Liquid Chromatography Orbitrap Mass Spectrometry Research Articles

Monitoring of organophosphate and pyrethroid metabolites in human urine samples by an automated method (TurboFlow™) coupled to ultra-high performance liquid chromatography-Orbitrap mass spectrometry

There is a growing concern to pesticide exposure in humans, being the organophosphate (OP), carbamate and pyrethroid the most commonly used insecticides. Therefore, a fast, simple and automated screening method based on ultra-high performance liquid chromatography coupled to Orbitrap high resolution mass spectrometry (UHPLC-Orbitrap-MS) has been developed to determine six OPs metabolites (3,5,6-trichloro-2-pyridinol “TCPy”, 2-isopropyl-6-methyl-4-pyrimidinol “IMPY”, 4-nitrophenol “PNP”, 4-nitrophenol-d4 “PNP-d4″, 3-chloro-7-hydroxy-4-methylcoumarin “CMHC” and 2-diethylamino-6-methylpyrimidin-4ol “DEAMPY”) and three pyrethroid metabolites (3-phenoxybenzoic acid “3-PBA”, 4-fluoro-3-phenoxybenzoic acid “4-F-3-PBA” and 3-(2,2-dichlorovinyl)-2,2-dimethyl-1-cyclopropanecarboxylic acid “cis/trans-DCCA”) in human urine samples. Off-line solid phase extraction (SPE), on-line SPE and turbulent flow chromatography (TurboFlow™) extraction methodologies were compared, obtaining better results when TurboFlow™ was applied, achieving a total analysis time of 13.83 min. Several validation parameters were tested, obtaining suitable results for all studied compounds. Recoveries ranged from 70 to 116%, meanwhile repeatability and reproducibility were lower than or equal to 7% and 13%, respectively. The limits of quantification (LOQs) were set in the range from 1 μg/L to 5 μg/L (except for DCCA at 10 μg/L). Finally, thirty seven urine samples from women and men living near the agricultural areas of Almeria (Spain) were analyzed to know and control the contamination on the human health. PNP, DEAMPY and IMPY were detected in six urine samples at trace levels except IMPY, which was detected at 3.44 μg/L in one of the samples. Additionally, a post-targeted and untargeted analysis were carried out in the samples, identifying eleven insecticides such as bensulide or cinerin II, one insecticide metabolite, fipronil sulfone, one drug metabolite, carboxyibuprofen and two nicotine metabolites, cotinine and 2-(1-pyrrolidinyl)isonicotinic acid, in several samples.

Lipidomic profiles disturbed by the internet gaming disorder in young Korean males

Internet Gaming Disorder (IGD) is characterized by uncontrollable and persistent playing of internet games despite the occurrence of negative consequences. Although there is a worldwide treatment demand, IGD still doesn't have an explicit biomarker.The primary goal of the study is to characterize lipidomic profiles specific to internet gaming disorder (IGD) based on liquid-chromatography Orbitrap mass-spectrometry (LC Orbitrap MS). Primarily, a total of 19 lipids were significantly dys-regulated in the IGD group compared to healthy controls. The lipidomic feature was mainly characterized by various types of phosphatidylcholines (PCs) and lyso-phosphatidylcholines (LysoPCs). Subsequent multivariate statistical model and linear regression model prioritized two LysoPCs (C16:0 and C18:0) for potential biomarker. Receiver operating characteristic (ROC) analysis demonstrated excellent performance of the combined lipid set for discriminating the IGD group from healthy controls (AUC: 0.981, 95% confidence interval: 0.958–1.000). Additional evaluation with potential confounders and clinical parameters suggested robustness and potential applicability of the outcome as biomarkers which may aid diagnosis.

Highly geographical specificity of metabolomic traits among Korean domestic soybeans (Glycine max)

Classification and characterization of agricultural products at molecular levels are important but often impractical with genotyping, particularly for soybeans that have numerous types of variety and landraces. Alternatively, metabolic signature, a determinant for nutritional value, can be the good molecular indicator, which reflects cultivation region-dependent factors such as climate and soil.Accordingly, we analyzed the integrative metabolic profiles of Korean soybeans cultivated in 7 different provinces (representative production areas), and explored the potential association with geographic traits. A total of 210 primary and secondary metabolites were profiled using gas-chromatography time-of-flight mass spectrometry (GC-TOF MS) and liquid-chromatography Orbitrap mass spectrometry (LC-Orbitrap MS). Despite the partial heterogeneity of the soybean varieties, the metabolomic phenotypic analysis based on multivariate statistics inferred the chemical compositional characteristics was primarily governed by the regional specificity. The OPLS-DA model proposed biomarker cluster re-composed with 5 metabolites (tryptophan, malonylgenistin, malonyldaidzin, N-acetylornithine, and allysine) (AUCs = 0.870–1.0). The most distinctive metabolic profiles were identified with the soybeans of Gunsan (middle-western coast) and Daegu (east-southern inland area), which were best characterized by the highest contents of isoflavones and amino acids, respectively. Further interrogation on geographic data suggested the combinatorial association of region-specific metabolic features with general soil texture and climate traits (total rainfall and average annual temperature).

Analysis of O-Acetylated Sialic Acids in Dried Blood Spots.

Sialic acid is a family of N- and O-substitutions of neuraminic acid. Plasma or serum sialic acid has been established as a potential disease marker. For example, the presence of 9- O-acetyl on the sialic acid of some glycans and glycoconjugates (e.g., 9- O-acetyl GD3 ganglioside) could be related to cancer occurrence. A variety of assays are available to measure serum or plasma sialic acid; however, sample preparation and storage can alter the O-acetylation profile due to the loss of O-acetyl groups and/or the migration of O-acetyl groups. Herein, we report dried blood spot (DBS) sampling, in combination with diamino-4,5-methylenedioxybenzene derivatization, for profiling sialic acids in blood samples with minimal alteration in O-acetylation patterns. The feasibility of the method was first evaluated by analyzing sialic acids in crucian carp blood and comparing with traditional blood/plasma sample preparation procedures. A total of 19 different sialic acids were identified by using liquid chromatography-Orbitrap mass spectrometry, including four mono-O-acetylated N-acetylneuraminic acids, four mono-O-acetylated N-glycolylneuraminic acids, six di-O-acetylated N-acetylneuraminic acids, and three tri-O-acetylated N-acetylneuraminic acids. The long-term storage study indicated that DBS sampling could effectively preserve the O-acetylation information for at least 6 weeks. Thus, it is demonstrated that this method is a valuable tool for the study of sialic acid diversity, especially for the characterization of isomeric structures.

Evaluation of bisphenol A exposure induced oxidative RNA damage by liquid chromatography-mass spectrometry

Highlighted evidence suggests the possible implication of bisphenol A (BPA) exposure on a variety of biological functions, such as DNA damage. Similar to DNA, exposed to BPA may also have potential risks to RNA damage due to its induction of reactive oxygen species. However, there are no related research reports about such health risks of BPA. Therefore, this work tried to investigate the BPA exposure induced oxidative RNA damage by detecting urinary nucleosides, the end-products of RNA metabolism. An ultra-high performance liquid chromatography-Orbitrap mass spectrometry method was applied to selectively and sensitively determine urinary nucleosides. As a result, 66 nucleosides were identified and the effects of BPA exposure on these nucleosides in rat urine samples were evaluated. The nucleosides showed different changing tendency along with different exposure dose of BPA. The strongest effect was observed in high does-exposure rats, indicating dose-response relationship between BPA-treatment and urinary nucleosides. Significant change of some nucleosides, including 8-oxoguanosine, was observed in the high-dose exposure group, suggesting obvious RNA damage to rats. To the best of our knowledge, it is the first study about the RNA damage induced by BPA exposure. The results provided a new perspective on the toxic effects of BPA exposure.

Quantification of known and unknown terpenoid organosulfates in PM10 using untargeted LC–HRMS/MS: contrasting summertime rural Germany and the North China Plain

Environmental contextSecondary organic aerosols account for a major fraction of atmospheric particulate matter, affecting both climate and human health. Organosulfates, abundant compounds in organic aerosols, are difficult to measure because of the lack of authentic standards. Here we quantify terpene-derived organosulfates in atmospheric particulate matter at a rural site in Germany and at the North China Plain using a combined target/non-target high-resolution mass spectrometry approach. AbstractOrganosulfates (OSs) are a ubiquitous class of compounds in atmospheric aerosol particles. However, a detailed quantification of OSs is commonly hampered because of missing authentic standards and the abundance of unknown OSs. Using a combined targeted and untargeted approach of high-resolution liquid chromatography–Orbitrap mass spectrometry (LC–Orbitrap MS), we quantified for the first time the total concentrations of known and unknown monoterpene (MT) and sesquiterpene (SQT) OSs in summertime PM10 particulate matter from field studies in rural Germany (MEL) and the North China Plain (NCP). At each site, we observed more than 50 MT-OSs, 13 of which were detectable at both sites. For both locations, median concentrations of MT-OSs were in the range of 10 to 40ngm−3, to which the 13 common MT-OSs contributed on average >50%. The main contributor to MT-OSs was C9H16O7S (MT-OS 267) with average mass concentrations of 2.23 and 6.38ngm−3 for MEL and NCP respectively. The concentrations of MT-OSs correlated with the concentrations of MT oxidation products only for MEL. For NCP, the low concentrations of MT oxidation products (i.e. typically <1ngm−3) suggested a suppression of carboxylic acid formation under high concentrations of NOx and particulate sulfate. Furthermore, we observed 17 SQT-OSs for the MEL samples, whereas 40 SQT-OSs were detected in the NCP samples. Only five of these SQT-OSs were detectable at both sites. Correspondingly, the total concentrations of SQT-OSs were larger for NCP than for MEL, which suggested large differences in the particle chemistry. In particular, aerosol acidity was found to be a key factor during SQT-OS formation, and was probably not sufficient in the PM10 from MEL.

Development and optimization of confirmatory liquid chromatography—Orbitrap mass spectrometry method for the determination of 17 anticoccidials in poultry and eggs

A new sensitive and selective multi-residue method based on liquid chromatography - Orbitrap high resolution mass spectrometry (LC-Orbitrap-HRMS) was developed and validated for the determination of 17 anticoccidials in poultry and eggs. Instrumental parameters were optimized by the means of statistical experimental designs to improve the sensitivity, precision, and repeatability of the method. Further optimization of auto-tuned MS parameters led to an increase of signal intensity by 10% to 99% for 16 out of 17 analytes. The sample preparation procedure included extraction from muscle tissue and egg samples with acetonitrile, followed by preconcentration, reconstitution, and filtration. Validation was performed according to the Commission Decision 2002/657/EC. The occurrence of anticoccidials in eggs and poultry was assessed by using the developed analytical procedure within the Latvian national monitoring program, revealing quantifiable residues for 6 analytes (marker residue of nicarbazin - 4,4′-dinitrocarbanilide (DNC), salinomycin, narasin, toltrazuril, and its two metabolites).

Determination of steviol glycosides in commercial extracts of Stevia rebaudiana and sweeteners by ultra-high performance liquid chromatography Orbitrap mass spectrometry

Stevia rebaudiana extracts are used as sweeteners in several countries worldwide. Several extracts of diverse composition are available on the market, and their taste depends on the contents of the various steviol glycosides. This study presents an accurate method for the qualitative and quantitative determination of steviol glycosides in 40 Stevia extracts, 7 sweeteners and 3 Stevia-sweetened beverages by a UHPLC coupled to an Orbitrap mass spectrometer. The sub-2 μm amide column provided the separation of all the target analytes in a run time of 30 min with high resolution. The effect of different eluent compositions on the ionisation efficiency of the steviol glycosides was studied. The optimal ionisation conditions were achieved in negative mode using 0.05% formic acid. Under this condition, adducts were not found, [M-H]− were the main ions and the spontaneous loss of a glucose residue at C19 was reduced. The %RSD for intra- and inter-day precision for all eleven analytes varied from 2.1 to 4.2% and 3.0–5.1%, respectively. The recoveries from spiked Stevia extract samples were greater than 95% for all analytes. Rebaudioside A was the most abundant, ranging from 23 to 102%. Nine Stevia extracts and one drink were not compliant with the European Regulation. Isosteviol was under the LOD in all samples and steviol was found in four samples in quantities in the range 0.01–0.03%.

Analysis of calystegines in tomato-based products by liquid chromatography–Orbitrap mass spectrometry

Taking into account the high consumption of tomato and tomato-based products, a study of the occurrence of 7 nortropane alkaloids (calystegines A3, A5, B1, B2, B3, B4 and C1) in this type of matrix has been performed. For that purpose, a simple and fast solid-liquid extraction with methanol/water (50/50, v/v) has been developed, and then liquid chromatography, using a HILIC-A as stationary phase, coupled to high resolution mass spectrometry (LC-HRMS-Orbitrap) has been used for their determination. The developed method was validated and recoveries ranged from 96 to 121%, and relative standard deviations lower than or equal to 16% were obtained. Limits of quantification (LOQ) were established at 0.1 (B4), 0.25 (B3) and 0.5 (A3, A5, B1, B2 and C1) mg/kg. Twelve different samples were analyzed, comprising 4 crushed tomatoes, 4 fried tomatoes and 4 jam tomatoes. Calystegines A3 and B2 were identified in all the analyzed samples, while calystegine B3 was determined in all the fried tomatoes as well as in two crushed tomatoes and one jam tomato at concentrations ranging from 0.4 mg/kg (calystegine B2) to 19.0 mg/kg (calystegine A3). Although calystegine A5 was also detected in all the samples, only 3 of them gave concentrations higher than LOQ. Additionally, calystegine B1 was found in one jam tomato at 1.9 mg/kg.

Enantioselectivity and residue analysis of fipronil in tea (Camellia sinensis) by ultra-performance liquid chromatography Orbitrap mass spectrometry

ABSTRACTThis study aimed to determine the residues of fipronil, metabolites, and enantiomers in tea (Camellia sinensis) during tea planting and green tea manufacture. An AD-RH chiral column was used to separate the fipronil enantiomers. During tea planting, the half-lives of the sum of fipronil and metabolites were similar to those of fipronil, which were 2.37 and 3.88 days for tea shoots and mature leaves, respectively. The residues of fipronil and its metabolites increased 2.3–3.6-fold during green tea manufacture. The values for the processing factors of fipronil and metabolites ranged from 1.0 to 2.1. A slightly significant enantioselectivity of (R)- and (S)-fipronil was observed during tea planting and green tea manufacturing. The residue pattern indicated that fipronil should not be applied in tea gardens due to its long persistence. The maximum residual limits of fipronil and metabolites at 2 μg kg−1 were considered optimal.

A novel liquid chromatography Orbitrap mass spectrometry method with full scan for simultaneous determination of multiple bioactive constituents of Shenkang injection in rat tissues: Application to tissue distribution and pharmacokinetic studies.

Shenkang injection is a traditional Chinese formula with good curative effect on chronic renal failure. In this paper, a novel, rapid and sensitive ultra-high-performance liquid chromatography coupled with Q Exactive hybrid quadrupole Orbitrap high-resolution accurate mass spectrometry was developed and validated for simultaneous determination of seven bioactive constituents of Shenkang injection in rat plasma and tissues after intravenous administration. Acetonitrile was used as a protein precipitation agent in biological samples disposal with carbamazepine as internal standard. The chromatographic separation was carried out on a C18 column with a gradient mobile phase consisting of acetonitrile and water (containing 0.1% formic acid). The MS analysis was performed in the full-scan positive and negative ion mode. The lower limits of quantification for the seven analytes in rat plasma and tissues were 0.1-10 ng/mL. The validated method was successfully applied to tissue distribution and pharmacokinetic studies of Shenkang injection after intravenous administration. The results of the tissue distribution study showed that the high concentrations of seven constituents were primarily in the kidney tract. This is the first report of the application of Q-Orbitrap with full-scan mass spectrometry in tissue distribution and pharmacokinetic studies of Shenkang injection.

MS-Based Metabolomics for the Investigation of Neuro-Metabolic Changes Associated with BDE-47 Exposure in C57BL/6 Mice

Polybrominated diphenyl ethers (PBDE), as one kind of the major persistent organic pollutants (POP), have potential adverse effects on human health. 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) has been identified as one of the dominant PBDE congener in environmental and human samples. In this paper, liquid chromatography–orbitrap mass spectrometry (LC-Orbitrap MS) was applied for the profiling analysis of small metabolites in mice serum and striatum. The metabolic characteristics of adult male C57BL/6 mice exposed to BDE-47 were investigated with a non-targeted metabolomics method. The partial least-squares discriminant analysis (PLS-DA) indicated that the metabolites profile was significantly changed due to the exposure. Fifty-seven differential metabolites that significantly altered in mice serum and striatum were identified through databases searching and authentic standards confirmation. The related metabolic pathways mainly involved purine metabolism, alanine, aspartate and glutamate metabolism, tryptophan metabolism, phenylalanine metabolism, and glutathione metabolism. On the basis of metabolic variations, molecular mechanisms including disturbance of dopaminergic system, neurotransmitters regulation, DNA methylation, and oxidative stress were proposed. The obtained results suggested that LC-Orbitrap MS-based metabolomics had the great potential for molecular understanding of metabolic regulation linked to the exposure of environment pollutants.

Metabolic characterization of (1-(5-fluoropentyl)-1H-indol-3-yl)(4-methyl-1-naphthalenyl)-methanone (MAM-2201) using human liver microsomes and cDNA-overexpressed cytochrome P450 enzymes.

MAM-2201 is a synthetic cannabinoid that is increasingly found in recreational drug abusers and cases of severe intoxication. Thus, characterization of the metabolic pathways of MAM-2201 is necessary to predict individual pharmacokinetics and toxicity differences, and to avoid toxic drug-drug interactions. Collectively, 19 phase 1 metabolites of MAM-2201 were identified using liquid chromatography-Orbitrap mass spectrometry following human liver microsomal incubations in the presence of NADPH: 7 hydroxy-MAM-2201 (M1-M7), 4 dihydroxy-MAM-2201 (M8-M11), dihydrodiol-MAM-2201 (M12), N-(5-hydroxypentyl)-MAM-2201 (M13), hydroxy-M13 (M14), N-dealkyl-MAM-2201 (M15), 2 hydroxy-M15 (M16, M17), MAM-2201 N-pentanoic acid (M18), and hydroxy-M18 (M19). On the basis of intrinsic clearance values in human liver microsomes, hydroxy-MAM-2201 (M1), N-(5-hydroxypentyl)-MAM-2201 (M13), and hydroxy-M13 (M14) were the major metabolites. Based on an enzyme kinetics study using human cDNA-expressed cytochrome P450 (CYP) enzymes and an immunoinhibition study using selective CYP antibodies in human liver microsomes, CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 enzymes were responsible for MAM-2201 metabolism. The CYP3A4 enzyme played a prominent role in MAM-2201 metabolism, and CYP1A2, CYP2B6, CYP2C8, and CYP2C9 enzymes played major roles in the formation of some metabolites. MAM-2201 is extensively metabolized by multiple CYP enzymes, indicating that MAM-2201 and its metabolites should be used as markers of MAM-2201 abuse and toxicity. Graphical abstract In vitro metabolic pathways of MAM-2201 were characterized in human liver microsomes and recombinant CYPs using LC-HRMS analysis. Total 19 phase I metabolites were identified with predominant contribution of CYP3A4.

Direct sample analysis-mass spectrometry vs separation mass spectrometry techniques for the analysis of writing inks

Direct sample analysis (DSA) coupled with high resolution, high mass accuracy time of flight mass spectrometry (TOF-MS) is an emerging technique for the in situ analysis of various substances. DSA is similar to desorption electrospray ionization (DESI) and direct analysis in real time (DART), however, it uses an atmospheric pressure chemical ionization source for ionization of organic compounds. In this study, DSA was applied to the identification of 80 black and blue writing inks from both ballpoint and non-ballpoint pens. Approximately 3mm sections of each writing was placed on the DSA sample holder and analyzed in an automated fashion. The holder can analyze 13 samples in a serial manner. Utilizing a high mass accuracy TOF-MS and using internal calibration, mass accuracy of <6ppm was achieved. Moreover, the results obtained from DSA analysis were compared with the extraction of the ink from the paper and its analysis by gas chromatography–mass spectrometry (GC–MS) and nano ultra-high performance liquid chromatography–Orbitrap mass spectrometry (nLC–MS). Analysis by DSA and LC-MS resulted in the identification of colorants as well as vehicles and additives while analysis by GC–MS mainly resulted in the identification of the non-colorant ingredients. Of the three techniques utilized, DSA provided the greatest number of ink compound identifications and in more samples. However, no single method detected all the components of the ink formulations. For profiling purposes, it is therefore necessary to utilize more than one method, such as the combination of DSA, LC-MS, and GC–MS presented in this study.

Quantitative profiling of bile acids in rat bile using ultrahigh-performance liquid chromatography-orbitrap mass spectrometry: Alteration of the bile acid composition with aging.

Bile acids (BAs) play important roles in physiological functions, including the homeostasis of cholesterol and lipids and as ligands for G protein-coupled receptors (GPCRs). With the increasing importance of BAs, analytical methods for their quantification and screening have been developed. However, due to the diverse range and variety of BAs with different activation potency, a simple, effective, and sensitive method is required to screen BAs for accurate quantification and identification. This paper presents an application of ultrahigh-performance liquid chromatography-orbitrap mass spectrometry (UHPLC-LTQ-Orbitrap MS) for profiling BAs in bile. Using this method, along with the accurate quantification of 19 targeted BAs, 22 unknown BAs were detected and characterized by their fragmentation patterns. The method is beneficial for screening most of the BAs (quantitatively and qualitatively) in rat bile with simple preparation in a single run. The sample dilution ranges of each BA were optimized depending on the concentration of BAs in the bile to obtain good peak separation and accurate data. The method validation was performed successfully using charcoal-treated bile and the intra and inter-day coefficients of variation were less than 20% for all BAs while the recovery were above 88.5% except for the lithocholic acid. The method was applied to profile the age-dependent changes in the contents of rat BAs. Through statistical analysis, we found that as the rats aged, unconjugated BAs and glycine-conjugated BAs decreased or were unaffected, while taurine-conjugated BAs were increased in general. Among the unknown BAs, 5 of the taurine-conjugated BAs increased, while a glycine-conjugated BA decreased, in agreement with the trends of the targeted BAs.

Improving global feature detectabilities through scan range splitting for untargeted metabolomics by high-performance liquid chromatography-Orbitrap mass spectrometry

Untargeted metabolomics aims at obtaining quantitative information on the highest possible number of low-molecular biomolecules present in a biological sample. Rather small changes in mass spectrometric spectrum acquisition parameters may have a significant influence on the detectabilities of metabolites in untargeted global-scale studies by means of high-performance liquid chromatography-mass spectrometry (HPLC-MS). Employing whole cell lysates of human renal proximal tubule cells, we present a systematic global-scale study of the influence of mass spectrometric scan parameters and post-acquisition data treatment on the number and intensity of metabolites detectable in whole cell lysates.Ion transmission and ion collection efficiencies in an Orbitrap-based mass spectrometer basically depend on the m/z range scanned, which, ideally, requires different instrument settings for the respective mass ranges investigated. Therefore, we split a full scan range of m/z 50–1000 relevant for metabolites into two separate segments (m/z 50–200 and m/z 200–1,000), allowing an independent tuning of the ion transmission parameters for both mass ranges. Three different implementations, involving either scanning from m/z 50–1000 in a single scan, or scanning from m/z 50–200 and from m/z 200–1000 in two alternating scans, or performing two separate HPLC-MS runs with m/z 50–200 and m/z 200–1000 scan ranges were critically assessed. The detected features were subjected to rigorous background filtering and quality control in order to obtain reliable metabolite features for subsequent differential quantification.The most efficient approach in terms of feature number, which forms the basis for statistical analysis, identification, and for generating biological hypotheses, was the separate analysis of two different mass ranges. This lead to an increase in the number of detectable metabolite features, especially in the higher mass range (m/z greater than 400), by 2.5 (negative mode) to 6-fold (positive mode) as compared to analysis involving a single scan range. The total number of features confidently detectable was 560 in positive ion mode, and 436 in negative ion mode.

In Vitro Metabolic Pathways of the New Anti-Diabetic Drug Evogliptin in Human Liver Preparations.

Evogliptin ((R)-4-((R)-3-amino-4-(2,4,5-trifluorophenyl)butanoyl)-3-(tert-butoxymethyl)-piperazin-2-one), is a new dipeptidyl peptidase IV inhibitor used for the treatment of type II diabetes mellitus. The in vitro metabolic pathways of evogliptin were identified in human hepatocytes, liver microsomes, and liver S9 fractions using liquid chromatography-Orbitrap mass spectrometry (LC-HRMS). Five metabolites of evogliptin-4-oxoevogliptin (M1), 4(S)-hydroxyevogliptin (M2), 4(R)-hydroxyevogliptin (M3), 4(S)-hydroxyevogliptin glucuronide (M4), and evogliptin N-sulfate (M5)—were identified in human liver preparations by comparison with authentic standards. We characterized the cytochrome P450 (CYP) enzymes responsible for evogliptin hydroxylation to 4(S)-hydroxyevogliptin (M2) and 4(R)-hydroxyevogliptin (M3) and the UGT enzymes responsible for glucuronidation of 4(S)-hydroxyevogliptin (M2) to 4(S)-hydroxy-evogliptin glucuronide (M4). CYP3A4/5 played the major role in the hydroxylation of evogliptin to 4(S)-hydroxyevogliptin (M2) and 4(R)-hydroxyevogliptin (M3). Glucuronidation of 4(S)-hydroxy-evogliptin (M2) to 4(S)-hydroxyevogliptin glucuronide (M4) was catalyzed by the enzymes UGT2B4 and UGT2B7. These results suggest that the interindividual variability in the metabolism of evogliptin in humans is a result of the genetic polymorphism of the CYP and UGT enzymes responsible for evogliptin metabolism.

Detection of N-(1-deoxy-D-fructos-1-yl) Fumonisins B₂ and B₃ in Corn by High-Resolution LC-Orbitrap MS.

The existence of glucose conjugates of fumonisin B2 (FB2) and fumonisin B3 (FB3) in corn powder was confirmed for the first time. These “bound-fumonisins” (FB2 and FB3 bound to glucose) were identified as N-(1-deoxy-d-fructos-1-yl) fumonisin B2 (NDfrc-FB2) and N-(1-deoxy-d-fructos-1-yl) fumonisin B3 (NDfrc-FB3) respectively, based on the accurate mass measurements of characteristic ions and fragmentation patterns using high-resolution liquid chromatography-Orbitrap mass spectrometry (LC-Orbitrap MS) analysis. Treatment on NDfrc-FB2 and NDfrc-FB3 with the o-phthalaldehyde (OPA) reagent also supported that d-glucose binding to FB2 and FB3 molecules occurred to their primary amine residues.

A method for simultaneous determination of 20 Fusarium toxins in cereals by high-resolution liquid chromatography-Orbitrap mass spectrometry with a pentafluorophenyl column.

A high-resolution liquid chromatography-Orbitrap mass spectrometry (LC-Orbitrap MS) method was developed for simultaneous determination of 20 Fusarium toxins (nivalenol, fusarenon-X, deoxynivalenol, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, HT-2 toxin, T-2 toxin, neosolaniol, diacetoxyscirpenol, fumonisin B1, fumonisin B2, fumonisin B3, fumonisin A1, fumonisin A2, fumonisin A3, zearalenone, α-zearalenol, β-zearalenol, α-zearalanol, and β-zearalanol) in cereals. The separation of 20 Fusarium toxins with good peak shapes was achieved using a pentafluorophenyl column, and Orbitrap MS was able to detect accurately from cereal matrix components within ±0.77 ppm. The samples were prepared using a QuEChERS kit for extraction and a multifunctional cartridge for purification. The linearity, repeatability, and recovery of the method were >0.9964, 0.8%–14.7%, and 71%–106%, respectively. Using this method, an analysis of 34 commercially available cereals detected the presence of deoxynivalenol, 15-acetyl deoxynivalenol, fumonisin B1, fumonisin B2, fumonisin B3, fumonisn A1, fumonisin A2, fumonisin A3, and zearalenone in corn samples with high concentration and frequency. Trichothecenes was detected from wheat samples with high frequency; in particular, the concentration of deoxynivalenol was high. Conversely, α-zearalenol, β-zearalenol, α-zearalanol, and β-zearalanol were not detected in any of the samples.

Liquid chromatography Orbitrap mass spectrometry with simultaneous full scan and tandem MS/MS for highly selective pesticide residue analysis.

This paper describes the application of LC/Q-Orbitrap MS for the analysis of pesticide residues in fruit and vegetable commodities. LC/Q-Orbitrap MS working in full scan simultaneously with a single MS/MS scan was used to analyse 139 pesticide residues in QuEChERS extracts of tomato, pepper, orange and green tea. Full scan data were obtained at a resolution of 70,000 whereas MS/MS data were obtained at a resolution of 17,500. Quantitation and detection was carried out using full scan data while MS/MS data were used only for identification. MS/MS scans did not have a negative influence on quantitation under the applied conditions. Some peak area reproducibility problems were the consequence of the low sensitivity for some compounds (aldicarb, chlorpyriphos methyl, fenitrothion and fipronil) under the applied conditions. The relation between the operational parameters (viz. automatic gain control (AGC) target, maximum injection time (IT), underfill ratio, isolation window and apex trigger) and the number of automatically identified compounds was investigated. Mass error and minimal intensity of selected fragment ions were also studied. Various working modes were compared, such as full scan with single MS/MS scan and full scan with multiple MS/MS scans. In both cases, the number of automatically reported pesticides was the same. However full scan with single MS/MS scan ensured more points per peak in full scan mode and better peak area reproducibility. The evaluation of the identification and quantitation capabilities of the instrument was performed through the analysis of 100 real samples. The samples were also analysed by LC-QqQ MS/MS and the results of both analytical systems were compared. The comparison revealed that the two instruments were consistent with each other. They found the same pesticides and neither false positive nor false negatives were reported. Nevertheless the Q-Orbitrap MS allowed one to work in high resolution mass spectrometry, increasing the selectivity and, in full scan mode, permitting the retrospective analysis of the data feature that cannot be achieved with QqQ.