Validation of a confirmatory method for lipophilic marine toxins in shellfish using UHPLC-HR-Orbitrap MS

Lipophilic marine toxins are produced by harmful microalgae and can accumulate in edible filter feeders such as shellfish, leading to an introduction of toxins into the human food chain, causing different poisoning effects. During the last years, analytical methods, based on liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), have been consolidated by interlaboratory validations. However, the main drawback of LC-MS/MS methods remains the limited number of compounds that can be analyzed in a single run. Due to the targeted nature of these methods, only known toxins, previously considered during method optimization, will be detected. Therefore in this study, a method based on ultra-high-performance liquid chromatography coupled to high-resolution Orbitrap mass spectrometry (UHPLC-HR-Orbitrap MS) was developed. Its quantitative performance was evaluated for confirmatory analysis of regulated lipophilic marine toxins in shellfish flesh according to Commission Decision 2002/657/EC. Okadaic acid (OA), dinophysistoxin-1 (DTX-1), pectenotoxin-2 (PTX-2), azaspiracid-1 (AZA-1), yessotoxin (YTX), and 13-desmethyl spirolide C (SPX-1) were quantified using matrix-matched calibration curves (MMS). For all compounds, the reproducibility ranged from 2.9 to 4.9 %, repeatability from 2.9 to 4.9 %, and recoveries from 82.9 to 113 % at the three different spiked levels. In addition, confirmatory identification of the compounds was effectively performed by the presence of a second diagnostic ion (¹³C). In conclusion, UHPLC-HR-Orbitrap MS permitted more accurate and faster detection of the target toxins than previously described LC-MS/MS methods. Furthermore, HRMS allows to retrospectively screen for many analogues and metabolites using its full-scan capabilities but also untargeted screening through the use of metabolomics software. Figure

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Interpretation of NMR Relaxation as a Tool for Characterising the Adsorption Strength of Liquids inside Porous Materials

Nuclear magnetic resonance (NMR) relaxation times are shown to provide a unique probe of adsorbate–adsorbent interactions in liquid‐saturated porous materials. A short theoretical analysis is presented, which shows that the ratio of the longitudinal to transverse relaxation times (T₁/T₂) is related to an adsorbate–adsorbent interaction energy, and we introduce a quantitative metric e_surf (based on the relaxation time ratio) characterising the strength of this surface interaction. We then consider the interaction of water with a range of oxide surfaces (TiO₂ anatase, TiO₂ rutile, γ‐Al₂O₃, SiO₂, θ‐Al₂O₃ and ZrO₂) and show that e_surf correlates with the strongest adsorption sites present, as determined by temperature programmed desorption (TPD). Thus we demonstrate that NMR relaxation measurements have a direct physical interpretation in terms of the characterisation of activation energy of desorption from the surface. Further, for a series of chemically similar solid materials, in this case a range of oxide materials, for which at least two calibration values are obtainable by TPD, the e_surf parameter yields a direct estimate of the maximum activation energy of desorption from the surface. The results suggest that T₁/T₂ measurements may become a useful addition to the methods available to characterise liquid‐phase adsorption in porous materials. The particular motivation for this work is to characterise adsorbate–surface interactions in liquid‐phase catalysis.     

High-resolution Orbitrap mass spectrometry for the analysis of carotenoids in tomato fruit: validation and comparative evaluation towards UV–VIS and tandem mass spectrometry

In this study, a generic extraction protocol and full-scan high-resolution Orbitrap-mass spectrometry (MS) detection method were developed, enabling the metabolomic screening for carotenoids in tomato fruit tissue. To this end, the carotenoids lutein, zeaxanthin, α-carotene, β-carotene, and lycopene (representing both xanthofylls and carotenes) were considered. The extraction procedure was optimized by means of a D-optimal design and consisted of a liquid–liquid extraction with methanol/tert-butyl methyl ether (1:1, v/v). The considered compounds were detected by a single-stage Exactive^TM mass spectrometer, operating at a mass resolution of 100,000 full width at half maximum. The validation study demonstrated excellent performance in terms of linearity (R ²?>?0.99), repeatability (CV?≤?10.6 %), within-laboratory reproducibility (CV?≤?12.2 %), and mean corrected recovery (ranging from 85 to 106 %). Additionally, a comparative evaluation towards well-established detection techniques, i.e., tandem mass spectrometry (MS/MS) and ultraviolet-visible spectroscopy (UV–VIS) photodiode array, indicated superior performance of high-resolution Orbitrap-MS with regard to specificity/selectivity and sensitivity (with limits of detection ranging from 1.0 to 3.8 pg μL^?1). As a result, it may be concluded that high-resolution Orbitrap-MS is a suited alternative for UV–VIS or MS/MS in analyzing carotenoids and may offer significant value in carotenoid research because of the metabolomic screening possibilities.

Probing local environments in paramagnetic europium-substituted Keggin solids by 31P magic angle spinning NMR spectroscopy

Paramagnetic Eu-substituted Keggin oxopolytungstates crystallize in different forms, determined by the nature of the counterions. The crystal packing is in turn responsible for the variations in the geometry of paramagnetic Eu sites with respect to the anion core. We probed the paramagnetic environments in a series of Eu-substituted Keggin solids, by 31P magic angle spinning NMR spectroscopy. 31P spinning sideband envelopes are dominated by the electron-nuclear dipolar interaction. For the compounds under investigation, both the magnitude and the asymmetry parameter of the electron-nuclear dipolar coupling tensor are sensitive to the mutual arrangements of paramagnetic Eu sites in the crystal lattice. and also report on the stoichiometry of the anion. The electron-nuclear dipolar coupling tensors were calculated from the crystallographic coordinates and the experimentally determined effective magnetic moments, assuming a point dipole approximation. The computed tensors are in very good agreement with the experimental spectra. Furthermore, the P-Eu distance estimates, accurate to within 0.06-0.12 A, can be obtained directly from the magnitude of the electron-nuclear dipolar coupling. This work demonstrates that 31P MAS NMR spectroscopy is a useful probe for investigating local environments in paramagnetic Keggin solids.     

Multiple cation binding by bola-amphiphilic crown ethers: Assessment by fast atom bombardment mass spectrometry

Abstract

Bola-amphiphiles having two aza-18-crown-6 ether rings attached by a covalent spacer (O—O) and an analogous tris (macrocycle) (O—O—O) have been prepared and their cation complexation behavior has been assessed by fast atom bombardment and collisionally activated dissociation tandem mass spectrometry; the key finding is that two cations may simultaneously be complexed by a two- or three-crown system and that this complexation may also involve an anion.     

Influence of the Lanthanide Ion and Solution Conditions on Formation of Lanthanide Wells–Dawson Polyoxotungstates

Lanthanide complexes of polyoxometalates, including the α₂-P₂W₁₇O₆₁ ¹⁰⁻ ligand, have been pioneered by Michael T. Pope, to whom this paper is dedicated. Examination of the solid-state and solution behavior of lanthanide complexes of the α₂-P₂W₁₇O₆₁ ¹⁰⁻ ligand are reported here to identify trends that will facilitate rational synthesis of hybrid organic lanthanide polyoxometalate complexes. Therefore, combining our data with that obtained by Pope and others a number of trends come into view. It is clear that there are two structural types for the 1:1 or 2:2 [Ln(H₂O)_X(α₂-P₂W₁₇O₆₁)]₂ ¹⁴⁻ species. The early lanthanides show a “cap to cap” structure that allows the Ln ion to be 9 coordinate and accommodates the longer bond lengths. The mid-late lanthanides show a “cap to belt” structure that allows the lanthanides to be 8 coordinate; this structural type is appropriate for the shorter bond lengths of the later lanthanides. The 1:1⇌1:2 equilibrium, that was observed by Pope for the Ce(III) analog is prevalent for the early- mid lanthanides. This equilibrium is slightly dependent on pH; however, cations have a major influence on this equilibrium. Larger, poorly hydrated cations appear to favor the 1:2 species for the early to mid lanthanides. Cations do not appear to influence the equilibrium for the later lanthanides; for all counterions, the 1:1 species was stable with no trace of the 1:2 species. Stability constants, K1 and K2, for the early to mid lanthanides were measured in this study by a competitive method and compared well with other published stability constant determinations. We suggest that the stability constants are not only dependent on the strength of interaction of the Ln with the α₂-P₂W₁₇O₆₁ ¹⁰⁻ ligand, but are also significantly influenced by the medium. The medium may bias the equilibria of the early-mid lanthanides and later lanthanides. The log K₁/log K₂ ratios are very close, suggesting that it is difficult to separate the 1:1 and 1:2 Ln: α₂-P₂W₁₇O₆₁ ¹⁰⁻ species.Electronic Supplementary Material Supplementary material for this article is available at and is accessible for authorized users.This paper is dedicated to Professor Michael T. Pope in honor of his substantial and sustained contributions to polyoxometalate chemistry and his inspiration to scientists working in the field.     

Europium(III) reduction and speciation within a Wells-Dawson heteropolytungstate

The redox speciation of Eu(III) in the 1:1 stoichiometric complex with the alpha-1 isomer of the Wells-Dawson anion, [alpha-1-P 2W 17O 61] (10-), was studied by electrochemical techniques (cyclic voltammetry and bulk electrolysis), in situ XAFS (X-ray absorption fine structure) spectroelectrochemistry, NMR spectroscopy ( (31)P), and optical luminescence. Solutions of K 7[(H 2O) 4Eu(alpha-1-P 2W 17O 61)] in a 0.2 M Li 2SO 4 aqueous electrolyte (pH 3.0) show a pronounced concentration dependence to the voltammetric response. The fully oxidized anion and its reduced forms were probed by Eu L 3-edge XANES (X-ray absorption near edge structure) measurements in simultaneous combination with controlled potential electrolysis, demonstrating that Eu(III) in the original complex is reduced to Eu(II) in conjunction with the reduction of polyoxometalate (POM) ligand. After exhaustive reduction, the heteropoly blue species with Eu(II) is unstable with respect to cluster isomerization, fragmentation, and recombination to form three other Eu-POMs as well as the parent Wells-Dawson anion, alpha-[P 2W 18O 62] (6-). EXAFS data obtained for the reduced, metastable Eu(II)-POM before the onset of Eu(II) autoxidation provides an average Eu-O bond length of 2.55(4) A, which is 0.17 A longer than that for the oxidized anion, and consistent with the 0.184 A difference between the Eu(II) and Eu(III) ionic radii. The reduction of Eu(III) is unusual among POM complexes with Lindqvist and alpha-2 isomers of Wells-Dawson anions, that is, [Eu(W 5O 18) 2] (9-) and [Eu(alpha-2-As 2W 17O 61) 2] (17-), but not to the Preyssler complex anion, [EuP 5W 30O 110] (12-), and fundamental studies of materials based on coupling Eu and POM redox properties are still needed to address new avenues of research in europium hydrometallurgy, separations, and catalysis sciences.     

The synthesis of azadirachtin: a potent insect antifeedant

We describe in full the first synthesis of the potent insect antifeedant azadirachtin through a highly convergent approach. An O-alkylation reaction is used to unite decalin ketone and propargylic mesylate fragments, after which a Claisen rearrangement constructs the central C8-C14 bond in a stereoselective fashion. The allene which results from this sequence then enables a second critical carbon-carbon bond forming event whereby the [3.2.1] bicyclic system, present in the natural product, is generated via a 5-exo-radical cyclisation process. Finally, using knowledge gained through our early studies into the reactivity of the natural product, a series of carefully designed steps completes the synthesis of this challenging molecule.     

Using a two-step hydride transfer to achieve 1,4-reduction in the catalytic hydrogenation of an acyl pyridinium cation

The stoichiometric reduction of N-carbophenoxypyridinium tetraphenylborate (6) by CpRu(P-P)H (Cp = eta(5)-cyclopentadienyl; P-P = dppe, 1,2-bis(diphenylphosphino)ethane, or dppf, 1,1'-bis(diphenylphosphino)ferrocene), and Cp*Ru(P-P)H (Cp* = eta(5)-pentamethylcyclopentadienyl; P-P = dppe) gives mixtures of 1,2- and 1,4-dihydropyridines. The stoichiometric reduction of 6 by Cp*Ru(dppf)H (5) gives only the 1,4-dihydropyridine, and 5 catalyzes the exclusive formation of the 1,4-dihydropyridine from 6, H(2), and 2,2,6,6-tetramethylpiperidine. In the stoichiometric reductions, the ratio of 1,4 to 1,2 product increases as the Ru hydrides become better one-electron reductants, suggesting that the 1,4 product arises from a two-step (e(-)/H(*)) hydride transfer. Calculations at the UB3LYP/6-311++G(3df,3pd)//UB3LYP/6-31G* level support this hypothesis, indicating that the spin density in the N-carbophenoxypyridinium radical (13) resides primarily at C4, while the positive charge in 6 resides primarily at C2 and C6. The isomeric dihydropyridines thus result from the operation of different mechanisms: the 1,2 product from a single-step H(-) transfer and the 1,4 product from a two-step (e(-)/H(*)) transfer.