Characterization of halogen species in seaweeds from the Antarctic using a multi-technique approach.

BACKGROUND: Despite the recognized importance, the determination of halogens in Antarctic seaweeds remains understudied. Limited research exists due to challenges associated with sample preparation, and reliable analytical techniques for this type of analysis. Therefore, further investigations are necessary to bridge this knowledge gap and gain a comprehensive understanding of halogen metabolism in Antarctic seaweeds. METHODS: In this study, seaweeds from the coast of the Antarctic continent were characterized concerning the total content of halogens and their species. For this purpose, different sample preparation methods, based on extraction and combustion, combining highly selective and sensitive chromatographic and spectrometric multi-technique approaches were used. RESULTS: By using optimized methods, it was possible to determine total halogens content, the distribution of bromine and iodine in different classes of species (lipids, water-soluble, proteins, carbohydrates, and residue), as well as the identification of iodinated amino acids (MIT and DIT) in ten brown and red seaweeds. Bromate and iodate were not detected in the samples, which presented only bromide and iodide species in their composition. Additionally, unknown bromine and iodine species were observed in different extracts evaluated. Furthermore, 25 halogenated polyphenols were identified in seaweeds, of which only four were already reported in the literature. CONCLUSION: The results obtained in this study comprise unprecedented data in the literature on species of halogens present in seaweeds from the Antarctic environment.

Study of metalation of thioredoxin by gold(I) therapeutic compounds using combined liquid chromatography/capillary electrophoresis with inductively coupled plasma/electrospray MS/MS detection.

The reactivity of thioredoxin (Trx1) with the Au(I) drug auranofin (AF) and two therapeutic N-heterocyclic carbene (NHC)2-Au(I) complexes (bis [1-methyl-3-acridineimidazolin-2-ylidene]gold(I) tetrafluoroborate (Au3BC) and [1,3-diethyl-4,5-bis(4methoxyphenyl)imidazol-2-ylidene]gold(I) (Au4BC)) was investigated. Direct infusion (DI) electrospray ionization (ESI) mass spectrometry (MS) allowed information on the structure, stoichiometry, and kinetics of formation of Trx-Au adducts. The fragmentation of the formed adducts in the gas phase gave insights into the exact Au binding site within the protein, demonstrating the preference for Trx1 Cys32 or Cys35 of AF or the (NHC)2-Au(I) complex Au3BC, respectively. Reversed-phase HPLC suffered from the difficulty of elution of gold compounds, did not preserve the formed metal-protein adducts, and favored the loss of ligands (phosphine or NHC) from Au(I). These limitations were eliminated by capillary electrophoresis (CE) which enabled the separation of the gold compounds, Trx1, and the formed adducts. The ICP-MS/MS detection allowed the simultaneous quantitative monitoring of the gold and sulfur isotopes and the determination of the metallation extent of the protein. The hyphenation of the mentioned techniques was used for the analysis of Trx1-Au adducts for the first time.

Unveiling the Pool of Metallophores in Native Environments and Correlation with Their Potential Producers.

For many organisms, metallophores are essential biogenic ligands that ensure metal scavenging and acquisition from their environment. Their identification is challenging in highly organic matter rich environments like peatlands due to low solubilization and metal scarcity and high matrix complexity. In contrast to common approaches based on sample modification by spiking of metal isotope tags, we have developed a two-dimensional (2D) Solid-phase extraction-Liquid chromatography-mass spectrometry (SPE-LC-MS) approach for the highly sensitive (LOD 40 fmol per g of soil), high-resolution direct detection and identification of metallophores in both their noncomplexed (apo) and metal-complexed forms in native environments. The characterization of peat collected in the Bernadouze (France) peatland resulted in the identification of 53 metallophores by a database mass-based search, 36 among which are bacterial. Furthermore, the detection of the characteristic (natural) metal isotope patterns in MS resulted in the detection of both Fe and Cu potential complexes. A taxonomic-based inference method was implemented based on literature and public database (antiSMASH database version 3.0) searches, enabling to associate over 40% of the identified bacterial metallophores with potential producers. In some cases, low completeness with the MIBiG reference BCG might be indicative of alternative producers in the ecosystem. Thus, coupling of metallophore detection and producers' inference could pave a new way to investigate poorly documented environment searching for new metallophores and their producers yet unknown.

Biodegradation of saturate fraction of crude oil and production of signature carboxylic acids.

Bacteria degrading large portion of saturated hydrocarbons are important for crude oil bioremediation. This study investigates Novosphingobium sp. S1, Gordonia amicalis S2 and Gordonia terrae S5 capability of degrading wide range of saturated hydrocarbons from Congo Bilondo crude oil and discusses the degradation pathway. A parallel analytical approach combining GC-MS and LC-HRMS enabled characterization of saturated hydrocarbons and comprehensive determination of carboxylic acid metabolites produced during biodegradation, respectively. Results showed that the three strains could efficiently degrade the n-alkanes (C10-C28) as well as methyl-substituted alkanes (C11-C26). The series of mono-, hydroxy- and dicarboxylic acids identified in this study confirmed the active biodegradation of the saturate fraction and suggest their degradation was via the bi-terminal oxidation pathway. This is the first study linking these bacterial species to bi-terminal oxidation of the saturated hydrocarbons. The study highlights the potential application of the bacterial strains in the bioremediation of crude oil contaminated sites. Additionally, while carboxylic acids is indicated as a suitable and valuable metabolic biomarker, its application is considered feasible and cost effective for rapid monitoring and evaluation of hydrocarbon biodegradation.

Detection of secondary cyanobacterial metabolites using LC-HRMS in Lake Karaoun.

Harmful algal blooms events have been reported worldwide and during the last decades are occurred with increasing frequency and intensity due to the climate change and the high inputs of nutrients in freshwaters from anthropogenic activities. During blooms cyanobacteria release in water their toxic secondary metabolites, known as cyanotoxins, along with other bioactive metabolites. Due to the negative impacts of these compounds on aquatic ecosystems and public health, there is an urgent need to detect and identify known and unknown cyanobacterial metabolites in surface waters. In the frame of the present study, a method based on liquid chromatography - high resolution mass spectrometry (LC-HRMS) was developed to investigate the presence of cyanometabolites in bloom samples from Lake Karaoun, Lebanon. Data analysis was performed using Compound Discoverer software with related tools and databases in combination to the CyanoMetDB mass list for detection, identification and structural elucidation of the cyanobacterial metabolites. In the course of this study, 92 cyanometabolites were annotated including 51 cyanotoxins belonging to microcystins, 15 microginins, 10 aeruginosins, 6 cyclamides, 5 anabaenopeptins, a cyanopeptolin, the dipeptides radiosumin B and dehydroradiosumin, the planktoncyclin and a mycosporine-like amino acid. Out of them, 7 new cyanobacterial metabolites, the chlorinated MC-ClYR, [epoxyAdda5]MC-YR, MC-LI, aeruginosin 638, aeruginosin 588, microginin 755C and microginin 727 were discovered. Moreover, the presence of anthropogenic contaminants was recorded indicating the pollution of the lake and emphasizing the need for assessment of the co-occurrence of cyanotoxins, other cyanobacterial metabolites and other compounds hazardous to the environment. Overall, results prove the suitability of the proposed approach for the detection of cyanobacterial metabolites in environmental samples but also highlight the necessity of spectral libraries for these compounds, considering the absence of their reference standards.

Study of metal and organic contaminants transported by microplastics in the Lebanese coastal environment using ICP MS, GC-MS, and LC-MS.

An untargeted study of multiclass contaminants associated with microplastics (MPs) in the East Mediterranean was carried out. Samples were collected in 2020-2021 from the shoreline at 14 different locations, along the Lebanese coast. Attenuated Total Reflectance (ATR) FTIR spectroscopy showed the predominant presence of polyethylene and polypropylene among plastic debris. The non-polar and polar organic compounds sorbed on the MPs were identified and quantified by GC - TOF MS and LC - electrospray MS/MS, respectively. Deconvolution of accurate GC-MS scan data allowed the identification of >130 organic pollutants, 64 of which could be confirmed by matching with the authentic standards, among which a number of previously unreported in targeted GC-MS(MS) methods. In addition to highly toxic, legacy chlorinated pollutants, high levels (average values from 0.8 to 4.0 µg g-1) of some musks, UV filters and UV absorbers were detected. Untargeted LC-MS demonstrated the persistence of several pesticides (i.e., chlorpyrifos) and pharmaceuticals, such as phenacetin and minoxidil, which were quantified. In addition, a study of metals associated with microplastics using ICP-MS confirmed the high potential of microplastics to act as a vector of, among others, toxic metals, such as Cd, Pb, Bi or Hg.

Nontargeted Screening for Flavonoids in Salicornia Plant by Reversed-Phase Liquid Chromatography-Electrospray Orbitrap Data-Dependent MS2/MS3.

The Salicornia genus has great potential in agrifood industries because of its nutritional benefits related to its high content of antioxidant compounds, including flavonoids. A nontargeted method based on reversed-phase liquid chromatography-electrospray orbitrap data-dependent MS2/MS3 and the fragment ion search (FISh) strategy was developed to screen flavonoids in Salicornia plants. An extensive study of fragmentation of a set of flavonoid standards allowed for the definition of 15 characteristic fragment ions for flagging flavonoids in the plant matrix. The nontargeted analysis was applied to Salicornia europaea species and allowed for the annotation of 25 candidate flavonoids, including 14 that had not been reported previously. Structural prediction of two unreported flavonoids and their isomeric forms was based on an advanced data processing method using an in silico approach and in-house databases compiling flavonoid-specific chemical substitution. Finally, the method developed allowed for the optimization of extraction yields of flavonoids from the plant matrix.

Isotopologue pattern based data mining for selenium species from HILIC-ESI-Orbitrap-MS-derived spectra.

Automated and specific picking of selenium-containing molecular entities has not been an obvious option for software tools associated with electrospray high-resolution mass spectrometry (MS). In our study, a comprehensive pattern matching approach based on intra-isotopologue distance and isotopologue ratio data was critically evaluated in terms of reproducibility and selenium isotope selection on three samples, including selenized Torula yeast and the selenium hyperaccumulator plant Cardamine violifolia. Hydrophilic interaction liquid chromatography was applied to provide a one-step separation for water soluble metabolites to put an end to the need for either orthogonal setups or poor retention on reversed phase chromatography. Assistance from inductively coupled plasma-MS was taken only for chromatographic verification purposes, and the involvement of absolute mass defect (MD) data in selenometabolite-specific screening was assessed by multivariate statistical tools. High focus was placed on screening efficiency and on the validation of discovered selenized molecules to avoid reporting of artefacts. From the >1000 molecular entries detected, selenium-containing molecules were picked up with a recovery rate of >88% and a false positive rate of <10%. Isotop(ologu)e pairs of 78Se-80Se and 80Se-82Se proved to be the most performant in the detection. On the basis of accurate mass information and hypothetical deamination processes, elemental composition could be proposed for 72 species out of the 75 selenium species encountered without taking into account selenocompound databases. Absolute MD data were used to significantly differentiate a potentially sample-specific subgroup of false positive molecular entities from non-selenized and selenized entities.

Resolving Severe Elemental Isobaric Interferences with a Combined Atomic and Molecular Ionization Source-Orbitrap Mass Spectrometry Approach: The 87Sr and 87Rb Geochronology Pair.

Many fields of basic and applied sciences, including geochronology, astronomy, metabolism, etc., rely on the ability of mass spectrometry to obtain isotope ratio measurements having a high degree of certainty. The inability to resolve difficult isobaric interferences plagues certain measurements. A combined atomic and molecular (CAM) ionization source has been interfaced to a high-field Orbitrap mass spectrometer to alleviate severe atomic, isobaric interferences. This work examines the geochronologically significant 87Sr and 87Rb isotope pair. The mass difference between 87Sr and 87Rb is approximately 0.3 mDa, requiring a minimum resolving power (R = m/Δm) of ∼290,000, a value ∼30× higher than available with sector-field elemental mass spectrometers. Under ultrahigh-resolution conditions, Sr isotope ratio accuracy and precision were evaluated using NIST Sr SRM 987, yielding precision values of <0.1% relative standard deviation (RSD) for the major isotopes and a calculated LOD of 2 pg mL-1 (120 fg of Sr for a 60 µL injection). In addition to manipulating the signal transient length, the total number of ions in the electrostatic trap and the 87Sr/87Rb concentration ratio were found to influence resolution. Ultimately, the isotopes were baseline-resolved with a calculated mass resolution of >1.7M. At equal 87Sr and 87Rb intensities, 87Sr/86Sr was measured as 0.71294 (a relative error of only 0.37%) with a precision of 0.097% RSD, clearly reflecting the alleviation of the isobaric interference.

Untargeted Analysis for Mycosporines and Mycosporine-Like Amino Acids by Hydrophilic Interaction Liquid Chromatography (HILIC)-Electrospray Orbitrap MS2/MS3.

Mycosporines and mycosporine-like amino acids have been described as natural sunscreens and antioxidant compounds presenting a great potential for health and cosmetic applications. Herein, an untargeted screening approach for mycosporines and mycosporine-like amino acids (MAAs) was developed by the coupling of zwitterionic hydrophilic interaction liquid chromatography (HILIC) with multistage electrospray mass spectrometry MS2/MS3 using an Orbitrap analyzer and fragment ion search (FISh). This method was applied to study the mycosporine and MAA contents of five algae extracted using a 50% methanol solution and sonication. Candidate-MAAs were detected by mining eight characteristic fragment ions in their HILIC data-dependent MS2 mass spectrum. Their exact masses were measured with 3 ppm mass accuracy and their structures were elucidated on the basis of the MS3/MS4 mass spectra. The method developed was validated with a targeted analysis using an extract of Gymnogongrus devoniensis which confirmed the detection of 14 MAAs reported in the literature. In addition, 23 previously unreported MAAs were detected and the structures could be assigned for seven of them. The developed method was applied to the analysis of four algae: Gelidium sesquipedale, Halopithys incurva, Porphyra rosengurtii and Cystoseira tamariscifolia allowing the detection of MAAs, including some reported here for the first time.

Nickel Nanoparticles Induce the Synthesis of a Tumor-Related Polypeptide in Human Epidermal Keratinocytes.

Although nickel allergy and carcinogenicity are well known, their molecular mechanisms are still uncertain, thus demanding studies at the molecular level. The nickel carcinogenicity is known to be dependent on the chemical form of nickel, since only certain nickel compounds can enter the cell. This study investigates, for the first time, the cytotoxicity, cellular uptake, and molecular targets of nickel nanoparticles (NiNPs) in human skin cells in comparison with other chemical forms of nickel. The dose-response curve that was obtained for NiNPs in the cytotoxicity assays showed a linear behavior typical of genotoxic carcinogens. The exposure of keratinocytes to NiNPs leads to the release of Ni2+ ions and its accumulation in the cytosol. A 6 kDa nickel-binding molecule was found to be synthesized by cells exposed to NiNPs at a dose corresponding to medium mortality. This molecule was identified to be tumor-related p63-regulated gene 1 protein.

A chemical speciation insight into the palladium(ii) uptake and metabolism by Sinapis alba. Exposure to Pd induces the synthesis of a Pd-histidine complex.

Palladium is recognized as a technologically critical element (TCE) because of its massive use in automobile exhaust gas catalytic converters. The release of Pd into the environment in the form of nanoparticles of various size and chemical composition requires an understanding of their metabolism by leaving organisms. We provide here for the first time a chemical speciation insight into the identity of the ligands produced or used by a plant Sinapis alba L. exposed in hydropony to Pd nanoparticles and soluble Pd (nitrate). The analytical method developed was based on the concept of 2D HPLC with parallel inductively coupled plasma mass spectrometry (ICP MS) and electrospray MS detection. Size exclusion chromatography - ICP MS of the plant extracts showed no difference between the speciation of Pd after the exposure to nanoparticles and after that to Pd2+ which indicated the reactivity and dissolution of Pd nanoparticles. A comparative investigation of the Pd speciation in a control plant extract spiked with Pd2+ and of an extract of a plant having metabolized palladium indicated the response of the Sinapis alba by the formation of a Pd-histidine complex. The complex was identified via Orbitrap MS; the HPLC-MS chromatogram produced two peaks at m/z 415.0341 each corresponding to a Pd-His2 complex. An investigation by ion-mobility MS revealed a difference in their collision cross section indicating that the complexes present varied in terms of spatial conformation. A number of other Pd complexes with different ligands (including nicotianamine) circulating in the plant were detected but these ligands were already observed in a control plant and their concentrations were not affected by the exposure to Pd.

Ultra-High Resolution Elemental/Isotopic Mass Spectrometry (m/Δm > 1,000,000): Coupling of the Liquid Sampling-Atmospheric Pressure Glow Discharge with an Orbitrap Mass Spectrometer for Applications in Biological Chemistry and Environmental Analysis.

Many fundamental questions of astrophysics, biochemistry, and geology rely on the ability to accurately and precisely measure the mass and abundance of isotopes. Taken a step further, the capacity to perform such measurements on intact molecules provides insights into processes in diverse biological systems. Described here is the coupling of a combined atomic and molecular (CAM) ionization source, the liquid sampling-atmospheric pressure glow discharge (LS-APGD) microplasma, with a commercially available ThermoScientific Fusion Lumos mass spectrometer. Demonstrated for the first time is the ionization and isotopically resolved fingerprinting of a long-postulated, but never mass-spectrometrically observed, bi-metallic complex Hg:Se-cysteine. Such a complex has been implicated as having a role in observations of Hg detoxification by selenoproteins/amino acids. Demonstrated as well is the ability to mass spectrometrically-resolve the geochronologically important isobaric 87Sr and 87Rb species (Δm ~ 0.3 mDa, mass resolution m/Δm ≈ 1,700,000). The mass difference in this case reflects the beta-decay of the 87Rb to the stable Sr isotope. These two demonstrations highlight what may be a significant change in bioinorganic and atomic mass spectrometry, with impact expected across a broad spectrum of the physical, biological, and geological sciences. Graphical Abstract "".

Direct screening of food packaging materials for post-polymerization residues, degradation products and additives by liquid extraction surface analysis nanoelectrospray mass spectrometry (LESA-nESI-MS).

Materials in direct contact with food should be monitored for the presence of species able to migrate into food. A direct method based on liquid extraction surface analysis nanoelectrospray mass spectrometry (LESA-nanoESI-MS) was developed for the analysis of the migrating species from a polymer film. Different types of molecules: post-polymerization residues, degradation products (oligomers resulting from polymer recycling, products of polymer oxidative degradation) and anti-oxidant additives (vitamin E) were demonstrated to be detected and identified, and determined quantitatively if relevant calibration standards are available. The method was validated by a comparison a standard method based on with bulk extraction mass spectrometry. It offers considerable advantages over the latter in terms of drastically reduced analysis time and solvent consumption. Also, LESA-nanoESI-MS produced simpler spectra (limited to compounds able to migrate into food) than Direct Analysis in Real Time (DART).

Deproteinization assessment using isotopically enriched compounds to trace the coprecipitation of low-molecular-weight selenium species with proteins.

Studies have shown that information related to the presence of low-molecular-weight metabolites is frequently lost after deproteinization of complex matrices, such as blood and plasma, during sample preparation. Therefore, the effect of several deproteinization reagents on low-molecular-weight selenium species has been compared by species-specific isotope labeling. Two isotopically enriched selenium tracers were used to mimic models of small inorganic anionic (77Se-selenite) and organic zwitterionic (76Se-selenomethionine) species. The results presented here show that the use of a methanol-acetonitrile-acetone (1:1:1 v/v/v) mixture provided approximately two times less tracer loss from plasma samples in comparison with the classic procedure using acetonitrile, which may not be optimal as it leads to important losses of low-molecular-weight selenium species. In addition, the possible interactions between selenium tracers and proteins were investigated, revealing that both coprecipitation phenomena and association with proteins were potentially responsible for selenite tracer losses during protein precipitation in blood samples. However, coprecipitation phenomena were found to be fully responsible for losses of both tracers observed in plasma samples and of the selenomethionine tracer in blood samples. This successfully applied strategy is anticipated to be useful for more extensive future studies in selenometabolomics.

Influence of Dietary Selenium Species on Selenoamino Acid Levels in Rainbow Trout.

Two forms of selenium (Se) supplementation of fish feeds were compared in two different basal diets. A 12-week feeding trial was performed with rainbow trout fry using either a plant-based or a fish meal-based diet. Se yeast and selenite were used for Se supplementation. Total Se and Se speciation were determined in both diets and whole body of trout fry using inductively coupled plasma mass spectrometry (ICP MS) and high-performance liquid chromatography (HPLC). The two selenoamino acids, selenomethionine (SeMet) and selenocysteine (SeCys), were determined in whole body of fry after enzymatic digestion using protease type XIV with a prior derivatization step in the case of SeCys. The plant-based basal diet was found to have a much lower total Se than the fish meal-based basal diet with concentrations of 496 and 1222 µg(Se) kg(-1), respectively. Dietary Se yeast had a higher ability to raise whole body Se compared to selenite. SeMet concentration in the fry was increased only in the case of Se yeast supplementation, whereas SeCys levels were similar at the end of the feeding trial for both Se supplemented forms. The results show that the fate of dietary Se in fry is highly dependent on the form brought through supplementation and that a plant-based diet clearly benefits from Se supplementation.

Influence of the forms and levels of dietary selenium on antioxidant status and oxidative stress-related parameters in rainbow trout (Oncorhynchus mykiss) fry.

Se is an essential micronutrient required for normal growth, development and antioxidant defence. The objective of the present study was to assess the impact of dietary Se sources and levels on the antioxidant status of rainbow trout (Oncorhynchus mykiss) fry. First-feeding fry (initial body weight: 91 mg) were fed either a plant- or fishmeal-based diet containing 0·5 or 1·2 mg Se/kg diet supplemented or not with 0·3 mg Se/kg diet supplied as Se-enriched yeast or sodium selenite for 12 weeks at 17°C. Growth and survival of rainbow trout fry were not significantly affected by dietary Se sources and levels. Whole-body Se was raised by both Se sources and to a greater extent by Se-yeast. The reduced:oxidised glutathione ratio was raised by Se-yeast, whereas other lipid peroxidation markers were not affected by dietary Se. Whole-body Se-dependent glutathione peroxidase (GPX) activity was enhanced in fish fed Se-yeast compared to fish fed sodium selenite or non-supplemented diets. Activity and gene expression of this enzyme as well as gene expression of selenoprotein P (SelP) were reduced in fish fed the non-supplemented plant-based diet. Catalase, glutamate-cysteine ligase and nuclear factor-erythroid 2-related factor 2 (Nrf2) gene expressions were reduced by Se-yeast. These results suggest the necessity to supplement plant-based diets with Se for rainbow trout fry, and highlight the superiority of organic form of Se to fulfil the dietary Se requirement and sustain the antioxidant status of fish. GPX and SelP expression proved to be good markers of Se status in fish.