Tomas Cajka

MS-DIAL: data-independent MS/MS deconvolution for comprehensive metabolome analysis

Data-independent acquisition (DIA) in liquid chromatography (LC) coupled to tandem mass spectrometry (MS/MS) provides comprehensive untargeted acquisition of molecular data. We provide an open-source software pipeline, which we call MS-DIAL, for DIA-based identification and quantification of small molecules by mass spectral deconvolution. For a reversed-phase LC-MS/MS analysis of nine algal strains, MS-DIAL using an enriched LipidBlast library identified 1,023 lipid compounds, highlighting the chemotaxonomic relationships between the algal strains.

Ambient mass spectrometry employing direct analysis in real time (DART) ion source for olive oil quality and authenticity assessment.

A novel approach for the authentication of olive oil samples representing different quality grades has been developed. A new type of ion source, direct analysis in real time (DART), coupled to a high-resolution time-of-flight mass spectrometer (TOFMS) was employed for the comprehensive profiling of triacylglycerols (TAGs) and/or polar compounds extracted with a methanol-water mixture. The main parameters influencing the ionization efficiency of TAGs were the type of sample solvent, degree of sample dilution, ion beam temperature, and presence of a dopant (ammonia vapors). The ionization yield of polar compounds depended mainly on a content of water in the extract and ion beam temperature. Using DART-TOFMS, not only differentiation among extra virgin olive oil (EVOO), olive pomace oil (OPO) and olive oil (OO) could be easily achieved, but also EVOO adulteration with commonly used adulterant, hazelnut oil (HO), was feasible. Based on the linear discriminant analysis (LDA), the introduced method allowed detection of HO addition of 6 and 15% (v/v) when assessing DART-TOFMS mass profiles of polar compounds and TAGs, respectively.

Comprehensive analysis of lipids in biological systems by liquid chromatography-mass spectrometry

Liquid chromatography-mass spectrometry (LC-MS)-based lipidomics has been a subject of dramatic developments over the past decade. This review focuses on state of the art in LC-MS-based lipidomics, covering all the steps of global lipidomic profiling. On the basis of review of 185 original papers and application notes, we can conclude that typical LC-MS-based lipidomics methods involve: (1) extraction using chloroform/MeOH or MTBE/MeOH protocols, both with addition of internal standards covering each lipid class; (2) separation of lipids using short microbore columns with sub-2-μm or 2.6-2.8-μm (fused-core) particle size with C18 or C8 sorbent with analysis time <30 min; (3) electrospray ionization in positive- and negative-ion modes with full spectra acquisition using high-resolution MS with capability to MS/MS. Phospholipids (phosphatidylcholines, phosphatidylethanolamines, phosphatidylinositols, phosphatidylserines, phosphatidylglycerols) followed by sphingomyelins, di- and tri-acylglycerols, and ceramides were the most frequently targeted lipid species.

Toward Merging Untargeted and Targeted Methods in Mass Spectrometry-Based Metabolomics and Lipidomics

in Mass Spectrometry-Based Metabolomics and Lipidomics Tomas Cajka† and Oliver Fiehn*,†,‡ †UC Davis Genome Center−Metabolomics, University of California Davis, 451 Health Sciences Drive, Davis, California 95616, United States ‡King Abdulaziz University, Faculty of Science, Biochemistry Department, P.O. Box 80203, Jeddah 21589, Saudi Arabia ■ CONTENTS Sample Extraction 525 Extraction of Polar Metabolites (Metabolomics) 525 Extraction of Lipids (Lipidomics) 527 Combined Extraction of Amphiphilic and Lipophilic Metabolites 527 Mass Spectrometry-Based Metabolomics and Lipidomics 528 Direct Infusion MS 528 Ion Mobility-Mass Spectrometry (IM-MS) 529 Liquid Chromatography−Mass Spectrometry (LC−MS) 533 Reversed-Phase Liquid Chromatography (RPLC) 533 Hydrophilic Interaction Chromatography (HILIC) 534 Normal-Phase Liquid Chromatography (NPLC) 535 Supercritical Fluid Chromatography (SFC) 535 Two-Dimensional Liquid Chromatography (2D-LC) 535 Mass Spectrometric Detection 536 Data Processing 540 Quality Control 541 Conclusions 541 Author Information 542 Corresponding Author 542 Notes 542 Biographies 542 Acknowledgments 542 References 542

Evaluation of two-dimensional gas chromatography–time-of-flight mass spectrometry for the determination of multiple pesticide residues in fruit

In recent years, comprehensive two-dimensional gas chromatography (GC x GC) has attained increasing attention for its outstanding separation potential and capability to solve demanding analytical tasks. Trace level analysis of pesticides residues in complex food matrices represents such a demanding task. For some commodities, such as baby food, the requirements on method detection limits are very strict and the unambiguous confirmation of the pesticide presence based on mass spectrometric detection is required. In this work, GC x GC coupled to time-of-flight mass spectrometry (TOF MS) has been evaluated for the determination of pesticides residues in fruit samples. Twenty modern pesticides with a broad range of physico-chemical properties were analysed in apple and peach samples. It has been demonstrated that the application of comprehensive two-dimensional gas chromatography brings distinct advantages such as enhanced separation of target pesticides from matrix co-extracts as well as their improved detectability. The limits of detection of the pesticides comprised in the study (determined at S/N = 5) ranged from 0.2 to 30 pg, injected with the exception of the last eluted deltamethrin, for which 100 pg could be detected. When compared to one-dimentional GC-TOF MS analysis under essentially the same conditions the detectability enhancement was 1.5-50-fold. Full mass spectral information by time-of-flight mass spectrometry and the deconvolution capability of the dedicated software allowed for reliable identification of most pesticides at levels below 0.01 mg/kg (< 10 pg injected) in fruit. Performance characteristics of the GC x GC-TOF MS method, such as linearity of calibration curves, repeatability of (summed) peak areas, as well as repeatability of first and second dimension retention times, were shown to fully satisfy the requirements for trace level analysis of the pesticide residues in food.

Streamlining sample preparation and gas chromatography–tandem mass spectrometry analysis of multiple pesticide residues in tea

In this work, a new rapid method for the determination of 135 pesticide residues in green and black dry tea leaves and stalks employing gas chromatography coupled to tandem mass spectrometry (GC-MS/MS) with a triple quadrupole was developed and validated. A substantial simplification of sample processing prior to the quantification step was achieved: after addition of water to a homogenised sample, transfer of analytes into an acetonitrile layer was aided by the addition of inorganic salts. Bulk co-extracts, contained in the crude organic extract obtained by partition, were subsequently removed by liquid-liquid extraction using hexane with the assistance of added 20% (w/w) aqueous NaCl solution. The importance of matrix hydration prior to the extraction for achieving good recoveries was demonstrated on tea samples with incurred pesticide residues. For most of the analytes, recoveries in the acceptable range of 70-120% and repeatabilities (relative standard deviations, RSDs) ≤20% were achieved for both matrices at spiking levels of 0.01, 0.1 and 1 mg kg(-1). Under optimised GC-MS/MS conditions, most of the analytes gave lowest calibration level ≤0.01 mg kg(-1), permitting the control at the maximum residue levels (MRLs) laid down in Regulation (EC) No 396/2005. The developed method was successfully applied to the determination of pesticide residues in real tea samples.

Analysis of multiple mycotoxins in beer employing (ultra)-high-resolution mass spectrometry

The objective of the presented study was to develop and optimize a simple, high-throughput method for the control of 32 mycotoxins (Fusarium and Alternaria toxins, aflatoxins, ergot alkaloids, ochratoxins, and sterigmatocystin) in beer. Due to the broad range of their physicochemical properties, the sample preparation step was simplified as much as possible to avoid analyte losses. The addition of acetonitrile to beer samples enabled precipitation of abundant matrix components. The clean-up efficiency was controlled by ambient mass spectrometry employing a direct analysis in real time (DART) ion source. For determination of analytes, ultra-high-performance liquid chromatography hyphenated with high-resolution mass spectrometry utilizing an orbitrap (U-HPLC-orbitrapMS) or time-of-flight (TOFMS) technology was used. Because of significantly better detection capabilities of the orbitrap technology, the U-HPLC-orbitrapMS method was chosen as a determinative step and fully validated. To compensate matrix effects, matrix-matched calibration was employed. The lowest calibration levels for most of the target mycotoxins ranged from 1 to 8  µg  L(-1) beer and the recoveries of analytes were in range from 86 to 124%.

Simplified and rapid determination of polychlorinated biphenyls, polybrominated diphenyl ethers, and polycyclic aromatic hydrocarbons in fish and shrimps integrated into a single method.

In this study, a new rapid and flexible method for the simultaneous determination of 18 key representatives of polychlorinated biphenyls (PCBs), 7 polybrominated diphenyl ethers (PBDEs), and 32 polycyclic aromatic hydrocarbons (PAHs) in fish and shrimps by gas chromatography coupled to mass spectrometry (GC-MS) was developed and validated. A substantial simplification of sample processing prior to quantification step was achieved: after addition of water to homogenized sample, transfer of hydrophobic analytes into ethyl acetate was supported by added inorganic salts. Bulk fat, contained in crude organic extract obtained by partition, was subsequently removed on a silica minicolumn. This approach enabled to process six samples in less than 1h; moreover, the volume of an extraction solvent and consumption of other chemicals can be significantly reduced compared to, e.g., traditional Soxhlet extraction followed by gel permeation chromatography. The recoveries of target analytes were in the range of 73-120% even at the lowest spiking level (1 μg kg(-1)), repeatabilities (relative standard deviations, RSDs) ranged from 1 to 20%. Under optimized GC-MS conditions (time-of-flight mass analyzer, TOF), the limits of quantification (LOQs) were as follows: PCBs 0.1-0.5 μg kg(-1), PBDEs 0.5 μg kg(-1), and PAHs 0.05-0.25 μg kg(-1). Ambient mass spectrometry employing a direct analysis in real time (DART) ion source was shown as an effective tool for fat control in extract, which is needed during the method development and examination of unknown samples prior to the analysis. Further extension of a method scope by other similar analytes is easily possible.

Direct analysis of dithiocarbamate fungicides in fruit by ambient mass spectrometry

Dithiocarbamates (DTCs) are fungicides that require a specific single-residue method for detection and verification of compliance with maximum residue limits (MRLs) as established for fruit and vegetables in the EU. In this study, the use of ambient mass spectrometry was investigated for specific determination of individual DTCs (thiram, ziram) in fruit. Two complementary approaches have been investigated for their rapid analysis: (i) direct analysis in real time (DART) combined with medium-high resolution/accurate mass time-of-flight mass spectrometry (TOFMS) and high-resolution/accurate mass Orbitrap MS, and (ii) desorption electrospray ionization (DESI) combined with tandem-in-time mass spectrometry (MS2). With both techniques, thiram deposited on a glass surface (DART) or Teflon (DESI) could be directly detected. With DART, this was also possible for ziram. Before the instrumental analysis of fruit matrix, an extract had to be prepared following a straightforward procedure. The raw extracts were deposited on a slide (DESI), or rods were dipped into the extracts (DART), after which thiram and ziram could be rapidly detected (typically 10 samples in a few minutes). In the case of thiram, the lowest calibration levels were 1 mg kg−1 (DART–TOFMS, DESI–MS2) and 0.1 mg kg−1 (DART–Orbitrap MS). For ziram, the achieved lowest calibration levels were 0.5 mg kg−1 (DART–TOFMS) and 1 mg kg−1 (DART–Orbitrap MS). In all cases, this was sufficiently low to test samples against EU-MRLs for a number of fruit crops. Using an internal standard, (semi)quantitative results could be obtained.

Liquid chromatography–mass spectrometry-based metabolomics for authenticity assessment of fruit juices

High performance liquid chromatography–mass spectrometry (HPLC–MS) technique, employing a hybrid triple quadrupole/linear ion trap (QqQ/LIT) mass analyzer, was used for comprehensive metabolomic fingerprinting of several fruit juices types, prepared from expensive (orange) or relatively low-priced (apple, grapefruit) fruits. Following the automated data mining and pre-treatment step, the suitability of the multivariate HPLC–MS metabolomic data for authentication, i.e., classification of fruit juice and adulteration detection, was assessed with the use of advanced chemometric tools (principal component analysis, PCA, and linear discrimination analysis, LDA). The LDA classification model, constructed and validated employing a highly variable samples set, was able to reliably detect 15% addition of apple or grapefruit juice to orange juice. In the final stage of this study, high performance liquid chromatography–quadrupole–quadrupole-time-of-flight mass spectrometry (HPLC–QqTOFMS) measurements were performed in order to obtain data for identification of pre-selected marker compounds using elemental formula calculation and online databases search.