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Inductively coupled plasma with time-of-flight mass spectrometry (ICP-ToF-MS) is currently setting new benchmarks for the analysis of single particles (SP) and elemental mapping using laser ablation (LA). The rapid collection of full elemental mass spectra promotes non-target approaches, fast imaging as well as inquiries of particle composition. However, one shortcoming often associated with ICP-ToF-MS is a lack of detection power due to lower duty cycles relative to sequentially operating mass analysers targeting only a limited number of elements. The sensitivity of ICP-ToF-MS can be increased using two strategies, which are detailed in this study. First, instead of analysing full mass spectra, elements in the low and high mass ranges were excluded from analysis using a Bradbury–Nielsen gate. The resulting restricted mass range was acquired up to 5 times faster increasing duty cycles and sensitivity accordingly. Second, isotopes of polyisotopic elements recorded simultaneously were accumulated to increase signal to noise ratios. In a proof of concept, we applied SP ICP-ToF-MS for the first time to characterise upconversion nanoparticles (UCNPs) that contained Gd and Yb. Both signal amplification strategies were combined and the consequences for detection limits and signal to noise ratios were considered and compared to a standard method. Sensitivities were increased up to factor 27 when accumulating all Gd and Yb isotopes at 177 kHz, and size detection limits decreased by a factor of approximately 3. Improved figures of merit promoted more accurate investigations of UCNPs, which were characterised regarding size distributions and composition. As a second application, we demonstrated the utility of the described strategies in LA-ICP-ToF-MS. Mo and Se were targeted as relatively rare elements in rat brain tissue. Increased acquisition frequencies of 180 kHz and isotope accumulation resulted in drastically improved signal to noise ratios and enabled the mapping of both elements while still considering relevant neuroanatomical elements such as Fe and Zn. [ABSTRACT FROM AUTHOR]

Many oil and gas developments will soon be decommissioned and, knowledge on the accumulation of mercury (Hg), throughout offshore infrastructure is limited. Any release of Hg could have a detrimental impact on marine ecosystems. To bridge this knowledge gap, a fractionation approach was taken on steel samples exposed to Hg 0 and H 2 S, separating Hg compounds removed from the surface into polar, non-polar and insoluble fractions. Hg 0 reacted on corroded surfaces to form several compounds, over 50 % of which were removed by seawater. This suggests that pipelines on the seabed could release a dramatic amount of Hg into the sea if they are left in place. Furthermore, a Cu-Hg amalgam, was identified to be a dominant species, by a combination of XFM, XANES and LA-ICP-TOFMS. Seawater-soluble and amalgam-bound Hg were present regardless of co-exposure to H 2 S. When H 2 S was present Hg nanoparticles accounted for up to 1 % of the total Hg on the steel. This investigation has shown that the Hg speciation on the surfaces of pipelines is complex and future decommissioning strategies should consider a range of Hg species beyond only Hg 0 and metacinnabar (β-HgS), all of which could interact with biota and impact Hg biomagnification through the marine the food web., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Breast cancer (BC) is the leading cause of cancer death among women worldwide, making the discovery and quantification of new biomarkers essential for improving diagnostic and preventive strategies to limit dissemination and improve prognosis. Essential trace metals such as Fe, Cu, and Zn may play critical roles in the pathophysiology of both benign and malignant breast tumors. However, due to the high metabolic activity and reduced element selectivity of cancer cells, also non-essential elements may be taken up and may even be implicated with disease progression. This study investigates the spatial distribution and concentrations of both essential and non-essential elements in breast tissues, assessing their potential for diagnostic applications. Laser ablation (LA)-inductively coupled plasma-mass spectrometry (ICP-MS) with a time-of-flight (ToF) mass analyzer (LA-ICP-ToF-MS) was used to inquire the distribution of almost all elements across the periodic table and their abundance in metastatic (n = 11), non-metastatic (n = 7), and healthy (n = 4) breast tissues. Quantification was achieved using gelatine-based standards for external calibration to quantitatively map various elements. Overall, the Fe, Cu, Zn, Sr, and Ba levels were significantly increased in tumor samples with Sr and Ba showing strong correlation, likely due to their similar chemistry. Comparison of calibrated LA-ICP-ToF-MS data with a histologic staining demonstrated the possibility to clearly differentiate between various tissue types and structures in breast tissues such as tumor niche and stroma. The levels of the studied elements were significantly higher in the tumor niche areas compared to the stroma, and for Fe, a significant accumulation was observed in the tumor niche areas from the metastatic patient group relative to the levels found in the same areas of the non-metastatic group., Competing Interests: Declarations Ethical approval This study was conducted in accordance with national regulations and received approval from the Ethics and Investigation Committee of the Hospital de Jove Foundation (PI02/2018). Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

The unique properties of per- and polyfluoroalkyl substances (PFAS) have led to their extensive use in consumer products, including ski wax. Based on the risks associated with PFAS, and to align with PFAS regulations, the international ski federation (FIS) implemented a ban on products containing "C 8 fluorocarbons/perfluorooctanoate (PFOA)" at all FIS events from the 2021/2022 season, leading manufactures to shift their formulations towards short-chain PFAS chemistries. To date, most studies characterising PFAS in ski waxes have measured a suite of individual substances using targeted analytical approaches. However, the fraction of total fluorine (TF) in the wax accounted for by these substances remains unclear. In this study, we sought to address this question by applying a multi-platform, fluorine mass balance approach to a total of 10 commercially available ski wax products. Analysis of TF by combustion ion chromatography (CIC) revealed concentrations of 1040-51700 μg F g -1 for the different fluorinated waxes. In comparison, extractable organic fluorine (EOF) determined in methanol extracts by CIC (and later confirmed by inductively-coupled plasma-mass spectrometry and 19 F- nuclear magnetic resonance spectroscopy) ranged from 92 to 3160 μg g -1 , accounting for only 3-8.8 % of total fluorine (TF). Further characterisation of extracts by cyclic ion mobility-mass spectrometry (IMS) revealed 15 individual PFAS with perfluoroalkyl carboxylic acid concentrations up to 33 μg F g -1 , and 3 products exceeding the regulatory limit for PFOA (0.025 μg g -1 ) by a factor of up to 100. The sum of all PFAS accounted for only 0.01-1.0 % of EOF, implying a high percentage of unidentified PFAS, thus, pyrolysis gas chromatography-mass spectrometry was used to provide evidence of the nature of the non-extractable fluorine present in the ski wax products., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Nanoparticles are produced at accelerating rates, are increasingly integrated into scientific and industrial applications, and are widely discharged into the environment. Analytical techniques are required to characterize parameters such as particle number concentrations, mass and size distributions, molecular and elemental compositions, and particle stability. This is not only relevant to investigate their utility for various industrial or medical applications and for controlling the manufacturing processes but also to assess toxicity and environmental fate. Different analytical strategies aim to characterize certain facets of particles but are difficult to combine to retrieve relevant parameters coherently and to provide a more comprehensive picture. In this work, we demonstrate the first online hyphenation of optofluidic force induction (OF2i) with Raman spectroscopy and inductively coupled plasma-time-of-flight-mass spectrometry (ICP-TOFMS) to harness their complementary technology-specific advantages and to promote comprehensive particle characterizations. We optically trapped individual particles on a weakly focused vortex laser beam by aligning a microfluidic flow antiparallelly to the laser propagation direction. The position of particles in this optical trap depended on the hydrodynamic diameter and therefore enabled size calibration as well as matrix elimination. Additionally, laser light scattered on particles was analyzed in a single particle (SP) Raman spectroscopy setup for the identification of particulate species and phases. Finally, particles were characterized regarding elemental composition and their distributions in mass and size using SP ICP-TOFMS. In a proof of concept, we analyzed polystyrene-based microplastic and TiO 2 nanoparticles and demonstrated the opportunities provided through the coupling of OF2i with SP Raman and SP ICP-TOFMS.

Innate immune systems alter the concentrations of trace elements in host niches in response to invading pathogens during infection. This work reports the interplay between d-block metal ions and their associated biomolecules using hyphenated elemental techniques to spatially quantify both elemental distributions and the abundance of specific transport proteins. Here, lung tissues were collected for analyses from naïve and Streptococcus pneumoniae-infected mice fed on a zinc-restricted or zinc-supplemented diet. Spatiotemporal distributions of manganese ( 55 Mn), iron ( 56 Fe), copper ( 63 Cu), and zinc ( 66 Zn) were determined by quantitative laser ablation-inductively coupled plasma-mass spectrometry. The murine transport proteins ZIP8 and ZIP14, which are associated with zinc transport, were also imaged by incorporation of immunohistochemistry techniques into the analytical workflow. Collectively, this work demonstrates the potential of a single instrumental platform suitable for multiplex analyses of tissues and labelled antibodies to investigate complex elemental interactions at the host-pathogen interface. Further, these methods have the potential for broad application to investigations of biological pathways where concomitant measurement of elements and biomolecules is crucial to understand the basis of disease and aid in development of new therapeutic approaches., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Wild boars have been reported as bioindicators for per- and polyfluoroalkyl substances (PFAS) in a variety of studies. However, data about PFAS levels in wild boars from sites with limited industrial and general human activity is scarce. In this study, wild boar (Sus scrofa) organs from the Bohemian Forest National Park (Czech Republic) were used as bioindicators for PFAS pollution. In this work, 29 livers and 24 kidneys from 30 wild boars (0.5-5 years) were investigated using a fluorine mass balance approach. For this, the samples were measured using high performance liquid chromatography with electrospray ionisation tandem mass spectrometry (HPLC-ESI-MS/MS), targeting 30 PFAS, including legacy and replacement PFAS, direct total oxidisable precursor assay (dTOPA) and combustion ion chromatography (CIC). Perfluorocarboxylic acids (PFCAs) from C 7 to C 14 and perfluorooctanesulfonic acid (PFOS) were detected in >50 % of samples. In the livers, PFCAs dominated the profile with median concentrations of 230 μg/kg for perfluorononanoic acid (PFNA) and 75 μg/kg perfluorooctanoic acid (PFOA). PFOA and PFNA concentrations in the livers were one order of magnitude higher than in livers from wild boars caught in rural NE Germany considered as background concentration. PFOS in liver contributed only 30 % to the Σc(PFAS Target ) with a median concentration of 170 μg/kg. Kidneys and livers contain an average of 2460 μg F/kg and 6800 μg F/kg extractable organic fluorine (EOF) respectively. Σc(PFAS Target ) add up to a maximum of 10 % of the extractable organic fluorine. After oxidisation of the samples, PFOA, PFNA and Σc(PFAS dTOPA ) increased in livers, but could not explain the EOF. The elevated concentration of PFOA and PFNA may indicate differences in biomagnification for different habitats or an unidentified PFAS source in proximity to the national park., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Multiple sclerosis (MS) is a prevalent immune-mediated inflammatory disease of the central nervous system inducing a widespread degradation of myelin and resulting in neurological deficits. Recent advances in molecular and atomic imaging provide the means to probe the microenvironment in affected brain tissues at an unprecedented level of detail and may provide new insights. This study showcases state-of-the-art spectroscopic and mass spectrometric techniques to compare distributions of molecular and atomic entities in MS lesions and surrounding brain tissues. MS brains underwent post-mortem magnetic resonance imaging (MRI) to locate and subsequently dissect MS lesions and surrounding white matter. Digests of lesions and unaffected white matter were analysed via ICP-MS/MS revealing significant differences in concentrations of Li, Mg, P, K, Mn, V, Rb, Ag, Gd and Bi. Micro x-ray fluorescence spectroscopy (μXRF) and laser ablation - inductively coupled plasma - time of flight - mass spectrometry (LA-ICP-ToF-MS) were used as micro-analytical imaging techniques to study distributions of both endogenous and xenobiotic elements. The essential trace elements Fe, Cu and Zn were subsequently calibrated using in-house manufactured gelatine standards. Lipid distributions were studied using IR-micro spectroscopy and matrix assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI). MALDI-MSI was complemented with high-resolution tandem mass spectrometry and trapped ion mobility spectroscopy for the annotation of specified phospho- and sphingolipids, revealing specific lipid species decreased in MS lesions compared to surrounding white matter. This explorative study demonstrated that modern molecular and atomic mapping techniques provide high-resolution imaging for relevant bio-indicative entities which may complement our current understanding of the underlying pathophysiological processes., Competing Interests: Declaration of competing interest Marten Seeba works for Bruker Corporation., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Formation of new C(sp 3 )-C(sp 3 ) bonds is a powerful synthetic tool to increase molecular diversity, which is highly sought after in medicinal chemistry. Traditional generation of carbon nucleophiles and more modern cross-electrophile-coupling methods typically lack sufficient selectivity when cross-coupling of analogous C(sp 3 )-containing reactants is attempted. Herein, we present a nickel-catalyzed, electrochemically driven method for the coupling of alkyl bromides with alkyl tosylates. Selective cross-coupling transformations were achieved even between C(sp 3 )-secondary bromides and tosylates. Key to achieve high selectivity was the combination of the tosylates with sodium bromide as the supporting electrolyte, gradually generating small amounts of the more reactive bromide by substitution and ensuring that one of the reaction partners in the nickel-catalyzed electroreductive process is maintained in excess during a large part of the process. The method has been demonstrated for a wide range of substrates (>30 compounds) in moderate to good yields. Further expanding the scope of electroorganic synthesis to C(sp 3 )-C(sp 3 ) cross-coupling reactions is anticipated to facilitate the switch to green organic synthesis and encourage future innovative electrochemical transformations.

This work describes a novel automated and rapid method for bottom-up proteomics combining protein isolation with a micro-immobilised enzyme reactor (IMER). Crosslinking chemistry based on 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling was exploited to immobilise trypsin and antibodies onto customisable silica particles coated with carboxymethylated dextran (CMD). This novel silica-CMD solid-phase extraction material was characterised using Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), conductometric titrations and enzymatic colorimetric assays. Micro-solid-phase extraction (μSPE) cartridges equipped with the modified CMD material were employed and integrated into an automated and repeatable workflow using a sample preparation workstation to achieve rapid and repeatable protein isolation and pre-concentration, followed by tryptic digestion producing peptide fragments that were identified by liquid chromatography mass spectrometry (LC-MS)., (© 2022. The Author(s).)

Metal ions are required by all organisms for the chemical processes that support life. However, in excess they can also exert toxicity within biological systems. During infection, bacterial pathogens such as Streptococcus pneumoniae are exposed to host-imposed metal intoxication, where the toxic properties of metals, such as copper, are exploited to aid in microbial clearance. However, previous studies investigating the antimicrobial efficacy of copper in vivo have reported variable findings. Here, we use a highly copper-sensitive strain of S. pneumoniae, lacking both copper efflux and intracellular copper buffering by glutathione, to investigate how copper stress is managed and where it is encountered during infection. We show that this strain exhibits highly dysregulated copper homeostasis, leading to the attenuation of growth and hyperaccumulation of copper in vitro. In a murine infection model, whole-tissue copper quantitation and elemental bioimaging of the murine lung revealed that infection with S. pneumoniae resulted in increased copper abundance in specific tissues, with the formation of spatially discrete copper hot spots throughout the lung. While the increased copper was able to reduce the viability of the highly copper-sensitive strain in a pneumonia model, copper levels in professional phagocytes and in a bacteremic model were insufficient to prosecute bacterial clearance. Collectively, this study reveals that host copper is redistributed to sites of infection and can impact bacterial viability in a hypersusceptible strain. However, in wild-type S. pneumoniae, the concerted actions of the copper homeostatic mechanisms are sufficient to facilitate continued viability and virulence of the pathogen. IMPORTANCE Streptococcus pneumoniae (the pneumococcus) is one of the world's foremost bacterial pathogens. Treatment of both localized and systemic pneumococcal infection is becoming complicated by increasing rates of multidrug resistance globally. Copper is a potent antimicrobial agent used by the mammalian immune system in the defense against bacterial pathogens. However, unlike other bacterial species, this copper stress is unable to prosecute pneumococcal clearance. This study determines how the mammalian host inflicts copper stress on S. pneumoniae and the bacterial copper tolerance mechanisms that contribute to maintenance of viability and virulence in vitro and in vivo . This work has provided insight into the chemical biology of the host-pneumococcal interaction and identified a potential avenue for novel antimicrobial development.

Inductively coupled-plasma mass spectrometry (ICP-MS) has transformed our knowledge on the role of trace and major elements in biology and has emerged as the most versatile technique in elemental mass spectrometry. The scope of ICP-MS has dramatically changed since its inception, and nowadays, it is a mature platform technology that is compatible with chromatographic and laser ablation (LA) systems. Over the last decades, it kept pace with various technological advances and was inspired by interdisciplinary approaches which endorsed new areas of applications. While the first part of this review was dedicated to fundamentals in ICP-MS, its hyphenated techniques and the application in biomonitoring, isotope ratio analysis, elemental speciation analysis, and elemental bioimaging, this second part will introduce relatively current directions in ICP-MS and their potential to provide novel perspectives in the medical sciences. In this context, current directions for the characterisation of novel nanomaterials which are considered for biomedical applications like drug delivery and imaging platforms will be discussed while considering different facets of ICP-MS including single event analysis and dedicated hyphenated techniques. Subsequently, immunochemistry techniques will be reviewed in their capability to expand the scope of ICP-MS enabling analysis of a large range of biomolecules alongside elements. These methods inspired mass cytometry and imaging mass cytometry and have the potential to transform diagnostics and treatment by offering new paradigms for personalised medicine. Finally, the interlacing of immunochemistry methods, single event analysis, and functional nanomaterials has opened new horizons to design novel bioassays which promise potential as assets for clinical applications and larger screening programs and will be discussed in their capabilities to detect low-level proteins and nucleic acids., (© 2022. The Author(s).)

Since its inception in the early 80s, inductively coupled plasma-mass spectrometry has developed to the method of choice for the analysis of elements in complex biological systems. High sensitivity paired with isotopic selectivity and a vast dynamic range endorsed ICP-MS for the inquiry of metals in the context of biomedical questions. In a stand-alone configuration, it has optimal qualities for the biomonitoring of major, trace and toxicologically relevant elements and may further be employed for the characterisation of disrupted metabolic pathways in the context of diverse pathologies. The on-line coupling to laser ablation (LA) and chromatography expanded the scope and application range of ICP-MS and set benchmarks for accurate and quantitative speciation analysis and element bioimaging. Furthermore, isotopic analysis provided new avenues to reveal an altered metabolism, for the application of tracers and for calibration approaches. In the last two decades, the scope of ICP-MS was further expanded and inspired by the introduction of new instrumentation and methodologies including novel and improved hardware as well as immunochemical methods. These additions caused a paradigm shift for the biomedical application of ICP-MS and its impact in the medical sciences and enabled the analysis of individual cells, their microenvironment, nanomaterials considered for medical applications, analysis of biomolecules and the design of novel bioassays. These new facets are gradually recognised in the medical communities and several clinical trials are underway. Altogether, ICP-MS emerged as an extremely versatile technique with a vast potential to provide novel insights and complementary perspectives and to push the limits in the medical disciplines. This review will introduce the different facets of ICP-MS and will be divided into two parts. The first part will cover instrumental basics, technological advances, and fundamental considerations as well as traditional and current applications of ICP-MS and its hyphenated techniques in the context of biomonitoring, bioimaging and elemental speciation. The second part will build on this fundament and describe more recent directions with an emphasis on nanomedicine, immunochemistry, mass cytometry and novel bioassays., (© 2022. The Author(s).)

Breast cancer is the leading cause of cancer death in woman and tremendous efforts are undertaken to limit its dissemination and to provide effective treatment. Various histopathological parameters are routinely assessed in breast cancer biopsies to provide valuable diagnostic and prognostic information. MMP-11 and CD45 are tumor-associated antigens and potentially valuable biomarkers for grading aggressiveness and metastatic probability. This paper presents methods for quantitative and multiplexed imaging of MMP-11 and CD45 in breast cancer tissues and investigates their potential for improved cancer characterization and patient stratification. An immunohistochemistry-assisted laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) method was successfully developed and optimized using lanthanide-tagged monoclonal antibodies as proxies to determine spatial distributions and concentrations of the two breast cancer biomarkers. The labeling degree of antibodies was determined via size exclusion-ICP-tandem mass spectrometry (SEC-ICP-MS/MS) employing online calibration via post-column isotope dilution analysis (IDA). The calibration of spatial distributions of labeled lanthanides in tissues was performed by ablating mold-prepared gelatin standards spiked with element standards. Knowledge of labeling degrees enabled the translation of lanthanide concentrations into biomarkers concentrations. The k-means clustering was used to select tissue areas for statistical analysis and mean concentrations were compared for sets of metastatic, non-metastatic and healthy samples. MMP-11 was expressed in stroma surrounding tumor areas, while CD45 was predominantly found inside tumor areas with high cell density. There was no significant correlation between CD45 and metastasis (P = 0.70); however, MMP-11 was significantly up-regulated (202%) in metastatic samples compared to non-metastatic (P = 0.0077) and healthy tissues (P = 0.0087)., (© The Author(s) 2022. Published by Oxford University Press.)

The analysis of natural and anthropogenic nanomaterials (NMs) in the environment is challenging and requires methods capable to identify and characterise structures on the nanoscale regarding particle number concentrations (PNCs), elemental composition, size, and mass distributions. In this study, we employed single particle inductively coupled plasma-mass spectrometry (SP ICP-MS) to investigate the occurrence of NMs in the Melbourne area (Australia) across 63 locations. Poisson statistics were used to discriminate between signals from nanoparticulate matter and ionic background. TiO 2 -based NMs were frequently detected and corresponding NM signals were calibated with an automated data processing platform. Additionally, a method utilising a larger mass bandpass was developed to screen for particulate high-mass elements. This procedure identified Pb-based NMs in various samples. The effects of different environmental matrices consisting of fresh, brackish, or seawater were mitigated with an aerosol dilution method reducing the introduction of salt into the plasma and avoiding signal drift. Signals from TiO 2 - and Pb-based NMs were counted, integrated, and subsequently calibrated to determine PNCs as well as mass and size distributions. PNCs, mean sizes, particulate masses, and ionic background levels were compared across different locations and environments., (© 2022. The Author(s).)

This work introduces novel and universal workflows for the analysis of intact proteins by capillary electrophoresis and presents guidelines for the targeted selection of appropriate background electrolytes (BGEs) by consideration of the target proteins' isoelectric point (pI). The suitability of neutral dimethyl polysiloxane (PDMS) capillaries with dynamic coatings of cationic cetyltrimethylammonium bromide (CTAB) or anionic sodium dodecyl sulfate (SDS), and bare fused silica (BFS) capillaries were systematically evaluated for the analysis of histidine and seven model proteins in six BGEs with pH values between 3.0 and 9.6. Multiple capillary and BGE combinations were suitable for the analysis of all proteins with molecular weights ranging from 13.7-150 kDa, and pIs between 4.7 and 9.6. The CTAB-PDMS capillary was best suited for low pH BGEs, while the SDS-PDMS and BFS capillary were superior for high pH BGEs. These combinations consistently resulted in sharp peak shapes and rapid migration times. pH values of BGEs closer to the proteins' pI produced poorer peak shapes and decreased effective mobilities due to suppressed ionisation. Plots of mobility vs. pH crossed at approximately the pI of the protein in most cases. The workflow was applied to the analysis of caseins and whey proteins in milk for the separation of the seven most abundant proteins, including the isoforms of A1 and A2 β-casein and β-lactoglobulin A and B.

Background: Elements are the basis of life on Earth, whereby organisms are essentially evolved chemical substances that dynamically interact with each other and their environment. Determining species elemental quotas (their elementome) is a key indicator for their success across environments with different resource availabilities. Elementomes remain undescribed for functionally diverse dinoflagellates within the family Symbiodiniaceae that includes coral endosymbionts. We used dry combustion and ICP-MS to assess whether Symbiodiniaceae (ten isolates spanning five genera Breviolum, Cladocopium, Durusdinium, Effrenium, Symbiodinium) maintained under long-term nutrient replete conditions have unique elementomes (six key macronutrients and nine micronutrients) that would reflect evolutionarily conserved preferential elemental acquisition. For three isolates we assessed how elevated temperature impacted their elementomes. Further, we tested whether Symbiodiniaceae conform to common stoichiometric hypotheses (e.g., the growth rate hypothesis) documented in other marine algae. This study considers whether Symbiodiniaceae isolates possess unique elementomes reflective of their natural ecologies, evolutionary histories, and resistance to environmental change., Results: Symbiodiniaceae isolates maintained under long-term luxury uptake conditions, all exhibited highly divergent elementomes from one another, driven primarily by differential content of micronutrients. All N:P and C:P ratios were below the Redfield ratio values, whereas C:N was close to the Redfield value. Elevated temperature resulted in a more homogenised elementome across isolates. The Family-level elementome was (C 19.8 N 2.6 P 1.0 S 18.8 K 0.7 Ca 0.1 ) · 1000 (Fe 55.7 Mn 5.6 Sr 2.3 Zn 0.8 Ni 0.5 Se 0.3 Cu 0.2 Mo 0.1 V 0.04 ) mmol Phosphorous -1 versus (C 25.4 N 3.1 P 1.0 S 23.1 K 0.9 Ca 0.4 ) · 1000 (Fe 66.7 Mn 6.3 Sr 7.2 Zn 0.8 Ni 0.4 Se 0.2 Cu 0.2 Mo 0.2 V 0.05 ) mmol Phosphorous -1 at 27.4 ± 0.4 °C and 30.7 ± 0.01 °C, respectively. Symbiodiniaceae isolates tested here conformed to some, but not all, stoichiometric principles., Conclusions: Elementomes for Symbiodiniaceae diverge from those reported for other marine algae, primarily via lower C:N:P and different micronutrient expressions. Long-term maintenance of Symbiodiniaceae isolates in culture under common nutrient replete conditions suggests isolates have evolutionary conserved preferential uptake for certain elements that allows these unique elementomes to be identified. Micronutrient content (normalised to phosphorous) commonly increased in the Symbiodiniaceae isolates in response to elevated temperature, potentially indicating a common elemental signature to warming., (© 2022. The Author(s).)

Per- and polyfluoroalkyl substances (PFAS) represent a large group of synthetic organic compounds which exhibit unique properties and have been extensively used for consumer and industrial products, resulting in a widespread presence in the environment. Regulation requiring PFAS monitoring has been implemented worldwide due to their potential health and eco-toxicological effects. Targeted methods are commonly used to monitor between twenty to forty PFAS compounds, representing only a small fraction of the number of compounds that may be present. Consequently, there is an increasing interest in complementary non-targeted methods to screen and identify unknown PFAS compounds with the aim to improve knowledge and to generate more accurate models regarding their environmental mobility and persistence. This work details the development of a method that simultaneously provided targeted and non-targeted PFAS analysis. Ultra-high performance liquid chromatography (UHPLC) was coupled to ion mobility-quadrupole time of flight-mass spectrometry (IMS-QTOF-MS) and used to quantify known and screen unknown PFAS in environmental samples collected within the greater Sydney basin (Australia). The method was validated for the quantification of 14 sulfonate-based PFAS, and a non-targeted data analysis workflow was developed using a combination of mass defect analysis with common fragment and neutral loss filtering to identify fluorine-containing species. The optimised method was applied to the environmental samples and enabled the determination of 3-7 compounds from the targeted list and the detection of a further 56-107 untargeted PFAS. This simultaneous analysis reduces the complexity of multiple analyses, and allows for greater interrogation of the full PFAS load in environmental samples., Competing Interests: Declaration of Competing Interest The Vion IMS QTOF is a Waters Australia instrument installed at UTS for collaborative and research use., (Copyright © 2021. Published by Elsevier B.V.)

The discharge of plastic waste and subsequent formation and global distribution of microplastics (MPs) has caused great concern and highlighted the need for dedicated methods to characterise MPs in complex environmental matrices like seawater. Single particle inductively coupled plasma - mass spectrometry (SP ICP-MS) is an elegant method for the rapid analysis of nano- and microparticles and to characterise number concentrations, mass, and size distributions. However, the analysis of carbon (C)-based microstructures such as MPs by SP ICP-MS is at an early stage. This paper investigates various strategies to improve figures of merit to detect and characterise MPs in complex matrices, such as seawater. Ten methods operating distinct acquisition modes with various collision/reaction gases, tandem MS (ICP-MS/MS) and targeting 12 C or 13 C were developed and compared for the analysis of polystyrene-based MPs standards in ultra-pure water and seawater. The robust analysis of MPs in seawater was accomplished by on-line aerosol dilution enabling repeatable size calibration while minimising drift effects. However, the direct analysis of seawater decreased ion transmission and required matrix-matching for accurate size calibration. Analysis of the 12 C isotope instead of 13 C improved the size detection limits (sDL) to 0.62 μm in ultra-pure water and to 0.96 μm in seawater. ICP-MS/MS methods decreased ion transmission but also reduced background signal and increased selectivity, particularly in the presence of spectral interferences. In the second part of this study, it was demonstrated that the developed methods were applicable for the analysis of C in unicellular organisms and allowed calibration of physical dimensions. This is relevant for the investigation and understanding of phenotypical traits associated, for example, with climate change resilience as well as oceanic C storage. SP/SC ICP-MS was employed to target five different intact Symbiodiniaceae algae strains with diverse life-histories in seawater and polystyrene-based MPs were used to calibrate cellular C masses, which were between 51 and 83 pg. The C mass distribution across the analysed unicellular cells was used for modelling cell sizes, which were in the range of 7.6 and 10.1 μm. Determined values were in line with values obtained with complementary techniques (Coulter-counting, total organic C analysis and microscopic analysis)., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

This work demonstrates the first forensic application of GC-ICP-MS for improved investigations of volatile organic compounds originating from a decomposing body. Volatile organic compounds were extracted from the headspace of human remains using sorbent tubes over a total time of 39 days. To account for naturally abundant species, control sites were prepared and sampled accordingly. All samples were spiked with an internal standard to minimise drift effects and errors during sample preparation and further analysis. Compound independent quantification was possible from a single chromatogram with a standard mix containing volatile pesticide compounds representing different mass fractions of target elements for calibration. Phosphorus, sulphur and chlorine were investigated as biologically relevant elements, which potentially form detectable volatile species during decomposition. The limits of detection of these elements in the headspace were 0.7, 5.4 and 1.6 ng/L, respectively. For sulphur, we identified abundant species which increased in concentrations of up to 1310 ng/L in the headspace above the remains. The concentrations were time dependent and show potential as forensic markers to determine post-mortem intervals or decomposition states. The universal quantification, standardisation and the high sensitivity of GC-ICP-MS augments traditional GC-MS analyses., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Standard preparation for elemental bio-imaging by laser ablation-inductively coupled plasma-mass spectrometry is confounded by the chemical and physical differences between standard and sample matrices. These differences lead to variable ablation, aerosol generation and transportation characteristics and must be considered when designing matrix-matched standards for reliable calibration and quantification. The ability to precisely mimic sample matrices is hampered due to the complexity and heterogeneity of biological tissue and small variabilities in standard matrices and sample composition often negatively impact accuracy, precision and robustness. Furthermore, cumbersome preparation protocols may limit reproducibility and traceability. This work presents novel facile methods for the preparation of gelatine standards using both commercial and laboratory-made moulds. Surface roughness, thickness and robustness of the mould-prepared standards were compared against cryo-sectioned gelatine and homogenised brain tissue standards. The mould-prepared standards had excellent thickness accuracy and signal precision which allowed robust quantification, were easier to prepare and therefore easier to reproduce. We also compared gelatine standards prepared from a variety of animal sources and discuss their suitability to calibrate low level elemental concentrations. Finally, we present a simple method to remove background metals in gelatine using various chelating resins to increase the dynamic calibration range and to improve limits of analysis.

Human zinc deficiency increases susceptibility to bacterial infection. Although zinc supplementation therapies can reduce the impact of disease, the molecular basis for protection remains unclear. Streptococcus pneumoniae is a major cause of bacterial pneumonia, which is prevalent in regions of zinc deficiency. We report that dietary zinc levels dictate the outcome of S. pneumoniae infection in a murine model. Dietary zinc restriction impacts murine tissue zinc levels with distribution post-infection altered, and S. pneumoniae virulence and infection enhanced. Although the activation and infiltration of murine phagocytic cells was not affected by zinc restriction, their efficacy of bacterial control was compromised. S. pneumoniae was shown to be highly sensitive to zinc intoxication, with this process impaired in zinc restricted mice and isolated phagocytic cells. Collectively, these data show how dietary zinc deficiency increases sensitivity to S. pneumoniae infection while revealing a role for zinc as a component of host antimicrobial defences., Competing Interests: The authors have declared that no competing interests exist.

This study presents a novel size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) method for the characterisation and quantification of immunoassays with lanthanide-labelled antibodies. SEC-ICP-MS in combination with a double isotope dilution approach enabled facile validation of the antibodies' integrity, the determination of the batch to batch labelling efficiency, monitoring of each labelling step, and quantification of the immunocomplexes after incubation with the target protein. The addition of oxygen into the dynamic reaction cell improved the detection of sulphur as a marker for the antibodies and target protein via mass-shifting (LOD = 3.7 ng/mL), whilst maintaining sufficient sensitivity for the analysis of the lanthanides. Ultra-high performance liquid chromatography (UHPLC) SEC ensured a rapid chromatographic method with separation times under 7 min of the labelled and unlabelled antibodies, the immunocomplexes, and the unconjugated polymer used to lanthanide-label the antibodies. SEC calibration estimated the molecular weights of all peaks and provided valuable insights in immunochemical reactions and the stoichiometry of the reactants and products. A novel on-line isotope dilution analysis (IDA) enabled absolute quantification of sulphur and lanthanide signals and the protein of interest. The chromatographic separation of immunocomplexes and labelled antibodies allowed the simultaneous determination of the antibody/metal stoichiometry and target protein concentration from a single mass flow chromatogram. An immunoglobulin protein was quantified after incubation with an 153 Eu-labelled primary polyclonal antibody. The procedure was validated with direct labelling of the target protein with 156 Gd for parallel, simultaneous quantification. The concentration determined via direct labelling of the protein deviated 1.9% from the immunochemical approach employing 153 Eu-labelled polyclonal antibodies. Graphical abstract.