment profiles (chlorophylls and carotenoids) of Spanish Extra Virgin Olive Oils (EVOO) produced in different Spanish regions. The simultaneous qualitative and quantitative analysis of EVOO natural pigments has been performed by ultra-high-performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS) using atmospheric pressure chemical ionisation (APCI). The results showed a similar natural pigment pattern for all the analysed EVOOs, although the total pigments content differed significantly. Moreover, the chlorophyll/carotenoid ratio was close to 1, while the lutein/ß-carotene ratio was higher than 1, showing that lutein is the most abundant carotenoid in the studied Spanish EVOOs. Data from multivariate statistical approach demonstrated that the olive variety does not discriminate between EVOO samples. However, they were classified based on their origin allowing good differentiation of samples from the Basque Country and Canary Islands from the rest of regions. The results of this study show the differences of the nature and pigments concentration of Spanish EVOO samples, parameters that are of significance for reliable characterisation.

Impact of storage conditions on the human stool metabolome and lipidome: Preserving the most accurate fingerprint.

Faecal metabolomics markedly emerged in clinical as well as analytical chemistry through the unveiling of aberrations in metabolic signatures as reflection of variance in gut (patho)physiology and beyond. Logistic hurdles, however, hinder the analysis of stool samples immediately following collection, inferring the need of biobanking. Yet, the optimum way of storing stool material remains to be determined, in order to conserve an accurate snapshot of the metabolome and circumvent artifacts regarding the disease and parameter(s) under observation. To address this problem, this study scrutinised the impact of freeze-thaw cycling, storage duration, temperature and aerobicity, thereby using ultra-high performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS)-based polar metabolomics and lipidomics methodologies for faecal metabolomics. Both targeted (n > 400) and untargeted approaches were implemented to assess storage effects on individual chemical classes of metabolites as well as the faecal fingerprint. In general, recommendations are that intact stool samples should be divided into aliquots, lyophilised and stored at -80 °C for a period no longer than 18 weeks, and avoiding any freeze-thawing. The first preservation week exerted the most decisive impact regarding storage temperature, i.e. 12.1% and 6.4% of the polar metabolome experienced a shift at -20 °C and at -80 °C, respectively, whereas 8.6% and 7.9% was observed to be changed significantly for the lipidome. In addition, aside from the negligible impact of aerobicity, the polar metabolome appeared to be more dependent on the storage conditions applied compared to the lipidome, which emerged as the more stable fraction when assessing the storage duration for 25 weeks. If the interest would greatly align with particular chemical classes, such as branched-chain amino acids or short-chain fatty acids, specific storage duration recommendations are reported. The provided insights on the stability of the faecal metabolome may contribute to a more reasoned design of experiments in biomarker detection or pathway elucidation within the field of faecal metabolomics.

Target analysis and retrospective screening of mycotoxins and pharmacologically active substances in milk using an ultra-high-performance liquid chromatography/high-resolution mass spectrometry approach.

Milk is a nutritious food suitable for infants and adults, and it plays an important role in the human diet. However, it may also be a vehicle for food contaminants. In this report, we developed a method using ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-Q-Exactive Orbitrap HRMS; Thermo Fisher Scientific, Waltham, MA) for simultaneous identification of target pharmacologically active substances and mycotoxins in milk. We also used the Q-Orbitrap operating in full scan mode to identify other possible drugs and microbial metabolites that occurred in samples. Fifty-six commercially available milk samples from the Italian market were analyzed. Investigated analytes were extracted using a QuEChERS (quick, easy, cheap, effective, rugged, and safe) approach. Method detection and quantification limits and performance criteria set by European regulations were fulfilled. Pharmacologically active substances were detected in 49% of samples (range 0.007-4.53 ng/mL), including nontarget mycotoxins. Retrospective analysis allowed us to identify other antibiotics and pharmacologically active substances, as well as nonregulated fungal/bacterial metabolites at a relatively high incidence. From the obtained values, the need for continuous monitoring of contaminants in the milk production chain is clear. This is the first study to assess the presence of pharmacologically active substances, mycotoxins, and other microbial metabolites in Italian milk samples using the UHPLC-Q-Orbitrap HRMS system.

Metabolomics data fusion between near infrared spectroscopy and high-resolution mass spectrometry: A synergetic approach to boost performance or induce confusion.

In general, data fusion can improve the classification performance of the model, but little attention is paid to the influence of the data fusion on the spatial distribution of the modeling samples. In this paper, the effect of data fusion on sample spatial distribution was studied through integrating NIR data and UHPLC-HRMS data for sulfur-fumigated Chinese herb medicine. Twelve samples collected from four different geographical origins were sulfur fumigated in the lab, and then metabolomics analysis was conducted using NIR and UHPLC-LTQ-Orbitrap mass spectrometer. First of all, the discriminating power of each technique was respectively examined based on PCA analysis. Secondly, combining NIR and UHPLC-HRMS data sets together with or without variable selection was parallelly compared. The results demonstrated that the discriminable ability was remarkably improved after data fusion, indicating data fusion could visualize variable selection and enhance group separation. Samples in the margin between two classes of samples may increase the experience error but has positive effect on the separation direction. Besides, an interesting feature extraction could obtain better discriminable effect than common data fusion. This study firstly provided a new path to employ a comprehensive analytical approach for discriminating SF Chinese herb medicines to simultaneously benefit from the advantages of several technologies.

Analysis of Common Herbicides in Blood by UPLC-HRMS.

OBJECTIVES: To develop a method to screen and quantify 10 common herbicides （paraquat, diquat, glyphosate, glufosinate, cyanazine, atrazine, metazachlor, acetochlor, chlorsulfuron, and metsulfuron） in blood. METHODS: With acetonitrile-water solution [V（acetonitrile）∶V（water）=3∶1] as protein precipitant, 10 common herbicides in blood were detected using ultra-high performance liquid chromatography-high resolution mass spectrometry （UPLC-HRMS）. RESULTS: All the 10 herbicides had good linearity in their linear range （coefficient of determination R2≥0.993）, with the recovery rates 67.4%-111.9%, the relative standard deviations 1.5%-10.8%, the accuracies 85.1%-106.1%, intra-day precisions 2.7%-13.5%, and inter-day precisions 3.3%-13.3%. CONCLUSIONS: This method is easy to operate with high recovery rates. It enables rapid and accurate qualitative screening and quantitative analysis of various herbicides in blood simultaneously.

Analysis of Common Herbicides in Blood by UPLC-HRMS / 法医学杂志

Objective To develop a method to screen and quantify 10 common herbicides (paraquat, diquat, glyphosate, glufosinate, cyanazine, atrazine, metazachlor, acetochlor, chlorsulfuron, and metsulfuron) in blood.Methods With acetonitrile-water solution[V (acetonitrile) ∶V (water) =3∶1]as protein precipitant, 10 common herbicides in blood were detected using ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS).Results All the 10 herbicides had good linearity in their linear range (coefficient of determination R2≥0.993), with the recovery rates 67.4％-111.9％, the relative standard deviations 1.5％-10.8％, the accuracies 85.1％-106.1％, intra-day precisions 2.7％-13.5％, and inter-day precisions 3.3％-13.3％.Conclusion This method is easy to operate with high recovery rates.It enables rapid and accurate qualitative screening and quantitative analysis of various herbicides in blood simultaneously.