Liquid chromatographic/electrospray ionization tandem mass spectrometric study of the phenolic composition of cocoa (Theobroma cacao)

Liquid chromatography coupled with ionspray mass spectrometry in the tandem mode (LC/MS/MS) with negative ion detection was used for the identification of a variety of phenolic compounds in a cocoa sample. Gradient elution with water and acetonitrile, both containing 0.1% HCOOH, was used. Standard solutions of 31 phenolic compounds, including benzoic and cinnamic acids and flavonoid compounds, were studied in the negative ion mode using MS/MS product ion scans. At low collisional activation, the deprotonated molecule [M - H](-) was observed for all the compounds studied. For cinnamic and benzoic acids, losses of CO(2) or formation of [M - CH(3)](-*) in the case of methoxylated compounds were observed. However, for flavonol and flavone glycosides, the spectra present both the deprotonated molecule [M - H](-) of the glycoside and the ion corresponding to the deprotonated aglycone [A - H](-). The latter ion is formed by loss of the rhamnose, glucose, galactose or arabinose residue from the glycosides. Different fragmentation patterns were observed in MS/MS experiments for flavone-C-glycosides which showed fragmentation in the sugar part. Fragmentation of aglycones provided characteristic ions for each family of flavonoids. The optimum LC/MS/MS conditions were applied to the characterization of a cocoa sample that had been subjected to an extraction/clean-up procedure which involved chromatography on Sephadex LH20 and thin-layer chromatographic monitoring. In addition to compounds described in the literature, such as epicatechin and catechin, quercetin, isoquercitrin (quercetin-3-O-glucoside) and quercetin-3-O-arabinose, other compounds were identified for the first time in cocoa samples, such as hyperoside (quercetin-3-O-galactoside), naringenin, luteolin, apigenin and some O-glucosides and C-glucosides of these compounds.     

Comparison of gas chromatographic and gas chromatographic/mass spectrometric techniques for the analysis of TNT and related nitroaromatic compounds

The use of capillary column gas chromatography and gas chromatography/mass spectrometry for the analysis of a series of standard solutions (0.1 to 10 μg/ml) of 2,4,6-trinitrotoluene (TNT) and eight other nitroaromatic components was evaluated. The techniques included gas chromatography with electron capture detection (GC/ECD), full scan and selected ion monitoring gas chromatography/mass spectrometry with electron impact ionization (EI/FS and EI/SIM), full scan and selected ion monitoring gas chromatography/mass spectrometry with positive ion chemical ionization using methane reagent gas (PICI/FS and PICI/SIM), and full scan and selected ion monitoring gas chromatography/mass spectrometry with negative ion chemical ionization using methane reagent gas (NICI/FS and NICI/SIM). The performance of the techniques was comapared by determining the linear response range, precision, and detection limits of the analyses.     

MALDI‐TOF mass spectrometry of hordeins: rapid approach for identification of malting barley varieties

A procedure for identification of malting barley varieties using matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) of ethanol‐soluble barley proteins (hordeins) is described. The hordeins were first extracted from milled barley grains by several extraction protocols (using different extraction agents and conditions). Hordein extracts were then analyzed directly via MALDI‐TOF MS without any preliminary purification or separation step, and the protein profiles of analyzed hordein extracts were compared in order to find out the most suitable extraction procedure for mass spectrometric analysis. The optimized procedure was successfully applied to identification of 13 malting barley varieties. Our results revealed that the proposed mass spectrometry‐based approach provides characteristic mass patterns of extracted hordeins, which can be advantageously used for barley variety identification. Copyright © 2009 John Wiley & Sons, Ltd.     

LC‐PDA‐ESI‐MSn analysis of phenolic and iridoid compounds from Globularia spp.

Plants produce a great number of metabolites with potentially useful biological activities. Species from the genus Globularia (Plantaginaceae) are known as sources of different phenolic and iridoid compounds. Globularia alypum L. is a medicinal plant used as a healing agent in many Mediterranean countries. Similarities in phytochemical composition are often observed for related species. For Globularia spp., such findings were mostly based on identification of several isolated compounds from distinct species. To our knowledge, this is the first study that enables simultaneous comparison of phytochemical profiles from several members of the genus Globularia. Liquid chromatography‐photodiode array detection‐electrospray ionization‐tandem mass spectrometry was used for the analysis of methanolic extracts of aerial parts obtained from four Globularia species (G. alypum, G. punctata, G. cordifolia and G. meridionalis). In total, 85 compounds were identified or tentatively identified based on comparison of their retention time, UV and MSⁿ (up to MS⁴) spectra to those of standard compounds and/or to literature data. Among these, high relative amounts of bioactive molecules such as globularin, globularifolin, asperuloside and verbascoside (acteoside) were found. Apart from providing new insights into the phytochemistry and chemotaxonomy of selected Globularia species, results of this study complement existing MS/MS spectral data and could enable easier mass spectrometric profiling of certain bioactive compounds such as iridoids and phenylethanoids in related plant species, genera and families. Copyright © 2016 John Wiley & Sons, Ltd.     

Recent developments of nanoparticle-based enrichment methods for mass spectrometric analysis in proteomics

In proteome research, rapid and effective separation strategies are essential for successful protein identification due to the broad dynamic range of proteins in biological samples. Some important proteins are often expressed in ultra low abundance, thus making the pre-concentration procedure before mass spectrometric analysis prerequisite. The main purpose of enrichment is to isolate target molecules from complex mixtures to reduce sample complexity and facilitate the subsequent analyzing steps. The introd...     

Sensitive liquid chromatographic/tandem mass spectrometric method for the determination of beclomethasone dipropionate and its metabolites in equine plasma and urine

Beclomethasone dipropionate (BDP) is a potent pro-drug to beclomethasone (BOH) and is used in the treatment of chronic and acute respiratory disorders in the horse. The therapeutic dose of BDP (325 microg per horse) by inhalation results in very low plasma and urinary concentrations of BDP and its metabolites that pose a challenge to detection and confirmation by equine forensic laboratories. To solve this problem, a method involving the use of a liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) was developed for the detection, confirmation and quantification of the analytes in equine samples. Ammonium formate or acetate buffer added to LC mobile phase favored the formation of [M + H](+) ions from BDP and its metabolites, whereas formic acid led to the formation of sodium and potassium adduct ions ([M + Na](+), [M + K](+)) together with [M + H](+) ions. Acetonitrile, on the other hand, favored the formation of abundant solvent adduct ions [M + H + CH(3)CN](+) with the analytes under electrospray ionization (ESI) and atmospheric pressure chemical ionization conditions. In contrast, methanol formed much less solvent adduct ions than acetonitrile. The solvent adduct ions were thermally stable and could not be completely desolvated under the experimental conditions, but they were very fragile to collision-induced dissociation (CID). Interestingly, these solvent adduct ions were observed on a triple-quadrupole mass spectrometry but not on an ion trap instrument where helium used as a damping gas in the ion trap might cause the solvent adduct ions desolvated by collision. By CID studies on the [M + H](+) ions of BDP and its metabolites, their fragmentation paths were proposed. In equine plasma at ambient temperature over 2 h, BDP and B21P were hydrolyzed in part to B17P and BOH, respectively, but B17P was not hydrolyzed. Sodium fluoride added to equine plasma inhibited the hydrolysis of BDP and B21P. The matrix effect in ESI was evaluated in equine plasma and urine samples. The method involved the extraction of BDP and its metabolites from equine plasma and urine samples by methyl tert-butyl ether, resolution on a C(8) column with a mobile phase gradient consisting of methanol and ammonium formate (2 mmol l(-1), pH 3.4) and multiple reaction monitoring for the analytes on a triple-quadrupole mass spectrometer. The detection limit was 13 pg ml(-1) for BDP and B17P, 25 pg ml(-1) for BOH and 50 pg ml(-1) for B21P in plasma and 25 pg ml(-1) for BOH in urine. The method was successfully applied to the analysis of equine plasma and urine samples for the analytes following administration of BDP to horses by inhalation. B17P, the major and active metabolite of BDP, was detected and quantified in equine plasma up to 4 h post-administration by inhalation of a very low therapeutic dose (325 microg per horse) of BDP.     

Liquid chromatographic tandem mass spectrometric determination of trandolapril in human plasma

A liquid chromatographic tandem mass spectrometric method for the determination of trandolapril in human plasma has been developed and fully validated. The article describes, in detail, the bioanalytical procedure and summarizes the validation results obtained. The samples were extracted using HLB Oasis solid-phase extraction cartridges. The chromatographic separation was performed on X-Terra C8 MS column (150 mm × 4.6 mm i.d., 5 μm particle size) using a mobile phase consisting of acetic acid 20 mM and triethylamine 4.3 mM/acetonitrile (40:60 (v/v)), pumped isocratically at 0.35 ml/min.

The analytes were detected using a micromass quattro micro triple quadrupole mass spectrometer with positive electrospray ionization in multiple reaction-monitoring (MRM) mode. Tandem mass spectrometric detection enabled the quantitation of trandolapril down to 2.0 ng/ml. Calibration graphs were linear (r better than 0.996, n = 9) in the concentration ranges 2.0–750 ng/ml and the intra- and inter-day R.S.D. values were less than 3.83 and 3.86% for trandolapril.     

Simultaneous quantitative analysis of polyphenolic compounds in human plasma by liquid chromatography tandem mass spectrometry

A diet rich in polyphenolic compounds has recognized health benefits, and as such is routinely monitored as part of dietary intervention studies. A method for the simultaneous determination of 36 phenolic compounds, including phenolic acids and flavonoids, using liquid chromatography and tandem mass spectrometry is described here. The target analytes were quantified based on their specific mass spectral fragments using a selected reaction monitoring approach. A C18 column with embedded aromatic functionality ensured separation of all phenolic compounds studied which included several pairs of isomers. Sample preparation involved the use of β‐glucuronidase to release the phenolic compounds from their conjugated forms. The intra‐day and inter‐day precision and accuracy was less than 7% for all phenolic compounds studied. Recoveries, where plasma was spiked with three different concentrations of the analytes, ranged from 95–115%. The limits of detection and quantification were 0.23–3.89 and 1.15–7.79 nM, respectively. The method was successfully applied to real samples and the range reported for each phenolic compound, with the exception of hydroferulic acid, nordihydroguaiaretic acid, methylgallate, and m‐coumaric acid, was at least an order of magnitude higher than the limit of quantification for the method.     

Rapid determination of ginkgolic acids in Ginkgo biloba kernels and leaves by direct analysis in real time‐mass spectrometry

A novel method based on direct analysis in real time integrated with mass spectrometry was established and applied into rapid determination of ginkgolic acids in Ginkgo biloba kernels and leaves. Instrument parameter settings were optimized to obtain the sensitive and accurate determination of ginkgolic acids. At the sample introduction speed of 0.2 mm/s, high intensity of [M–H]^– ions for ginkgolic acids were observed in the negative ion mode by utilization of high‐purity helium gas at 450°C. Two microliters of methanol extract of G. biloba kernels or leaves dropped on the surface of Quick‐Strip module was analyzed after solvent evaporated to dryness. A series of standard solutions of ginkgolic acid 13:0 in the range of 2–50 mg/L were analyzed with a correlation coefficient r  = 0.9981 and relative standard deviation (n  = 5) from 12.5 to 13.7%. The limit of detection was 0.5 mg/L. The results of direct analysis in real time‐mass spectrometry were in agreement with those observed by thermochemolysis gas chromatography. The proposed method demonstrated significant potential in the application of the high‐throughput screening and rapid analysis for ginkgolic acids in dietary supplements.     

MASSyPup — an ‘Out of the Box’ solution for the analysis of mass spectrometry data

Mass spectrometry has evolved to a key technology in the areas of metabolomics and proteomics. Centralized facilities generate vast amount of data, which frequently need to be processed off‐site. Therefore, the distribution of data and software, as well as the training of personnel in the analysis of mass spectrometry data, becomes increasingly important. Thus, we created a comprehensive collection of mass spectrometry software which can be run directly from different media such as DVD or USB without local installation. MASSyPup is based on a Linux Live distribution and was complemented with programs for conversion, visualization and analysis of mass spectrometry (MS) data. A special emphasis was put on protein analysis and proteomics, encompassing the measurement of complete proteins, the identification of proteins based on Peptide Mass Fingerprints (PMF) or LC‐MS/MS data, and de novo sequencing. Another focus was directed to the study of metabolites and metabolomics, covering the detection, identification and quantification of compounds, as well as subsequent statistical analyses. Additionally, we added software for Mass Spectrometry Imaging (MSI), including hardware support for self‐made MSI devices. MASSyPup represents a ‘ready to work’ system for teaching or MS data analysis, but also represents an ideal platform for the distribution of MS data and the development of related software. The current Live DVD version can be downloaded free of charge from http://www.bioprocess.org/massypup . Copyright © 2014 John Wiley & Sons, Ltd.     

Induction based fluidics (IBF) for droplet‐based mass spectrometric analysis of oligonucleotides

Here, we report the utility of induction‐based fluidics (IBF) for the introduction of oligonucleotides to a mass spectrometer via charged droplets. The device produces nanoliter‐sized droplets, which are field transported with minimal concerns related to source plugging or sampling loss. The IBF source enabled detection of oligonucleotides at the nanomolar concentration level. Importantly, analysis of individual droplets revealed that oligonucleotide mixtures could be detected with ion abundance ratios that closely match the initial concentration ratios within the sample. Copyright © 2015 John Wiley & Sons, Ltd.     

In‐time and in‐space tandem mass spectrometry to determine the metabolic profiling of flavonoids in a typical sweet cherry (Prunus avium L.) cultivar from Southern Italy

This paper presents a comprehensive analytical methodology, based on ‘in‐time’ and ‘in‐space’ tandem mass spectrometry (MS) techniques, to identify and quantify flavonoid compounds in a typical Italian sweet cherry cultivar (cv. Ferrovia). Five anthocyanins, four flavan‐3‐ols and nine flavonols were determined by means of hyphenated high‐performance liquid chromatography – multi‐stage MS (HPLC‐MSⁿ) analyses (MSⁿ up to MS⁴), among which quercetin‐3‐O‐rutinoside‐7‐O‐glucoside, kaempferol‐3‐O‐rutinoside‐7‐O‐glucoside, quercetin‐3‐O‐galactosyl‐rhamnoside and quercetin‐3‐O‐coumaroylglucoside were tentatively identified in sweet cherries for the first time. Ultrafast HPLC and tandem MS (UHPLC‐MS/MS) analyses through multiple reaction monitoring experiments showed that cyanidin‐3‐O‐rutinoside and cyanidin‐3‐O‐glucoside were the main anthocyanins of cv. Ferrovia at maturity. Moreover, consistent levels of catechin and epicatechin as well as quercetin‐3‐O‐rutinoside and kaempferol‐3‐O‐rutinoside were also found. Because flavonoids have been ascribed as potential health‐promoting compounds, gathered findings provide new insight into the knowledge of the quali‐quantitative profile of these phytochemicals into a widespread fruit such as sweet cherry. Copyright © 2014 John Wiley & Sons, Ltd.     

Combining targeted and nontargeted data analysis for liquid chromatography/high‐resolution mass spectrometric analyses

Increasing importation of food and the diversity of potential contaminants have necessitated more analytical testing of these foods. Historically, mass spectrometric methods for testing foods were confined to monitoring selected ions (SIM or MRM), achieving sensitivity by focusing on targeted ion signals. A limiting factor in this approach is that any contaminants not included on the target list are not typically identified and retrospective data mining is limited. A potential solution is to utilize high‐resolution MS to acquire accurate mass full‐scan data. Based on the instrumental resolution, these data can be correlated to the actual mass of a contaminant, which would allow for identification of both target compounds and compounds that are not on a target list (nontargets). The focus of this research was to develop software algorithms to provide rapid and accurate data processing of LC/MS data to identify both targeted and nontargeted analytes. Software from a commercial vendor was developed to process LC/MS data and the results were compared to an alternate, vendor‐supplied solution. The commercial software performed well and demonstrated the potential for a fully automated processing solution.     

Mass spectrometric evidence for the modification of small molecules in a cobalt‐60‐irradiated rodent diet

The purpose of this study was to investigate the effect of radiation on the content of animal diet constituents using global metabolomics. Aqueous methanolic extracts of control and cobalt‐60‐irradiated Teklad 7001 diets were comprehensively analyzed using nano‐liquid chromatography–MS/MS. Among the over 2000 ions revealed by XCMS followed by data preprocessing, 94 positive and 143 negative metabolite ions had greater than 1.5‐fold changes and p‐values <0.01. Use of MetaboAnalyst statistical software demonstrated complete separation of the irradiated and non‐radiated diets in unsupervised principal components analysis and supervised partial least squares discriminant analysis. Irradiation led to an increase in the content of phytochemicals such as glucosinolates and oxidized lipids in the diet. Twenty‐eight metabolites that were significantly changed in the irradiated samples were putatively identified at the level of molecular formulae by MS/MS. MS/MS^ALL analysis of chloroform–methanol extracts of the irradiated diet showed increased levels of a number of unique linoleic acid‐derived branched fatty acid esters of hydroxy fatty acids. These data imply that gamma irradiation of animal diets causes chemical changes to dietary components, which in turn may influence the risk of mammary cancer. Copyright © 2017 John Wiley & Sons, Ltd.     

An approach to analysis of primary amines by a combination of thin‐layer chromatography and matrix‐assisted laser desorption ionization mass spectrometry in conjunction with post‐chromatographic derivatization

On‐spot derivatization has been suggested for the modification of primary amine containing compounds for their analysis by thin‐layer chromatography hyphenated with matrix‐assisted laser desorption ionization mass spectrometry. The proposed approach was based on post‐chromatographic treatment of separated analytes inside the chromatographic zones on the thin‐layer chromatography plates by tris(2,6‐dimethoxyphenyl)methilium reagent. The derivatives, containing permanent positive charge, reveal exceptionally intense peaks of their cationic moieties and high signal/noise ratio in mass spectra recorded directly from the plates. The method was tested on a series of aliphatic, aromatic, and amine‐containing pharmaceuticals.     

Evaluation of the chemical consistency of Yin‐Chen‐Hao‐Tang prepared by combined and separated decoction methods using high‐performance liquid chromatography and quadrupole time‐of‐flight mass spectrometry coupled with multivariate statistical analysis

In this study, Yin‐Chen‐Hao‐Tang prepared by two decoction methods, namely, combined decoction (modern decoction method) and separated decoction (traditional decoction method), was analyzed by high‐performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry. The acquired datasets containing sample codes, t_R‐m/z pairs and ion intensities were processed with multivariate statistical analyses, such as principal component analysis and an orthogonal partial least squared discriminant analysis model, to globally compare the chemical differences between the different decoction samples. Then, the chemical differences between the combined and separated decoctions were screened out by S‐plots generated from the orthogonal partial least squared discriminant analysis model and compared with chemical information from an established in‐house library. The six components that contributed the most to the chemical differences were identified as chlorogenic acid, caffeic acid, geniposide, genipin, scopoletin, and 3,5‐dicaffeoylquinic acid. The concentrations of genipin and caffeic acid from the separated decoction were higher than those from the combined decoction, indicating that the separated decoction may present a stronger hepatoprotective effect. However, the results still require further investigation through pharmacological and clinical studies. Our findings not only establish a strategy to evaluate chemical consistency of Yin‐Chen‐Hao‐Tang but also provide the scientific basis for using traditional separated decoction method.     

Improved liquid chromatographic tandem mass spectrometric determination and pharmacokinetic study of glimepiride in human plasma

An improved liquid chromatographic tandem mass spectrometric method for the determination of glimepiride in human plasma has been developed and fully validated. The article describes in detail the bioanalytical procedure and summarizes the validation results obtained. The samples were extracted using liquid--liquid extraction with a mixture of 1-chlorobutane-isopropanol-ethyl acetate (88:2:10, v/v/v). The chromatographic separation was performed on a reversed-phase Hypersil ODScolumn (250 x 4.6 mm i.d.; 5 microm particle size) using a mobile phase consisting of formic acid 0.05 M-acetonitrile (28:72, v/v), pumped at a flow rate of 0.3 ml min(-1) heated to 25 degrees C. The analytes were detected using an API 3000 triple quadrupole mass spectrometer with positive electrospray ionization in multiple reaction monitoring mode. Tandem mass spectrometric detection enabled the quantitation of glimepiride down to 0.50 ng mL(-1). Calibration graphs were linear (r better than 0.998, n=1), in concentration range 0.50--1000 ng mL(-1), and the intra- and inter- day RSD values were less than 10.37 and 11.55% for glimepiride. The method was successfully applied to a kinetic study in order to assess the main pharmacokinetic parameters of glimepiride.     

Comparison of the origin and phenolic contents of Lycium ruthenicum Murr. by high‐performance liquid chromatography fingerprinting combined with quadrupole time‐of‐flight mass spectrometry and chemometrics

The fruits of Lycium ruthenicum Murr. have long been used in folk medicine. Nevertheless, detailed information related to its phenolic composition and its quality control remains scarce. In this study, a simple and reproducible method, based on high‐performance liquid chromatography combined with chemometrics, was developed to authenticate 18 samples of L. ruthenicum Murr. collected from different parts of China through fingerprint analysis. The main peaks were identified by quadrupole time‐of‐flight electrospray ionization mass spectrometry. Four phenolics were quantified, and the most abundant phenolic compound in almost all samples was kukoamine A. Hierarchical cluster analysis and principal component analysis were applied to classify these samples. Also, a total of 26 compounds, which were mainly phenolic compounds and anthocyanins, were identified or tentatively identified based on the available literature and standard references. Among these, 16 were reported for the first time in the extract. The results showed that there was no significant difference between L. ruthenicum fruits from different provinces in terms of chemical composition. Also, the fingerprint together with chemometric analyses and quadrupole time‐of‐flight electrospray ionization mass spectrometry are promising methods for evaluating the quality consistency, identification, and comprehensive evaluation of L. ruthenicum .     

Mass spectrometric methods for the analysis of chlorinated and nitrated isoflavonoids: a novel class of biological metabolites

Electrospray ionization combined with tandem mass spectrometry was applied to a study of some representative chlorinated and nitrated isoflavones-potential metabolites of isoflavones in inflammatory cells. Upon collision-induced dissociation of deprotonated [M - H](-) ions of these compounds, a number of structurally characteristic product ions were produced. The product ion analysis of 3'- and 8-chlorodaidzein in the tandom mass spectra led to ready differentiation of these isomers. 3-Nitro derivatives of both genistein and daidzein have product ions due to the losses of HNO(2) and two OH groups. Chlorinated derivatives of isoflavones were detected in cell-based experiments and their structures were proposed by comparing the tandem mass spectra of their product ions with those of standards. This work provides a suitable analytical basis to aid the characterization of chlorinated and nitrated metabolites in studies in vivo and in vitro.     

Screening for pharmaco‐toxicologically relevant compounds in biosamples using high‐resolution mass spectrometry: a ‘metabolomic’ approach to the discrimination between isomers

High‐resolution mass spectrometry (HRMS) enables the identification of a chemical formula of small molecules through the accurate measurement of mass and isotopic pattern. However, the identification of an unknown compound starting from the chemical formula requires additional tools: (1) a database associating chemical formulas to compound names and (2) a way to discriminate between isomers. The aim of this present study is to evaluate the ability of a novel ‘metabolomic’ approach to reduce the list of candidates with identical chemical formula. Urine/blood/hair samples collected from real positive cases were submitted to a screening procedure using ESI‐MS‐TOF (positive‐ion mode) combined with either capillary electrophoresis or reversed phase liquid chromatography (LC). Detected peaks were searched against a Pharmaco/Toxicologically Relevant Compounds database (ca 50 500 compounds and phase I and phase II metabolites) consisting of a subset of PubChem compounds and a list of candidates was retrieved. Then, starting from the mass of unknown, mass shifts corresponding to pre‐defined biotransformations (e.g. demethylation, glucuronidation, etc.) were calculated and corresponding mass chromatograms were extracted from the total ion current (TIC) in order to search for metabolite peaks. For each candidate, the number of different functional groups in the molecule was automatically calculated using E‐Dragon software (Talete srl, Milan, Italy). Then, the presence of metabolites in the TIC was matched with functional groups data in order to exclude candidates with structures not compatible with observed biotransformations (e.g. loss of methyl from a structure not bearing methyls). The procedure was tested on 108 pharmaco‐toxicologically relevant compounds (PTRC) and their phase I metabolites were detected in real positive samples. The mean list length (MLL) of candidates retrieved from the database was 7.01 ± 4.77 (median, 7; range, 1–28) before the application of the ‘metabolomic’ approach, and after the application it was reduced to 4.08 ± 3.11 (median 3, range 1–17). HRMS allows a much broader screening for PTRC than other screening approaches (e.g. library search on mass spectra databases). The ‘metabolomic’ approach enables the reduction of the list of candidate isomers. Copyright © 2009 John Wiley & Sons, Ltd.