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The catalyst-free synthesis of urethanes from cyclic iminocarbonates, used as masked isocyanates, and soft nucleophiles (i.e. carboxylic acids and thiols) has been studied. Remarkably, the ring-opening reaction was fully site-selective (i.e. methylene). The disclosed method showed high functional group tolerance towards carboxylic acids bearing alkyl-, alkenyl-, ketone-, pyrone- and hydroxyl groups. This methodology was further applied for the construction of regioregular functional polyurethanes by step-growth copolymerization of cyclic bisiminocarbonates and dicarboxylic acids or dithiols. M-w up to 34 000 g mol(-1) was obtained in a fully atom-economical manner using an equimolar amount of both monomers.

Direct hydroxylation via the functionalization of tertiary benzylic C(sp(3))-H bonds is of great significance for obtaining tertiary alcohols, which find wide applications in pharmaceuticals as well as in fine chemical industries. However, current synthetic procedures use toxic reagents, and therefore, the development of a sustainable strategy for the synthesis of tertiary benzylic alcohols is highly desirable. To solve this problem, herein, we report a metal-free heterogeneous photocatalyst to synthesize the hydroxylated products using oxygen as the key reagent. Various benzylic substrates were employed into our mild reaction conditions to afford the desirable products in good to excellent yields. More importantly, the gram-scale reaction was achieved via harvesting direct solar energy and exhibited high quantity of the product. The high stability of the catalyst was proved via recycling the catalyst and spectroscopic analyses. Finally, a possible mechanism was proposed based on electron paramagnetic resonance and other experimental evidence.

Sensitivity to molecular ions remains a limiting factor for high resolution imaging mass spectrometry of organic and biological materials. Here, we investigate a variant of matrix-enhanced secondary ion mass spectrometry in which the transfer of matrix molecules to the analyte sample is carried out in situ (in situ ME-SIMS). This approach is therefore compatible with both 2D and 3D imaging by SIMS. In this exploratory study, nanoscale matrix layers were sputter-transferred inside our time-of-flight (ToF)-SIMS to a series of thin films of biomolecules (proteins, sugars, lipids) adsorbed on silicon, and the resulting layers were analyzed and depth-profiled. For this purpose, matrix molecules were desorbed from a coated target (obtained by drop-casting or sublimation) using 10 keV Ar-3000(+) ion beam sputtering, followed by redeposition on a collector carrying the sample to be analyzed. After evaluating the quality of the transfer of six different matrices on bare Si collectors, alpha-cyano-4-hydroxycinnamic acid (CHCA) was selected for further experiments. The mass spectra and depth profiles obtained from the organic layer prior to and after the sputter-transfer of CHCA were compared, along with those obtained from regular ME-SIMS samples (dried droplets) and, finally, with MALDI data for the same matrix-analyte combinations. Signal amplification factors were calculated by dividing the integrated molecular intensities obtained with or without matrix transfer. While the amplification factors are between 0.5 and 2 for molecules already detected with high intensities in SIMS, such as cholesterol or human angiotensin, other compounds show very large integrated signal amplification, even above two orders of magnitude. This is the case for d-glucose and cardiolipin, for which the molecular ion intensity is low (or very low) under normal SIMS analysis conditions. For such low ionization probability compounds, the beneficial effect of the matrix is unquestionable. Test experiments on mouse brain tissue sections also indicate signal enhancement with the matrix, especially for high mass lipid ions.

The number of open access databases containing experimental and predicted collision cross section (CCS) values is rising and leads to their increased use for compound identification. However, the reproducibility of reference values with different instrumental designs and the comparison between predicted and experimental CCS values is still under evaluation. This study compared experimental CCS values of 56 small molecules (Contaminants of Emerging Concern) acquired by both drift tube (DT) and travelling wave (TW) ion mobility mass spectrometry (IM-MS). The TWIM-MS included two instrumental designs (Synapt G2 and VION). The experimental TWCCSN2 values obtained by the TWIM-MS systems showed absolute percent errors (APEs) < 2% in comparison to experimental DTIMS data, indicating a good correlation between the datasets. Furthermore, TWCCSN2 values of [M − H]- ions presented the lowest APEs. An influence of the compound class on APEs was observed. The applicability of prediction models based on artificial neural networks (ANN) and multivariate adaptive regression splines (MARS), both built using TWIM-MS data, was investigated for the first time for the prediction of DTCCSN2 values. For [M+H]+ and [M − H]- ions, the 95th percentile confidence intervals of observed APEs were comparable to values reported for both models indicating a good applicability for DTIMS predictions. For the prediction of DTCCSN2 values of [M+Na]+ ions, the MARS based model provided the best results with 73.9% of the ions showing APEs below the threshold reported for [M+Na]+. Finally, recommendations for database transfer and applications of prediction models for future DTIMS studies are made. L. Belova acknowledges funding through a Research Foundation Flanders (FWO) fellowship (11G1821N). L. Bijlsma acknowledges his fellowship funded by “la Caixa” Foundation. The project that gave rise to these results also received the support of a fellowship from “la Caixa” Foundation (ID 10 0 010434). The fellowship code is LCF/BQ/PR21/11840012. Jesse Sterckx is acknowledged for helping with the measurements on the Synapt G2 system. This work received financial support also from the University Jaume I (UJI-B2020-19). The graphical abstract was created with BioRender.com, license no 2641–5211.

Although long chained PFASs have been phased-out in several countries, their persistence in the environment and bioaccumulative potential cause the environmental and biotic concentrations to remain high, highlighting the need to further monitor these pollutants. Currently several methods are used for the quantification of perfluoroalkyl substances (PFASs) in biological matrices including different ways to correct for recovery losses, each with its specific pros and contras. With this paper we aim to re-evaluate current methodologies and to create an updated new analytical guideline that is applicable for both abiotic and biotic matrices. The developed LC/MS/MS method was validated and shown to be specific, selective, linear, robust and sensitive. Reliable results could still be obtained 6 days after extraction. The recoveries varied, depending on the matrix, between 1% and 100%, but nevertheless, a high accuracy was obtained even at the lowest recoveries. A reduction of sample mass could significantly increase method recoveries and therefore it is highly recommended to take less matrix. We confirmed that using the ISTD closest in terms of functional group and carbon chain length is a suitable method for the quantification of PFASs that lack a corresponding ISTD. The newly described method was, depending on the matrix, similar in terms of sensitivity and reliability compared to a frequently used method and could be used simultaneously in future monitoring studies. Therefore, we recommend to select the purification method based on the target analytes as well as the sample matrix. Capsule The newly described method was similar in terms of sensitivity and reliability compared to a frequently used method and a selection of purification methods should be based on the target analyte and sample matrix.

A green, efficient and cheap demethylation reaction of aromatic methyl ethers with mineral acid (HCl or H2SO4) as a catalyst in high temperature pressurized water provided the corresponding aromatic alcohols (phenols, catechols, pyrogallols) in high yield. 4-Propylguaiacol was chosen as a model, given the various applications of the 4-propylcatechol reaction product. This demethylation reaction could be easily scaled and biorenewable 4-propylguaiacol from wood and clove oil could also be applied as a feedstock. Greenness of the developed method versus state-of-the-art demethylation reactions was assessed by performing a quantitative and qualitative Green Metrics analysis. Versatility of the method was shown on a variety of aromatic methyl ethers containing (biorenewable) substrates, yielding up to 99% of the corresponding aromatic alcohols, in most cases just requiring simple extraction as work-up.

Understanding the fundamental mechanisms of oxidative ageing in bitumen is considered of paramount importance in order to take steps towards durable binders able to tackle distresses related to this phenomenon which deteriorates the asphalt performance. This paper focuses on the identification of the intermediate and final oxygenated products after short- and long-term laboratory ageing simulated with rolling thin-film oven testing (RTFOT) and pressurised ageing vessel (PAV) respectively. Three binders were investigated in this study, two originated from the same wax-free crude source, while the third was obtained from a different source, containing natural wax, and followed a different manufacturing process. Fourier-Transform Infrared (FTIR) spectroscopy demonstrated a clear increase of the sulfoxide and carbonyl functional groups upon ageing for all the binders independently of origin, manufacturing or performance. Electron Paramagnetic Resonance (EPR) spectroscopy showed an increase of the organic carbon-centred radicals after short-term ageing (RTFOT), whereas after PAV these radicals remained constant in the two wax-free binders originating from the same crude source, and even decreased for the third, waxy binder. Proton Nuclear Magnetic Resonance (1H NMR) spectroscopy reported differences in the relative distribution of protons between the binders in the unaged state, and similar minor changes after both ageing steps regardless of the binder's crude source and distillation. The results of Time-of-Flight Secondary Ion Mass Spectrometry (TOF-SIMS) revealed that SOx- and (OH)x-containing compounds are produced after the sequentially occurring short- and long-term ageing in both wax-free bitumens, whereas an almost constant behaviour of aliphatics after PAV ageing can be seen for the same bitumens. Finally, the strengths and weaknesses of each of these experimental techniques were reviewed and compared versus the obtained results and possible ageing mechanisms.

The intrinsically disordered protein &alpha, synuclein plays a major role in Parkinson&rsquo, s disease. The protein can oligomerize resulting in the formation of various aggregated species in neuronal cells, leading to neurodegeneration. The interaction of &alpha, synuclein with biological cell membranes plays an important role for specific functions of &alpha, synuclein monomers, e.g., in neurotransmitter release. Using different types of detergents to mimic lipid molecules present in biological membranes, including the presence of Ca2+ ions as an important structural factor, we aimed to gain an understanding of how &alpha, synuclein interacts with membrane models and how this affects the protein conformation and potential oligomerization. We investigated detergent binding stoichiometry, affinity and conformational changes of &alpha, synuclein taking detergent concentration, different detergent structures and charges into account. With native nano-electrospray ionization ion mobility-mass spectrometry, we were able to detect unique conformational patterns resulting from binding of specific detergents to &alpha, synuclein. Our data demonstrate that &alpha, synuclein monomers can interact with detergent molecules irrespective of their charge, that protein-micelle interactions occur and that micelle properties are an important factor.

The identification of unknown molecules has been one of the cornerstone applications of mass spectrometry for decades. This tutorial reviews the basics of the interpretation of electrospray ionization-based MS and MS/MS spectra in order to identify small-molecule analytes (typically below 2000 Da). Most of what is discussed in this tutorial also applies to other atmospheric pressure ionization methods like atmospheric pressure chemical/photoionization. We focus primarily on the fundamental steps of MS-based structural elucidation of individual unknown compounds, rather than describing strategies for large-scale identification in complex samples. We critically discuss topics like the detection of protonated and deprotonated ions ([M + H]+ and [M − H]−) as well as other adduct ions, the determination of the molecular formula, and provide some basic rules on the interpretation of product ion spectra. Our tutorial focuses primarily on the fundamental steps of MS-based structural elucidation of individual unknown compounds (eg, contaminants in chemical production, pharmacological alteration of drugs), rather than describing strategies for large-scale identification in complex samples. This tutorial also discusses strategies to obtain useful orthogonal information (UV/Vis, H/D exchange, chemical derivatization, etc) and offers an overview of the different informatics tools and approaches that can be used for structural elucidation of small molecules. It is primarily intended for beginning mass spectrometrists and researchers from other mass spectrometry sub-disciplines that want to get acquainted with structural elucidation are interested in some practical tips and tricks.

Pretargeted positron emission tomography (PET) imaging based on the bioorthogonal inverse-electron-demand Diels–Alder reaction between tetrazines (Tz) and trans-cyclooctenes (TCO) has emerged as a promising tool for solid tumor imaging, allowing the use of short-lived radionuclides in immune-PET applications. With this strategy, it became possible to achieve desirable target-to-background ratios and at the same time to decrease the radiation burden to nontargeted tissues because of the fast clearance of small PET probes. Here, we show the synthesis of novel 18F-labeled dTCO-amide probes for pretargeted immuno-PET imaging. The PET probes were evaluated regarding their stability, reactivity toward tetrazine, and pharmacokinetic profile. [18F]MICA-213 showed an extremely fast kinetic rate (10,553 M–1 s–1 in 50:50 MeOH/water), good stability in saline and plasma up to 4 h at 37 °C with no isomerization observed, and the biodistribution in healthy mice revealed a mixed hepatobiliary and renal clearance with no defluorination and low background in other tissues. [18F]MICA-213 was further used for in vivo pretargeted immune-PET imaging carried out in nude mice bearing LS174T colorectal tumors that were previously treated with a tetrazine-modified anti-TAG-72 monoclonal antibody (CC49). Pretargeted μPET imaging results showed clear visualization of the tumor tissue with a significantly higher uptake when compared to the control.

The identification of peptides and proteins from tandem mass spectra is a difficult task and multiple tools have been developed to aid this identification. We present a new method called quantum chemical mass spectrometry for materials science (QCMS

A new approach to synthesize valuable 3,4- dialkoxyanilines and alkyl propionates from lignin-derived 4- propylguaiacol and -catechol with overall isolated yields up to 65% has been described. The strategy is based on the introduction of nitrogen via a Beckmann rearrangement. Amino introduction therefore coincides with a C-defunctionalization reaction; overall a replacement of the propyl chain by an amino group is obtained. The process only requires cheap bulk chemicals as reagents/reactants and does not involve column chromatography to purify the reaction products. Furthermore, all carbon atoms from the biorenewable lignin-derived monomers are transformed into valuable compounds. Greenness was assessed by performing a Green Metrics analysis on two dialkoxyanilines. A comparison was made with literature routes for these compounds starting from a petrochemical substrate. ispartof: ACS Sustainable Chemistry & Engineering vol:7 issue:7 pages:6906-6916 status: published

Rationale Melphalan is a frequently used chemotherapeutical agent for the treatment of myeloma, breast cancer, ovarian cancer and sarcoma of soft tissue. A good knowledge of the reactivity of the drug toward the different amino acids, e.g. covalent adduct formation, is crucial for the understanding of its activity and side effects during cancer treatment. Methods The reactivity of melphalan and sites of adduct formation were studied by in vitro incubation of melphalan with free amino acids and glutathione as a model peptide. The formed covalent adducts were investigated using ultra-performance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) using a triple-quadrupole instrument. Accurate mass measurements for the confirmation of characteristic product ions were performed on a quadrupole time-of-flight (QTOF) mass spectrometer. Results The incubation of melphalan with different classes of amino acids resulted in the formation of adducts on the amino and carboxyl termini, as well as adduct formation in the reactive side chains of Cys, Met, Tyr, His, Lys, Asp and Glu. All these melphalan adducts could be identified by their characteristic collision-induced dissociation (CID) product ion patterns. Conclusions The present study demonstrates the reactivity of melphalan towards the functional groups of amino acids. The different alkylation site products show distinctive fragmentation patterns, which enable a fast identification of the different melphalan adducts. This study is a first important step towards a better understanding of the adduct formation in more complex molecules, e.g. peptides and proteins. Copyright © 2016 John Wiley & Sons, Ltd.

Electrospray ion mobility-mass spectrometry (IM-MS) data show that for some small molecules, two (or even more) ions with identical sum formula and mass, but distinct drift times are observed. In spite of showing their own unique and characteristic fragmentation spectra in MS/MS, no configurational or constitutional isomers are found to be present in solution. Instead the observation and separation of such ions appears to be inherent to their gas-phase behaviour during ion mobility experiments. The origin of multiple drift times is thought to be the result of protonation site isomers ('protomers'). Although some important properties of protomers have been highlighted by other studies, correlating the experimental collision cross-sections (CCSs) with calculated values has proven to be a major difficulty. As a model, this study uses the pharmaceutical compound melphalan and a number of related molecules with alternative (gas-phase) protonation sites. Our study combines density functional theory (DFT) calculations with modified MobCal methods (e.g. nitrogen-based Trajectory Method algorithm) for the calculation of theoretical CCS values. Calculated structures can be linked to experimentally observed signals, and a strong correlation is found between the difference of the calculated dipole moments of the protomer pairs and their experimental CCS separation.

This work presents a strategy for elucidation of unknown migrants from plastic food contact materials (baby bottles) using a combination of analytical techniques in an untargeted approach. First, gas chromatography (GC) coupled to mass spectrometry (MS) in electron ionisation mode was used to identify migrants through spectral library matching. When no acceptable match was obtained, a second analysis by GC-(electron ionisation) high resolution mass spectrometry time of flight (TOF) was applied to obtain accurate mass fragmentation spectra and isotopic patterns. Databases were then searched to find a possible elemental composition for the unknown compounds. Finally, a GC hybrid quadrupole-TOF-MS with an atmospheric pressure chemical ionisation source was used to obtain the molecular ion or the protonated molecule. Accurate mass data also provided additional information on the fragmentation behaviour as two acquisition functions with different collision energies were available (MS(E) approach). In the low-energy function, limited fragmentation took place, whereas for the high-energy function, fragmentation was enhanced. For less volatile unknowns, ultra-high pressure liquid chromatography-quadrupole-TOF-MS was additionally applied. Using a home-made database containing common migrating compounds and plastic additives, tentative identification was made for several positive findings based on accurate mass of the (de)protonated molecule, product ion fragments and characteristic isotopic ions. Six illustrative examples are shown to demonstrate the modus operandi and the difficulties encountered during identification. The combination of these techniques was proven to be a powerful tool for the elucidation of unknown migrating compounds from plastic baby bottles.

The synthetic utility of electron-deficient alpha,alpha,gamma-trichloroaldimines was demonstrated by an indium(III) triflate-catalyzed cascade reaction with terminal alkynes allowing one to rapidly and selectively access 2-alkynyl-3,3-dichloropyrrolidines in good to excellent yields. The reaction proceeds in a single synthetic operation via an addition of acetylenes to alpha,alpha,gamma-trichloroaldimines, followed by a spontaneous cyclization of the in situ formed trichloropropargylic amines. The dichloromethylene moiety of the aldimine acts as an activating group to accomplish this transformation under very mild conditions. A broad variety of both aryl and alkyl acetylenes, as well as primary and secondary nitrogen substituents in the imine are well tolerated. The dichloromethylene group, which is conserved in the 2-alkynylpyrrolidine enhances the synthetic value of these pyrrolidines and allowed their conversion to (E/Z)-2-alkenyl-3-chloropyrroles by a base induced monodechlorination.

Molecular dynamics (MD) simulations were performed to provide atomic scale insight in the initial interaction between hydroxyl radicals (OH) and peptide systems in solution. These OH radicals are representative reactive oxygen species produced by cold atmospheric plasmas. The use of plasma for biomedical applications is gaining increasing interest, but the fundamental mechanisms behind the plasma modifications still remain largely elusive. This study helps to gain more insight in the underlying mechanisms of plasma medicine but is also more generally applicable to peptide oxidation, of interest for other applications. Combining both reactive and nonreactive MD simulations, we are able to elucidate the reactivity of the amino acids inside the peptide systems and their effect on their structure up to 1 μs. Additionally, experiments were performed, treating the simulated peptides with a plasma jet. The computational results presented here correlate well with the obtained experimental data and highlight the importance of the chemical environment for the reactivity of the individual amino acids, so that specific amino acids are attacked in higher numbers than expected. Furthermore, the long time scale simulations suggest that a single oxidation has an effect on the 3D conformation due to an increase in hydrophilicity and intra- and intermolecular interactions.

An iron-catalyzed aerobic oxidation of (alkyl)(aryl)azinylmethanes has been developed leading to tertiary alcohols in moderate to good yields. Hock rearrangement was identified as a major side reaction leading to a complex mixture of undesired products. Addition of thiourea sometimes allows inhibiting this side reaction and steers the reaction towards the desired products.

The identification of peptides and proteins from tandem mass spectra is a difficult task and multiple tools have been developed to aid this identification. We present a new method called quantum chemical mass spectrometry for materials science (QCMS(2)), which is based on quantum chemical calculations of bond orders, reaction, and transition-state energies at the DFT/B3LYP/6-311+G* level of theory. The method was used to describe the fragmentation pathways of five X-His-Ser tripeptides with X = Asn, Asp, Glu, Ser, and Trp, thereby focusing on the influence of the side chain and inter-side-chain interactions on the fragmentation. The main features in the mass spectra of the five tripeptides were correctly reproduced, and a number of fragments were assigned to fragmentations involving the side chain and the influence of inter-side-chain interactions. Product ion spectra were recorded to evaluate the capabilities and limitations of QCMS(2) and a number of conventional tools.

RATIONALE Melphalan is a widely used cytotoxic agent in cancer treatments. This phenylalanine analog has been shown an effective drug in the treatment of breast cancer, multiple myeloma and melanoma of the extremities. A good knowledge of the drug's degradation and metabolism are crucial for understanding its activity during cancer treatments. METHODS The formation of hydrolysis products of melphalan is studied using ultra-performance liquid chromatography (UPLC) tandem mass spectrometry (MS/MS). Aqueous melphalan solutions were incubated at elevated temperatures and analyzed by UPLC/MS/MS. Two previously described hydrolysis products, mono- and dihydroxymelphalan (MOH and DOH), were formed in vitro and could be characterized during MS/MS and high-resolution experiments. RESULTS Novel compounds with m/z values >500 Da were discovered. Comparison of the fragmentation patterns of these new molecules with those of MOH and DOH show great similarities. The higher masses are explained by the presence of two or more melphalan units. In total, more than 15 new hydrolysis products were found. Experiments were set up to study the formation and the chemical structures of these molecules. CONCLUSIONS The hydrolysis of melphalan is studied in the scope of a phase II clinical trial (isolated lung perfusion, ILuP). Patient samples were screened for the presence of all documented and novel melphalan hydrolysis products. This study reports the formation of a new class of oligomeric compounds in both in vivo and in vitro samples.

The use of traveling wave ion mobility mass spectrometry (TWIMS) is evaluated in conjunction with, and as a possible alternative to, conventional LC-MS(/MS) methods for the separation and characterization of drug-like compounds and metabolites. As a model system we use an in vitro incubation mixture of the chemotherapeutic agent melphalan, which results in more than ten closely related hydrolysis products and chain-like oligomers. Ion mobility as a filtering tool results in the separation of ions of interest from interfering ions, based on charge state and shape/size. Different classes of chemical compounds often display different mobilities even if they show the same LC behavior - thereby providing an orthogonal separation dimension. Small molecules with identical or similar m/z that only differ in shape/size (e.g. isomers and isobars, monomers/dimers) can also be distinguished using ion mobility. Similar to retention times and mass-to charge ratios, drift times are analyte-dependent and can be used as an additional identifier. We find that the compound melphalan shows two different drift times due to the formation of gas phase charge isomers (protomers). The occurrence of protomers has important implications for ion mobility characterization of such analytes, and also for the interpretation of their fragmentation behavior (CID) in the gas phase. (C) 2017 Elsevier B.V. All rights reserved.

Mannosylerythritol lipids (MELs) are one of the most promising biosurfactants because of their high fermentation yields (>100 g l−1) and during the last two decades they have gained a lot of attention due to their interesting self-assembling properties and biological activities. In this study, MELs were produced by fed-batch bioreactor fermentation of rapeseed oil with Pseudozyma aphidis MUCL 27852. This high-level MEL-producing yeast secretes four conventional MEL structures, -A, -B, -C and -D, which differ in their degree of acetylation. During our research, unknown compounds synthesized by P. aphidis were detected by thin-layer chromatography. The unknown compounds were separated by flash chromatography and identified as tri-acylated MELs by high-performance liquid chromatography tandem mass spectrometry (HPLC–MS/MS). The third fatty acid chain on the tri-acylated MELs was positioned on the primary alcohol of the erythritol moiety and comprised long-chain acids, mainly oleic and linoleic acid, which are not found in conventional di-acylated MELs. Furthermore, the LC–MS analysis time of conventional MELs was reduced to almost one-third by switching from HPLC–MS/MS to ultraperformance liquid chromatography tandem mass spectrometry (UPLC–MS/MS). Provided optimization of the fermentation yield, P. aphidis could be an interesting novel producer of tri-acylated MELs and, thereby expand the supply and applicability of biosurfactants.

The total 80% EtOH extract of stem bark of Nauclea pobeguinii (Rubiaceae), which is active against uncomplicated falciparum malaria as shown in previous clinical studies, was analysed by means of UPLC-MS and HPLC-SPE-NMR. Apart from the main constituent, strictosamide, a series of minor constituents was identified, including two new iridoid-indole alkaloids, i.e. naucleidinic acid and 19-O-methyl-3,14-dihydroangustoline, together with 8 known iridoid-indole alkaloids, i.e. naucleidinal, magniflorine, naucleofficine D, two diastereoisomers of 3,14-dihydroangustoline, strictosidine, desoxycordifoline, 3 alpha,5 alpha-tetrahydrodeoxycordifoline lactam, and a phenol glycoside 3,4,5-trimethoxyphenol beta-D-apiofuranosyl-(1-6)-beta-D-glucopyranoside (kelampayoside A). (C) 2012 Phytochemical Society of Europe. Published by Elsevier B. V. All rights reserved.

The morphological, physiological and behavioural differences between solitarious and gregarious desert locusts are so pronounced that one could easily mistake the two phases as belonging to different species, if one has no knowledge of the phenomenon of phenotypic plasticity. A number of phase-specific features are hormonally controlled. Juvenile hormone promotes several solitarious features, the green cuticular colour being the most obvious one. The neuropeptide corazonin elicits the dark cuticular colour that is typical for the gregarious phase, as well as particular gregarious behavioural characteristics. However, it had to be concluded, for multiple reasons, that the endocrine system is not the primary phase-determining system. Our observation that longevity gets imprinted in very early life by crowding of the young hatchlings, and that it cannot be changed thereafter, made us consider the possibility that, perhaps, epigenetic control of gene expression might be, if not the missing, a primary phase-determining mechanism. Imprinting is likely to involve DNA methylation and histone modification. Analysis of a Schistocerca EST database of nervous tissue identified the presence of several candidate genes that may be involved in epigenetic control, including two DNA methyltransferases (Dnmts). Dnmt1 and Dnmt2 are phase-specifically expressed in certain tissues. In the metathoracic ganglion, important in the serotonin pathway for sensing mechanostimulation, their expression is clearly affected by crowding. Our data urge for reconsidering the role of the endocrine system as being sandwiched in between genetics and epigenetics, involving complementary modes of action.

In our earlier LC–MS experiments on the analysis of phosphocompounds like nucleotides (mono-, di- and triphosphate) and phosphopeptides and in literature, low sensitivity and severe losses of analyte to the instrumental setup were observed. Since we noticed that the stainless steel parts of the setup (e.g., the electrospray needle) adsorbed important quantities of the phosphorylated analytes, we made a LC–ESI setup without metallic surfaces. The first results were disappointing since also the fused silica surface of the LC-ESI coupler adsorbed part of the nucleotides and phosphopeptides injected in flow analysis experiments. We present experiments documenting the contribution of the different components of the setup. A number of potential solutions to the adsorption problem are proposed and tested. Only dimethyldichlorosilane deactivation of fused silica capillaries gave satisfactory results as adsorption of nucleotides and phosphopeptide was minimised.

Mannosylerythritol lipids (MELs) are currently one of the most promising biosurfactants because of their multifunctional applications and good biodegradability. Depending on the yeast strain and the feedstock used for the fermentation process, structural variations in the MELs obtained occur. Therefore, MELs produced by Pseudozyma aphidis DSMZ 70725 with a soybean oil feedstock were characterized by chromatography and mass spectrometry (MS). Column chromatography with silica provided fractionation of the different types of MEL. High-performance liquid chromatography combined with MS was employed for the analysis of the MEL fractions and crude mixtures. A characteristic MS pattern for the MELs was obtained and indications of the presence of new MEL homologues, showing the incorporation of longer and more unsaturated fatty acid chains than previously reported, were given. Gas chromatographyMS analysis confirmed the presence of such unsaturated fatty acid chains in the MELs, demonstrating the incorporation of fatty acids with lengths ranging from C8 to C14 and with up to two unsaturations per chain. The incorporation of C16 and C18 fatty acid chains requires further investigation. MS/MS data allowed the unambiguous identification of the fatty acids present in the MELs. The product ion spectra also revealed the presence of a new isomeric class of MELs, bearing an acetyl group on the erythritol moiety.

D-ring chloro-substituted neocryptolepines have been synthesized in excellent yield starting from 3-bromo-2-chloro-1-methylquinolinium triflate via a one-pot condensation—Pd-catalyzed intramolecular direct arylation strategy involving chloroanilines. The 3-bromo-2-chloro-1-methylquinolinium triflate was obtained via methylation of commercial 3-bromo-2-chloroquinoline with methyl triflate.

A novel class of self-doping conjugated oligomers, E,E-2-(sulfoalkoxy)-5-alkoxy-1,4-bis[2-(2,4,6-trimethoxyphenyl) ethenyl]benzenes, is presented. The synthesis and spectroscopic characterisation of five such oligomers are described, and an electrochemical analysis using cyclic voltammetry is performed to determine the anodic peak potentials. A structural study is performed on six self-doping oligomers in which the structures and energies of the possible mono-molecular forms of the electrically conducting doped material are described and evaluated using Hirshfeld charges and the Quantum Theory of Atoms In Molecules. Copyright © 2009 John Wiley & Sons, Ltd.

Four furanoditerpenoids were isolated from roots of Croton jatrophoides (Euphorbiaceae) collected in Tanzania. In addition to the known compounds penduliflaworosin and teucvin (mallotucin A), a new teucvin isomer, which was named isoteucvin, and a furanoditerpenoid with a new skeleton, for which the name jatrophoidin was adopted, were isolated. Their structures were elucidated by spectroscopic methods such as ESI-MS and NMR, including H-1-, C-13-, and two-dimensional NMR. The crystal structures of isoteucvin and jatrophoidin were solved using single-crystal X-ray diffraction, by which we also established the absolute configuration of jatrophoidin. The refined crystal structure of isoteucvin has the same (absolute) configuration as jatrophoidin, although the X-ray diffraction data of isoteucvin were not conclusive with respect to the absolute configuration.

An automated online immobilized metal affinity chromatography/high-performance liquid chromatography mass spectrometric (IMAC-HPLC/MS/MS) method was developed to study cytidine 3',5'-cyclic monophosphate (cCMP)-specific protein phosphorylation, analogous to a previously successful offline IMAC method using microvolume IMAC pipette tips. The optimized method identified murine brain phosphoproteins selectively modified by challenge with cCMP, using manual interpretation of the results to confirm both phosphorylation and selectivity of response to cCMP. A number of proteins identified by this strategy have potential roles in hyperproliferation, a previously reported response to elevated levels of cCMP.

Urinary modified nucleosides have a potential role as cancer biomarkers, and most of the methods used in their study have utilized low-pressure phenylboronate affinity chromatography materials for the purification of the cis-diol-containing nucleosides. In this study, a boronate HPLC column was surprisingly shown not to trap the nucleosides as would be expected from experience with the classic Affigel 601 resin but showed only partial selectivity toward cis-diol groups while other groups exhibited better retention. In aprotic conditions, trapping of nucleosides was possible; however, the selectivity toward cis-diol-containing compounds was lost with the Lewis basicity of available nitrogens being the main determinant of retention. The experimental findings are compared to and confirmed by DFT calculations.

Melphalan is a frequently used chemotherapeutical agent for the treatment of myeloma, breast cancer, ovarian cancer and sarcoma of soft tissue. A good knowledge of the reactivity of the drug toward the different amino acids, e.g. covalent adduct formation, is crucial for the understanding of its activity and side effects during cancer treatment.The reactivity of melphalan and sites of adduct formation were studied by in vitro incubation of melphalan with free amino acids and glutathione as a model peptide. The formed covalent adducts were investigated using ultra-performance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) using a triple-quadrupole instrument. Accurate mass measurements for the confirmation of characteristic product ions were performed on a quadrupole time-of-flight (QTOF) mass spectrometer.The incubation of melphalan with different classes of amino acids resulted in the formation of adducts on the amino and carboxyl termini, as well as adduct formation in the reactive side chains of Cys, Met, Tyr, His, Lys, Asp and Glu. All these melphalan adducts could be identified by their characteristic collision-induced dissociation (CID) product ion patterns.The present study demonstrates the reactivity of melphalan towards the functional groups of amino acids. The different alkylation site products show distinctive fragmentation patterns, which enable a fast identification of the different melphalan adducts. This study is a first important step towards a better understanding of the adduct formation in more complex molecules, e.g. peptides and proteins.

An operationally simple and efficient method for the synthesis of 2-alkynyl-3,3-dichloropyrrolidines from imines and readily available acetylenes is developed using indium(III) triflate as a Lewis acid catalyst.

This work presents a strategy for elucidation of unknown migrants from plastic food contact materials (baby bottles) using a combination of analytical techniques in an untargeted approach. First, gas chromatography (GC) coupled to mass spectrometry (MS) in electron ionisation mode was used to identify migrants through spectral library matching. When no acceptable match was obtained, a second analysis by GC-(electron ionisation) high resolution mass spectrometry time of flight (TOF) was applied to obtain accurate mass fragmentation spectra and isotopic patterns. Databases were then searched to find a possible elemental composition for the unknown compounds. Finally, a GC hybrid quadrupole-TOF-MS with an atmospheric pressure chemical ionisation source was used to obtain the molecular ion or the protonated molecule. Accurate mass data also provided additional information on the fragmentation behaviour as two acquisition functions with different collision energies were available (MS(E) approach). In the low-energy function, limited fragmentation took place, whereas for the high-energy function, fragmentation was enhanced. For less volatile unknowns, ultra-high pressure liquid chromatography-quadrupole-TOF-MS was additionally applied. Using a home-made database containing common migrating compounds and plastic additives, tentative identification was made for several positive findings based on accurate mass of the (de)protonated molecule, product ion fragments and characteristic isotopic ions. Six illustrative examples are shown to demonstrate the modus operandi and the difficulties encountered during identification. The combination of these techniques was proven to be a powerful tool for the elucidation of unknown migrating compounds from plastic baby bottles.

A study of the interaction of phosphorylated organic compounds with the stainless components of a liquid chromatography-electrospray ionisation-mass spectrometry system (LC-ESI-MS) was carried out to disclose a (forgotten?) likely pitfall in the LC-ESI-MS analysis of phosphorylated compounds. The retention behaviour of some representative compounds of different important classes of phosphorylated biomolecules such as nucleotides, oligonucleotides, phosphopeptides, phospholipids and phosphorylated sugars was investigated during their passage through the injector and the stainless steel electrospray capillary. It became clear that the stainless steel components within the LC-ESI-MS setup were able to retain and trap phosphorylated compounds when these compounds were introduced under acidic conditions (0.1% acetic acid). Their release from these stainless steel parts was accomplished by applying an extreme basic mobile phase (25-50% ammonium hydroxide, ca. pH 12). From the data collected one could conclude that the availability of a primary phosphate group appeared imperative but was not always sufficient to realise adsorption on a stainless surface. Furthermore, the number of phosphate moieties seemed to enhance the adsorption properties of the molecules and hence roughly correlated with the analyte fraction lost. Corrosion of the inner surface caused by the mobile phase and the electrospray process was found to be an important factor in the course of these adsorption phenomena.

The development of an on-column focusing gradient capillary LC method coupled to tandem mass spectrometry (quadrupole-linear ion trap) for the quantitative determination of the anticancer agent ZD1839 (Gefitinib, Iressa) in blood plasma is described. Plasma samples (0.2 ml) were extracted with methyl tert-butyl ether. The analytes of interest, ZD1839 and the internal standard [(2)H8]ZD1839 (ZD1839-d8) were eluted on a 50 mm x 1 mm, 5 microm particle size, capillary ODS Hypersil column using an aqueous ammonium acetate gradient at 40 microl/min. Mass spectrometric detection was performed by a Q-Trap tandem mass spectrometer with electrospray positive ionisation, and monitored in the multiple reaction monitoring transitions 447 >128 and 455 >136, respectively. The limit of quantification of ZD18395 was 0.1 ng/ml. The method proved to be robust, allowing quantification of ZD1839 with sufficient precision, accuracy and sensitivity.

Calf thymus DNA was treated with melphalan, a nitrogen mustard, and the formation of melphalan cross-linked DNA adducts was investigated. These cross-linked adducts could not be detected either in the enzymatically or in the thermally generated DNA hydrolysates. However, a search for DNA cross-linked adducts in the hydrolysates obtained under acidic conditions revealed the presence of different types of cross-links, mainly containing an adenine moiety. These results are very important because they show that the detection of cross-links is dependent on the hydrolytic procedure used and that these cross-linked adducts are formed under totally different reaction conditions from those in in vivo situations. This can explain the very low abundance or even the absence of cross-linked adducts in nitrogen mustard treated animals. The generally accepted theory that the anti-cancer activity of bifunctional mustards such as melphalan is due to cross-linking of DNA strands remains therefore from our point of view questionable.

Xenobiotic and endobiotic molecules can react with DNA leading to formation of so-called DNA adducts. This modified DNA can be repaired enzymatically, but, if not, these modifications are believed to be responsible for the initiation of carcinogenic processes. Hence, we studied the interaction of 2′-deoxynucleosides and 2′-deoxynucleotides with 3,4-estronequinone (3,4-E1Q), a metabolite of estrone (E1) and a supposed carcinogen. These estrone–nucleic acid adducts were analysed by capillary liquid chromatography (CapLC) coupled to electrospray ionization mass spectrometry (ESI-MS). Knowledge of their behaviour from in vitro studies is a prerequisite for detecting adducts in in vivo studies. Our initial attempts to synthesise nucleos(t)ide adducts of 3,4-E1Q in an aprotic solvent (dimethylformamide) yielded no adducts. However, under acidic aqueous conditions, adducts were obtained. With dGuo, a dGuo adduct was found in addition to a Gua adduct. Earlier publications on adduct formation in protic solvents failed to report formation of any adduct with dAdo. A N3-Ade adduct was reported upon reaction of 3,4-E1Q with Ade base and with DNA. With dAdo, we obtained two nucleoside adducts and six Ade adducts due to loss of 2′-deoxyribose. Thus, contrary to general belief that only 2,3-E1Q can form stable adducts, we showed formation of substantial amounts of intact DNA adducts with 3,4-E1Q in addition to deglycosylated adducts. Adducts were also obtained with dGMP and dAMP, but no phosphate alkylation was found. Adducts of dCyd, dCMP, dThd, and dTMP were not detected. Using chromatographic-MS data a structural relationship between the 2′-deoxynucleoside, 2′-deoxynucleotide and base adducts was found in the various reaction mixtures. The adducts of dGuo and dGMP reaction mixtures were alkylated at the same N7-position of the nucleobase, as indicated by the occurrence of a rapid deglycosylation reaction. In dAdo and dAMP reaction mixtures, 14 adducts were detected; their relationships from the LC and MS data reduced the number of structures to six adenine base alkylated adducts with respect to alkylation between N1, N3, N7 and/or N6 in the adenine and C1, C2 and/or C6 in 3,4-E1Q. We could infer, in addition, whether they had an A ring attachment or a C6 attachment on the estrone moiety. Copyright © 2004 John Wiley & Sons, Ltd.

A liquid chromatography/mass spectrometry (LC/MS) method for the analysis of complex mixtures of nucleoside mono-, di- and triphosphates has been developed. A short capillary column (35mm × 0.3mm i.d.) was operated under ion-pair high-performance liquid chromatography conditions and hyphenated to (negative) electrospray (tandem) mass spectrometry. As such, the separation of 12 nucleotides was performed by a binary gradient elution using CH3OH/H2O and N,N-dimethylhexylamine (N,N-DMHA) as ion-pairing agent. The influence of different N,N-DMHA concentrations on the chromatographic and mass spectrometric performance was evaluated to achieve optimal LC/MS conditions. In addition it was demonstrated that a controlled admission of ammonium dihydrogen phosphate (NH4H2PO4) improved both chromatographic performance and mass spectrometric detection. Because the system was hyphenated to an orthogonal designed electrospray interface (Z-spray™), long acquisition times were possible without loss of sensitivity. Copyright © 2002 John Wiley & Sons, Ltd.

Background Mass spectrometry-based proteomics experiments generate spectra that are rich in information. Often only a fraction of this information is used for peptide/protein identification, whereas a significant proportion of the peaks in a spectrum remain unexplained. In this paper we explore how a specific class of data mining techniques termed “frequent itemset mining” can be employed to discover patterns in the unassigned data, and how such patterns can help us interpret the origin of the unexpected/unexplained peaks. Results First a model is proposed that describes the origin of the observed peaks in a mass spectrum. For this purpose we use the classical correlative database search algorithm. Peaks that support a positive identification of the spectrum are termed explained peaks. Next, frequent itemset mining techniques are introduced to infer which unexplained peaks are associated in a spectrum. The method is validated on two types of experimental proteomic data. First, peptide mass fingerprint data is analyzed to explain the unassigned peaks in a full scan mass spectrum. Interestingly, a large numbers of experimental spectra reveals several highly frequent unexplained masses, and pattern mining on these frequent masses demonstrates that subsets of these peaks frequently co-occur. Further evaluation shows that several of these co-occurring peaks indeed have a known common origin, and other patterns are promising hypothesis generators for further analysis. Second, the proposed methodology is validated on tandem mass spectrometral data using a public spectral library, where associations within the mass differences of unassigned peaks and peptide modifications are explored. The investigation of the found patterns illustrates that meaningful patterns can be discovered that can be explained by features of the employed technology and found modifications. Conclusions This simple approach offers opportunities to monitor accumulating unexplained mass spectrometry data for emerging new patterns, with possible applications for the development of mass exclusion lists, for the refinement of quality control strategies and for a further interpretation of unexplained spectral peaks in mass spectrometry and tandem mass spectrometry. Electronic supplementary material The online version of this article (doi:10.1186/s12953-014-0054-1) contains supplementary material, which is available to authorized users.

Spectral library searching is a popular approach for MS/MS-based peptide identification. Because the size of spectral libraries continues to grow, the performance of searching algorithms is an important issue. This technical note introduces a strategy based on a minimum shared peak count between two spectra to reduce the set of admissible candidate spectra when issuing a query. A theoretical validation through time complexity analysis and an experimental validation based on an implementation of the candidate reduction strategy show that the approach can achieve a reduction of the set of candidate spectra by (at least) an order of magnitude, resulting in a significant improvement in the speed of the search. Meanwhile, more than 99% of the positive search results is retained. This efficient strategy to drastically improve the speed of spectral library searching with a negligible loss of sensitivity can be applied to any current spectral library search tool, irrespective of the employed similarity metric.

The interaction of 4-hydroxy metabolites of estrogens with DNA leads to the formation of DNA adducts. These adducts are believed to play an important role in the incidence of breast and endometrial cancer. In order to be able to analyze these adducts in in vivo samples a method based upon the coupling of miniaturized liquid chromatography (LC) to electrospray tandem mass spectrometry (ES-MS/MS) was developed for the analysis of the adducts formed with 4-hydroxyequilenin. In vitro synthesized adducts obtained by the reaction of 4-hydroxyequilenin with the main 2'-deoxynucleosides were separated on a Hypersyl C(18) BDS nano-HPLC column (15 cm x 75 microm i.d.) at a flow-rate of 300 nl min(-1) using gradient elution with CH(3)OH--0.2% CH(3)COOH in H(2)O. The column was coupled, in combination with a column switching system, to a nano-electrospray interface. Analysis of the low- and high-resolution low-energy collision-activated dissociation product ion spectra of normal and deuterated adducts supported earlier data demonstrating equilenin to form different isomeric adducts, except with thymidine, for which no adducts were found. The nano-HPLC column-switching ES-MS system was tested for its sensitivity on a triple-quadrupole instrument, and detection limits down to 197 fg in the single reaction monitoring mode were obtained for semi-preparatively isolated equilenin--2'-deoxyguanosine adduct.

Calf thymus DNA was reacted in vitro with phenyl glycidyl ether (PGE) and was hydrolysed enzymatically, to the 5'-monophosphate nucleotides using deoxyribonuclease I (DNA-ase I) and nuclease P1. The adducts were concentrated using solid phase extraction (SPE), on a polystyrene divinylbenzene copolymer in order to remove the unmodified nucleotides. The adducts could be identified using capillary zone electrophoresis-electrospray tandem mass spectrometry (CZE ES-MS/MS), using sample stacking. In addition to the base alkylated 2'-deoxynucleotides present in the DNA-hydrolysate, also phosphate alkylated 2'-deoxynucleotide adducts were identified for TMP and dAMP. An additional adduct, dUMP alkylated on the uridine moiety was found originating from the hydrolytic deamination of dCMP alkylated on N3 of the cytosine moiety. Enzymatic hydrolysis using nuclease P1 was incomplete as shown by the presence of dinucleotides alkylated on the base moiety. They were successfully hydrolysed to the corresponding 2'-deoxynucleotides by snake venom phosphodiesterase (SVP). Data are shown indicating that alkylations on the pyrimidine bases were more resistant to enzymatic hydrolysis with nuclease P1 than the purine alkylated products.

Macromolecules such as deoxyribonucleic acid (DNA) and ribonucleic acid as well as their constituents play an important role in all kinds of biochemical reactions in nature. Hence their isolation and identification plays a major role in biochemical analysis. Mass spectrometry (MS) has gained an important position in this field because of the development of soft ionization techniques such as fast atom bombardment (FAB), liquid secondary mass spectrometry, thermospray ionization (TSP), the atmospheric pressure ionization techniques, electrospray ionization and atmospheric pressure chemical ionization and matrix assisted laser desorption. Because of their polar nature, mixtures of nucleosides, nucleotides and oligonucleotides are wel separated by liquid chromatography (LC) and electrophoretic techniques. Therefore it is not surprising to note that a lot of effort has been put into the development of LC–MS methods for the analysis of these compounds. In this review, covering the period 1990–1996, the LC–MS analysis of nucleobases, nucleosides, nucleotides, oligonucleotides and DNA adducts by TSP, continuous flow FAB and electrospray MS is discussed.

A phytochemical investigation of Striga asiatica and the leaves of Sauropus spatulifolius revealed seventeen compounds, three of which were reported for the first time from nature, i.e. two alkaloids N-hydroxyethyl-2-acetylpyrrole, N-(3-carboxypropyl)-2-acetylpyrrole, and 2-(4-hydroxy-2,2,6-trimethylcyclohexyl)acetic acid, for which the name sauropic acid was adopted. Their structures were established spectroscopically and by single-crystal X-ray diffraction analysis. All extracts and isolates were evaluated for antiplasmodial activity against Plasmodium falciparum strain K1 and for cytotoxicity against MRC-5 cells.

Extracts of urinary nucleosides have been sequentially purified and examined by mass spectrometric analysis. Seventeen modified nucleosides have been unequivocally identified and a further five provisionally identified. While several nucleosides were found only in a small number of extracts, the occurrence and levels of others were found to correlate with the tumour type and stage.