Evaluation of the sensitivity of the ‘Wiley registry of tandem mass spectral data,MSforID’ with MS/MS data of the ‘NIST/NIH/EPA mass spectral library’

Tandem mass spectral libraries are versatile tools for small molecular identification finding application in forensic science, doping control, drug monitoring, food and environmental analysis, as well as metabolomics. Two important libraries are the ‘Wiley Registry of Tandem Mass Spectral Data, MSforID’ (Wiley Registry MSMS) and the collection of MS/MS spectra part of the 2011 edition of the ‘NIST/NIH/EPA Mass Spectral Library’ (NIST 11 MSMS). Herein, the sensitivity and robustness of the Wiley Registry MSMS were evaluated using spectra extracted from the NIST 11 MSMS library. The sample set was found to be heterogeneous in terms of mass spectral resolution, type of CID, as well as applied collision energies. Nevertheless, sensitive compound identification with a true positive identification rate ≥95% was possible using either the MSforID Search program or the NIST MS Search program 2.0g for matching. To rate the performance of the Wiley Registry MSMS, cross‐validation experiments were repeated using subcollections of NIST 11 MSMS as reference library and spectra extracted from the Wiley Registry MSMS as positive controls. Unexpectedly, with both search algorithms tested, correct results were obtained in less than 88% of cases. We examined possible causes for the results of the cross validation study. The large number of precursor ions represented by a single tandem mass spectrum only was identified as the basic cause for the comparably lower sensitivity of the NIST library. Copyright © 2013 John Wiley & Sons, Ltd.     

On the inter‐instrument and the inter‐laboratory transferability of a tandem mass spectral reference library. 3. Focus on ion trap and upfront CID

Mass spectral libraries represent versatile tools for the identification of small bioorganic molecules. Libraries based on electron impact spectra are rated robust and transferable. Tandem mass spectral libraries are often considered to work properly only on the instrument that has been used to build the library. An exception from that rule is the ‘Wiley Registry of Tandem Mass Spectral Data, MSforID’. In various studies with data sets from different kinds of tandem mass spectrometric instruments, the outstanding sensitivity and robustness of this tandem mass spectral library search approach was demonstrated. The instrumental platforms tested, however, mainly included various tandem‐in‐space instruments. Herein, the results of a multicenter study with a focus on upfront and tandem‐in‐time fragmentation are presented. Five laboratories participated and provided fragment ion mass spectra from the following types of mass spectrometers: time‐of‐flight (TOF), quadrupole–hexapole–TOF, linear ion trap (LIT), 3‐D ion trap and LIT–Orbitrap. A total number of 1231 fragment ion mass spectra were collected from 20 test compounds (amiloride, buphenin, cinchocaine, cyclizine, desipramine, dihydroergotamine, dyxirazine, dosulepin, ergotamine, ethambutol, etofylline, mefruside, metoclopramide, phenazone, phentermine, phenytoin, sulfamethoxazole, sulfamoxole, sulthiame and tetracycline) on seven electrospray ionization instruments using 18 different instrumental configurations for fragmentation. For 1222 spectra (99.3%), the correct compound was retrieved as the best matching compound. Classified matches (matches with ‘relative average match probability’ >40.0) were obtained for 1207 spectra (98.1%). This high percentage of correct identifications clearly supports the hypothesis that the tandem mass spectral library approach tested is a robust and universal identification tool. Copyright © 2012 John Wiley & Sons, Ltd.     

On the inter‐instrument and inter‐laboratory transferability of a tandem mass spectral reference library: 1. Results of an Austrian multicenter study

The inter‐instrument and inter‐laboratory transferability of a tandem mass spectral reference library originally built on a quadrupole‐quadrupole‐time‐of‐flight instrument was examined. The library consisted of 3759 MS/MS spectra collected from 402 reference compounds applying several different collision‐energy values for fragmentation. In the course of the multicenter study, 22 test compounds were sent to three different laboratories, where 418 tandem mass spectra were acquired using four different instruments from two manufacturers. The study covered the following types of tandem mass spectrometers: quadrupole‐quadrupole‐time‐of‐flight, quadrupole‐quadrupole‐linear ion trap, quadrupole‐quadrupole‐quadrupole, and linear ion trap‐Fourier transform ion cyclotron resonance mass spectrometer. In each participating laboratory, optimized instrumental parameters were gathered solely from routinely applied workflows. No standardization procedure was applied to increase the inter‐instrument comparability of MS/MS spectra. The acquired tandem mass spectra were matched against the established reference library using a sophisticated matching algorithm, which is presented in detail in a companion paper. Correct answers, meaning that the correct compound was retrieved as top hit, were obtained in 98.1% of cases. For the remaining 1.9% of spectra, the correct compound was matched at second rank. The observed high percentage of correct assignments clearly suggests that the developed mass spectral library search approach is to a large extent platform independent. Copyright © 2009 John Wiley & Sons, Ltd.     

Tandem mass spectral studies of the fragmentation pathways of organotin compounds of general formula R3SnR′

Positive ion‐electron impact (EI) mass spectra of organotin compounds of the type R₃SnR′, where R = Ph, n−Bu, n−Pe and R′ = allyl, vinyl, H and Ph, have been recorded. The spectra were also examined by tandem mass spectrometry (MS‐MS) in order to establish fragmentation reaction mechanisms for compounds bearing mixed substituents. Fragmentation patterns of six organotin compounds, based on precursor‐product ion relationships are proposed. Significant differences were found in the reaction pathways of organotin compounds with different substituents. This technique has potential to predict the effect of substitution on the mass spectra of organotin compounds. Copyright © 2000 John Wiley & Sons, Ltd.     

Evaluation of mass spectral library search algorithms implemented in commercial software

Performance of several library search algorithms (against EI mass spectral databases) implemented in commercial software products ( acd/specdb , chemstation , gc/ms solution and ms search ) was estimated. Test set contained 1000 mass spectra, which were randomly selected from NIST'08 (RepLib) mass spectral database. It was shown that composite (also known as identity) algorithm implemented in ms search (NIST) software gives statistically the best results: the correct compound occupied the first position in the list of possible candidates in 81% of cases; the correct compound was within the list of top ten candidates in 98% of cases. It was found that use of presearch option can lead to rejection of the correct answer from the list of possible candidates (therefore presearch option should not be used, if possible). Overall performance of library search algorithms was estimated using receiver operating characteristic curves. Copyright © 2015 John Wiley & Sons, Ltd.     

Automatic recalibration and processing of tandem mass spectra using formula annotation

High accuracy, high resolution tandem mass spectrometry (MS/MS) is becoming more common in analytical applications, yet databases of these spectra remain limited. Databases require good quality spectra with sufficient compound information, but processing, calibration, noise reduction and retrieval of compound information are time‐consuming tasks that prevent many contributions. We present a comprehensive workflow for the automatic processing of MS/MS using formula annotation for recalibration and cleanup to generate high quality spectra of standard compounds for upload to MassBank ( www.massbank.jp ). Compound information is retrieved via Internet services. Reference standards of 70 pesticides were measured at various collision energies on an LTQ‐Orbitrap XL to develop and evaluate the workflow. A total of 944 resulting spectra are now available on MassBank. Evidence of nitrogen adduct formation during MS/MS fragmentation processes was found, highlighting the benefits high accuracy MS/MS offers for spectral interpretation. A database of recalibrated, cleaned‐up spectra resulted in the most correct spectra ranked in first place, regardless of whether the search spectra were recalibrated or not, whereas the average rank of the correct molecular formula was improved from 2.55 (uncalibrated) to 1.53 when using recalibrated MS/MS data. The workflow is available as an R package RMassBank capable of generating MassBank records from raw MS and MS/MS data and can be adjusted to process data acquired with different settings and instruments. This workflow is a vital step towards addressing the need for more high quality, high accuracy MS/MS spectra in spectral databases and provides important information for spectral interpretation. Copyright © 2012 John Wiley & Sons, Ltd.     

Quality evaluation of tandem mass spectral libraries

Tandem mass spectral libraries are gaining more and more importance for the identification of unknowns in different fields of research, including metabolomics, forensics, toxicology, and environmental analysis. Particularly, the recent invention of reliable, robust, and transferable libraries has increased the general acceptance of these tools. Herein, we report on results obtained from thorough evaluation of the match reliabilities of two tandem mass spectral libraries: the MSforID library established by the Oberacher group in Innsbruck and the Weinmann library established by the Weinmann group in Freiburg. Three different experiments were performed: (1) Spectra of the libraries were searched against their corresponding library after excluding either this single compound-specific spectrum or all compound-specific spectra prior to searching; (2) the libraries were searched against each other using either library as reference set or sample set; (3) spectra acquired on different mass spectrometric instruments were matched to both libraries. Almost 13,000 tandem mass spectra were included in this study. The MSforID search algorithm was used for spectral matching. Statistical evaluation of the library search results revealed that principally both libraries enable the sensitive and specific identification of compounds. Due to higher mass accuracy of the QqTOF compared with the QTrap instrument, matches to the MSforID library were more reliable when comparing spectra with both libraries. Furthermore, only the MSforID library was shown to be efficiently transferable to different kinds of tandem mass spectrometers, including “tandem-in-time” instruments; this is due to the coverage of a large range of different collision energy settings—including the very low range—which is an outstanding characteristics of the MSforID library.     

Rapid method for the confirmatory analysis of chrysoidine in aquaculture products by ultra‐performance liquid chromatography–tandem mass spectrometry

A sensitive and fast method for the quantification of the illegal dye chrysoidine in aquaculture products with ultra‐performance liquid chromatography–tandem mass spectrometry (UPLC‐MS/MS) is presented. Muscle tissues were made alkaline with sodium hydroxide and extracted with ethyl acetate. After evaporation and subsequent defatting with n‐hexane, extracts were directly injected onto the UPLC‐column. Chromatography was performed on a C₁₈ column using 0.1% formic acid in water and an acetonitrile gradient within 6 min. Mass spectrometric analysis was performed in the positive electrospray MS/MS mode. The limit of quantification was 0.25 ng/g, which was 30 times lower than the only previously published method with gas chromatographic detection. A complete validation according to the scientific literature and as defined by the European Union was performed. The applicability of the method was shown in the analysis of more than 50 unknown samples in the framework of a monitoring program. Copyright © 2010 John Wiley & Sons, Ltd.     

L‐Valine assisted distinction between the stereo‐isomers of D‐hexoses by positive ion ESI tandem mass spectrometry

The binary mixtures of 7 hexoses and 20 amino acids were investigated by electrospray ionization ion trap mass spectrometry (ESI‐ITMS). The adduct ions of the amino acid and the hexose were detected for 12 amino acids but not for the other 8 amino acids which are basic acidic amino acids and amides. The ions of amino acid–hexose complexes were further investigated by tandem mass spectrometry (MS/MS), and some of them just split easily into two parts whereas the others gave rich fragmentation, such as the complex ions of isoleucine, phenylalanie, tyrosine, and valine. We found that hexoses could be complexed by two molecules of valine but only by one molecule of the other amino acids. Among seven kinds of valine–hexose complexes coordinated by potassium ion, the MS² spectra of the ion at m/z 453 yielded unambiguous differentiation. And the fragmentation ions are sensitive to the stereochemical differences at the carbon‐4 of hexoses in the complexes, as proved by the MS². Copyright © 2010 John Wiley & Sons, Ltd.     

SANIST: optimization of a technology for compound identification based on the European Union directive with applications in forensic,pharmaceutical and food analyses

Electrospray Ionization and collision induced dissociation tandem mass spectrometry are usually employed to obtain compound identification through a mass spectra match. Different algorithms have been developed for this purpose (for example the nist match algorithm). These approaches compare the tandem mass spectra of the unknown analyte with the tandem mass spectra spectra of known compounds inserted in a database. The compounds are usually identified on the basis of spectral match value associated with a probability of recognition. However, this approach is not usually applied to multiple reaction monitoring transition spectra achieved by means of triple quadrupole apparatus, mainly due to the lack of a transition spectra database. The Surface Activated Chemical Ionization‐Electrospray‐NIST Bayesian model database search (SANIST) platform has been recently developed for new potential metabolite biomarker discovery, to confirm their identity and to use them for clinical and diagnostic applications. Here, we present an improved version of the SANIST platform that extends its application to forensic, pharmaceutical, and food analysis studies, where the compound identification rules are strict. The European Union (EU) has set directives for compound identification (EU directive 2002/657/EC). We have applied the SANIST method to identification of 11‐nor‐9‐carboxytetrahydro‐cannabinol in urine samples (an example of a forensic application), circulating levels of the immunosuppressive drug tacrolimus in blood (an example of a pharmaceutical application) and glyphosate in fruit juice (an example of a food analysis application) that meet the EU directive requirements. Copyright © 2016 John Wiley & Sons, Ltd.     

Mass spectrometry studies of organolead compounds

The mass spectra of alkyl‐, aryl‐ and chlorinated ‐alkyl, aryl organolead compounds were investigated. Positive and negative ion mass spectra of these compounds were recorded using conventional electron impact conditions. In common with the analogous tetraalkyltin and tetraalkylgermanium compounds, tetrabutyllead produced no negative ion spectra under these conditions. The spectra were also examined by tandem mass spectrometry in order to establish reaction mechanisms for these compounds. Fragmentation patterns of seven organolead compounds, based on precursor–product ion relationships, are proposed. Copyright © 2001 John Wiley & Sons, Ltd.     

Confirmation of drug delivery after liver chemoembolization: direct tissue doxorubicin measurement by UHPLC‐MS‐MS

Because liver cancer is rarely suitable for surgery, transcatheter arterial chemoembolization (TACE) is used for palliative therapy. In this procedure, an emulsion of doxorubicin in iodized oil is injected directly into liver tumors through a catheter positioned within the artery supplying blood flow to the tumor. At present, there is limited understanding of factors affecting the delivery and dispersion of doxorubicin within treated tumors during TACE. This study addresses the development and application of an ultrahigh‐pressure liquid chromatography–tandem mass spectrometry (UHPLC‐MS‐MS) method for rapid confirmation of drug delivery after TACE in a rabbit VX2 liver cancer model. Doxorubicin levels in liver tumors were measured using UHPLC‐MS‐MS and compared with computed tomography measured levels of iodized oil, a metric used clinically to indicate drug delivery. We found that tissue drug levels determined using UHPLC‐MS‐MS did not correlate with the regional iodized oil concentration (vehicle) within tumors following TACE, suggesting that chemotherapeutic drugs like doxorubicin spread throughout tumors, and that lack of iodized oil staining in portions of a tumor does not necessarily indicate inadequate therapy during TACE. Copyright © 2012 John Wiley & Sons, Ltd.     

Application of LC-MS/MS method for the in vivo metabolite determination of oleuropein after intravenous administration to rat

A highly sensitive, specific and simple LC‐MS/MS method was developed to investigate in vivo bio‐transformation of oleuropein in rat. Rat urine samples collected after the intravenous administrations were determined using liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the negative‐ion mode. The assay procedure involves a simple liquid–liquid extraction of parent oleuropein and the metabolite from rat urine with ethyl acetate. Chromatographic separation was operated with 0.1% formic acid aqueous and methanol in gradient program at a flow rate of 0.80 mL/min on an RP‐C₁₈ column with a total run time of 30 min. This method has been successfully applied to simultaneous determination of oleuropein and its metabolite in rat urine. Oxygenation was found to be the major metabolic pathway of the oleuropein in rat after intravenous administration. Copyright © 2011 John Wiley & Sons, Ltd.     

Determination of the relative ligand‐binding strengths in heteroleptic IrIII complexes by ESI‐Q‐TOF tandem mass spectrometry

An electrospray ionization quadrupole time‐of‐flight mass spectrometer has been utilized to investigate the relative ligand‐binding strengths in a series of heteroleptic‐charged iridium(III) complexes of the general formula [(C^N)₂Ir^III(S‐tpy)](PF₆) by using variable collision energies. Collision‐induced dissociation experiments were performed in order to study the stability of the Ir^III complexes that are, for instance, suitable phosphors in light‐emitting electrochemical cells. The ratio of signal intensities belonging to the fragment and the undissociated complex depends on the collision energy applied for the tandem mass spectra (MS/MS) analysis. By defining the threshold collision energy and the point of complete complex dissociation, it is possible to estimate the relative complex stabilities depending on the nature of the coordinated ligands [i.e. type of cyclometalating ligand (C^N), substituents on the S‐shaped terpyridine (S‐tpy)]. The collision energy values differed as a function of the coordination sphere of the Ir^III centers. Copyright © 2011 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.     

Electrospray ionization tandem mass spectrometry of the star‐shaped propoxylated diethylenetriamine polyols

The collision‐induced dissociation of the protonated five‐arm star propoxylated diethylenetriamine polyols was studied under electrospray conditions. Two product ion series were detected because of the cleavage of the C? N bonds in the initiator moiety. No backbone fragmentation of the polyether chains was observed, which allowed to explore the initiation and side‐chain propagation process of the oligomers. On the basis of MS/MS spectra, it is probable that the rate of the initiation is larger than that of the chain propagation. The propylene oxide repeat units attach to the five arms with approximately the same probability. Furthermore, it was found that the collision energy necessary to obtain 50% fragmentation (CE₅₀) was linearly dependent on the molecular weight of the polyols. Copyright © 2015 John Wiley & Sons, Ltd.     

Characterization of N,N‐dimethyl amino acids by electrospray ionization–tandem mass spectrometry

Methylation is an essential metabolic process for a number of critical reactions in the body. Methyl groups are involved in the healthy function of the body life processes, by conducting methylation process involving specific enzymes. In these processes, various amino acids are methylated, and the occurrence of methylated amino acids in nature is diverse. Nowadays, mass‐spectrometric‐based identification of small molecules as biomarkers for diseases is a growing research. Although all dimethyl amino acids are metabolically important molecules, mass spectral data are available only for a few of them in the literature. In this study, we report synthesis and characterization of all dimethyl amino acids, by electrospray ionization–tandem mass spectrometry (MS/MS) experiments on protonated molecules. The MS/MS spectra of all the studied dimethyl amino acids showed preliminary loss of H₂O + CO to form corresponding immonium ions. The other product ions in the spectra are highly characteristic of the methyl groups on the nitrogen and side chain of the amino acids. The amino acids, which are isomeric and isobaric with the studied dimethyl amino acids, gave distinctive MS/MS spectra. The study also included MS/MS analysis of immonium ions of dimethyl amino acids that provide information on side chain structure, and it is further tested to determine the N‐terminal amino acid of the peptides. Copyright © 2015 John Wiley & Sons, Ltd.     

Comparison of CID versus ETD‐based MS/MS fragmentation for the analysis of doubly derivatized steroids

Electrospray ionization coupled with collision‐induced dissociation (CID) and tandem mass spectrometry (MS/MS) is a commonly used technique to analyze the chemical composition of steroids. However, steroids are structurally similar compounds, making it difficult to interpret their product‐ion spectra. Electron transfer dissociation (ETD), a relatively new technique for protein and peptide fragmentation, has been shown to provide more detailed structural information. In this study, we compared the ability of CID with that of ETD to differentiate between eight 3,20‐dioxosteroids that had been derivatizated with a quaternary ammonium salt, Girard reagent P (GirP), at room temperature or after exposure to microwave irradiation to generate doubly charged ions. We found that the derivatization of steroid with GirP hydrazine occurred in less than 10 min when the reaction was carried out in the presence of microwave irradiation compared to 30 min when the reaction was carried out at room temperature. According to the MS/MS spectra, CID provided rich, structurally informative ions; however, the spectra were complex, thereby complicating the peak assignment. In contrast, ETD generated simpler spectra, making it easier to recognize individual peaks. Remarkably, both CID and ETD were allowed to differentiate of steroid isomers, 17α‐hydroxyprogesterone (17OHP) and deoxycorticosterone (DOC), but the signature ions obtained from CID were less intense than those generated by ETD, which generated much clearer spectra. These results indicate that ETD in conjunction with CID can provide more structural information for precise characterization of steroids. Copyright © 2013 John Wiley & Sons, Ltd.     

Photooxidation of glycated and non‐glycated phosphatidylethanolamines monitored by mass spectrometry

Phosphatidylethanolamines (PE) are one of the major components of cells membranes, namely in skin and in retina, that are continuously exposed to solar UV radiation being major targets of photooxidation damage. In addition, due to the presence of the free amine group, PE can also undergo glycation, in hyperglycemic conditions which may increase the susceptibility to oxidation. The aim of this study is to develop a model, based on mass spectrometry (MS) analysis, to identify photooxidative degradation of selected PE (POPE: PE 16:0/18:1, PLPE: PE 16:0/18:2, PAPE: PE 16:0/20:4) and glycated PEs due to UV irradiation. Photooxidation products were analysed by electrospray ionization MS (ESI‐MS) and tandem MS (ESI‐MS/MS) in positive and negative mode. Emphasis is placed in the influence of glycation in the generation of distinct photooxidation products. ESI‐MS spectra of PE after UV photo‐irradiation showed mainly hydroperoxy derivatives, due to oxidation of unsaturated fatty acyl chains. Glycated PE gave rise to several new photooxidation products formed due to oxidative cleavages of the glucose moiety, namely between C1 and C2, C2 and C3, and C5 and C6 of this sugar unit. These new products were identified by ESI‐MS/MS in positive mode showing distinct neutral loss depending on the different structure of the polar head group. These new identified advanced glycated photooxidation products may have a deleterious role in the etiology of diabetic retinopathy and in diabetic retinal microvascular complications. Copyright © 2013 John Wiley & Sons, Ltd.