Directly-Fused Ni(II)Porphyrin Conjugated Polymers with Blocked meso-positions: Impact on Electrocatalytic Properties.

The oxidative coupling reaction of two Ni(II) porphyrins meso-substituted with three and four phenyl groups, Ni(II) 5,10,15-(triphenyl)porphyrin (NiPh3P) and Ni(II) 5,10,15,20-(tetraphenyl)porphyrin (NiPh4P) respectively, was investigated in a oxidative chemical vapor deposition (oCVD) process. Irrespective of the number of meso-substituents, high-resolution mass spectrometry evidences the formation of oligomeric species containing up to five porphyrin units. UV-Vis-NIR and XPS analyses of the oCVD films highlighted a strong dependence of the intermolecular coupling reaction with the substrate temperature. Specifically, higher substrate temperatures yield lowering of valence band maxima and reduction of the band gap. The formation of conjugated polymeric assemblies results in increased conductivities as compared to their sublimed counterparts. Yet, electrocatalytic measurements exhibit water oxidation onset overpotentials (308 mV for pNiPh3P and 343 mV for pNiPh4P) comparatively higher than the onset overpotential measured for the oCVD film from Ni(II) 5,15-(diphenyl)porphyrin (pNiPh2P), i.e. 283 mV. Although DFT and comparative oCVD studies suggest the formation of directly fused porphyrins involving b-b  linkages when reacting tetra-meso-substituted porphyrins, the present findings highlight that multiple direct fusion (b-b/meso-meso/b-b or meso-b/b-meso) is essential for Ni(II) porphyrin-based conjugated polymers to enable a dinuclear radical oxo-coupling operating mechanism for water oxidation at low overpotential and durable catalytic activity.

Multimodal mass spectrometry imaging identifies cell-type-specific metabolic and lipidomic variation in the mammalian liver.

Spatial single-cell omics provides a readout of biochemical processes. It is challenging to capture the transient lipidome/metabolome from cells in a native tissue environment. We employed water gas cluster ion beam secondary ion mass spectrometry imaging ([H2O]n>28K-GCIB-SIMS) at ≤3 µm resolution using a cryogenic imaging workflow. This allowed multiple biomolecular imaging modes on the near-native-state liver at single-cell resolution. Our workflow utilizes desorption electrospray ionization (DESI) to build a reference map of metabolic heterogeneity and zonation across liver functional units at tissue level. Cryogenic dual-SIMS integrated metabolomics, lipidomics, and proteomics in the same liver lobules at single-cell level, characterizing the cellular landscape and metabolic states in different cell types. Lipids and metabolites classified liver metabolic zones, cell types and subtypes, highlighting the power of spatial multi-omics at high spatial resolution for understanding celluar and biomolecular organizations in the mammalian liver.

A Comprehensive Comparison of Tissue Processing Methods for High-Quality MALDI Imaging of Lipids in Reconstructed Human Epidermis.

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has become an important tool for skin analysis, as it allows the simultaneous detection and localization of diverse molecular species within a sample. The use of in vivo and ex vivo human skin models is costly and presents ethical issues; therefore, reconstructed human epidermis (RHE) models, which mimic the upper part of native human skin, represent a suitable alternative to investigate adverse effects of chemicals applied to the skin. However, there are few publications investigating the feasibility of using MALDI MSI on RHE models. Therefore, the aim of this study was to investigate the effect of sample preparation techniques, i.e., substrate, sample thickness, washing, and matrix recrystallization, on the quality of MALDI MSI for lipids analysis of the SkinEthic RHE model. Images were generated using an atmospheric pressure MALDI source coupled to a high-resolution mass spectrometer with a pixel size of 5 µm. Masses detected in a defined region of interest were analyzed and annotated using the LipostarMSI platform. The results indicated that the combination of (1) coated metallic substrates, such as APTES-coated stainless-steel plates, (2) tissue sections of 6 µm thickness, and (3) aqueous washing before HCCA matrix spraying (without recrystallization), resulted in images with a significant signal intensity as well as numerous m/z values. This refined methodology using AP-MALDI coupled to a high-resolution mass spectrometer should improve the current sample preparation workflow to evaluate changes in skin composition after application of dermatocosmetics.

Formate promotes invasion and metastasis in reliance on lipid metabolism.

Metabolic rewiring is essential for cancer onset and progression. We previously showed that one-carbon metabolism-dependent formate production often exceeds the anabolic demand of cancer cells, resulting in formate overflow. Furthermore, we showed that increased extracellular formate concentrations promote the in vitro invasiveness of glioblastoma cells. Here, we substantiate these initial observations with ex vivo and in vivo experiments. We also show that exposure to exogeneous formate can prime cancer cells toward a pro-invasive phenotype leading to increased metastasis formation in vivo. Our results suggest that the increased local formate concentration within the tumor microenvironment can be one factor to promote metastases. Additionally, we describe a mechanistic interplay between formate-dependent increased invasiveness and adaptations of lipid metabolism and matrix metalloproteinase activity. Our findings consolidate the role of formate as pro-invasive metabolite and warrant further research to better understand the interplay between formate and lipid metabolism.

Lipidomic Profiling of PFOA-Exposed Mouse Liver by Multi-Modal Mass Spectrometry Analysis.

Perfluorooctanoic acid (PFOA) is a synthetic perfluorinated chemical classified as a persistent organic pollutant. PFOA has been linked to many toxic effects, including liver injury. Many studies report that PFOA exposure alters serum and hepatic lipid metabolism. However, lipidomic pathways altered by PFOA exposure are largely unknown and only a few lipid classes, mostly triacylglycerol (TG), are usually considered in lipid analysis. Here, we performed a global lipidomic analysis on the liver of PFOA-exposed (high-dose and short-duration) and control mice by combining three mass spectrometry (MS) techniques: liquid chromatography with tandem mass spectrometry (LC-MS/MS), matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), and time-of-flight secondary ion mass spectrometry (TOF-SIMS). Among all hepatic lipids identified by LC-MS/MS analysis, more than 350 were statistically impacted (increased or decreased levels) after PFOA exposure, as confirmed by multi-variate data analysis. The levels of many lipid species from different lipid classes, most notably phosphatidylethanolamine (PE), phosphatidylcholine (PC), and TG, were significantly altered. Subsequent lipidomic analysis highlights the pathways significantly impacted by PFOA exposure, with the glycerophospholipid metabolism being the most impacted, and the changes in the lipidome network, which connects all the lipid species together. MALDI-MSI displays the heterogeneous distribution of the affected lipids and PFOA, revealing different areas of lipid expression linked to PFOA localization. TOF-SIMS localizes PFOA at the cellular level, supporting MALDI-MSI results. This multi-modal MS analysis unveils the lipidomic impact of PFOA in the mouse liver after high-dose and short-term exposure and opens new opportunities in toxicology.

MALDI Mass Spectrometry Imaging: A Potential Game-Changer in a Modern Microbiology.

Nowadays, matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) is routinely implemented as the reference method for the swift and straightforward identification of microorganisms. However, this method is not flawless and there is a need to upgrade the current methodology in order to free the routine lab from incubation time and shift from a culture-dependent to an even faster independent culture system. Over the last two decades, mass spectrometry imaging (MSI) gained tremendous popularity in life sciences, including microbiology, due to its ability to simultaneously detect biomolecules, as well as their spatial distribution, in complex samples. Through this literature review, we summarize the latest applications of MALDI-MSI in microbiology. In addition, we discuss the challenges and avenues of exploration for applying MSI to solve current MALDI-TOF MS limits in routine and research laboratories.

Electronic and energy level engineering of directly fused porphyrin-conjugated polymers - impact of the central metal cation.

The integration of porphyrins and their derivatives in functional devices for solar-assisted fuel production is both highly attractive and challenging due to the difficulties in processing them. This limitation is overcome in the gas-phase approach, particularly by oxidative chemical vapor deposition (oCVD), leading to the simultaneous synthesis and deposition of conjugated porphyrin coatings. We have investigated the impact of the metal cation of 5,15-diphenyl metalloporphyrins (MDPP; M = Co, Cu, Mg, Zn, Pd, Pt, Ag, Ru, Ag, and FeCl) on the dehydrogenative coupling reaction leading to fused-metalloporphyrin thin films via oCVD and on the optoelectronic properties of the resulting thin films. We found that the nature of the chelated cation strongly affects the intermolecular coupling efficiency, as well as the occurrence of side reactions such as chlorination, intramolecular cyclization, demetallation/re-metalation, and oxidation of the porphyrin core. Moreover, we discussed the influence of the above-mentioned reactions on the optoelectronic properties of the fused metalloporphyrin coatings, in view of their potential application in photo-electrocatalytic systems. This study paves the way toward the engineering and future implementation of porphyrin-based systems for clean and efficient solar fuel production.

Evaluation of 6 MALDI-Matrices for 10 µm Lipid Imaging and On-Tissue MSn with AP-MALDI-Orbitrap.

Mass spectrometry imaging is a technique uniquely suited to localize and identify lipids in a tissue sample. Using an atmospheric pressure (AP-) matrix-assisted laser desorption ionization (MALDI) source coupled to an Orbitrap Elite, numerous lipid locations and structures can be determined in high mass resolution spectra and at cellular spatial resolution, but careful sample preparation is necessary. We tested 11 protocols on serial brain sections for the commonly used MALDI matrices CHCA, norharmane, DHB, DHAP, THAP, and DAN in combination with tissue washing and matrix additives to determine the lipid coverage, signal intensity, and spatial resolution achievable with AP-MALDI. In positive-ion mode, the most lipids could be detected with CHCA and THAP, while THAP and DAN without additional treatment offered the best signal intensities. In negative-ion mode, DAN showed the best lipid coverage and DHAP performed superiorly for gangliosides. DHB produced intense cholesterol signals in the white matter. One hundred fifty-five lipids were assigned in positive-ion mode (THAP) and 137 in negative-ion mode (DAN), and 76 peaks were identified using on-tissue tandem-MS. The spatial resolution achievable with DAN was 10 µm, confirmed with on tissue line-scans. This enabled the association of lipid species to single neurons in AP-MALDI images. The results show that the performance of AP-MALDI is comparable to vacuum MALDI techniques for lipid imaging.

Insights on the Atmospheric-Pressure Plasma-Induced Free-Radical Polymerization of Allyl Ether Cyclic Carbonate Liquid Layers.

Plasma-induced free-radical polymerizations rely on the formation of radical species to initiate polymerization, leading to some extent of monomer fragmentation. In this work, the plasma-induced polymerization of an allyl ether-substituted six-membered cyclic carbonate (A6CC) is demonstrated and emphasizes the retention of the cyclic carbonate moieties. Taking advantage of the low polymerization tendency of allyl monomers, the characterization of the oligomeric species is studied to obtain insights into the effect of plasma exposure on inducing free-radical polymerization. In less than 5 min of plasma exposure, a monomer conversion close to 90% is obtained. The molecular analysis of the oligomers by gel permeation chromatography coupled with high-resolution mass spectrometry (GPC-HRMS) further confirms the high preservation of the cyclic structure and, based on the detected end groups, points to hydrogen abstraction as the main contributor to the initiation and termination of polymer chain growth. These results demonstrate that the elaboration of surfaces functionalized with cyclic carbonates could be readily elaborated by atmospheric-pressure plasmas, for instance, by copolymerization.

Visualizing Cholesterol in the Brain by On-Tissue Derivatization and Quantitative Mass Spectrometry Imaging.

Despite being a critical molecule in the brain, mass spectrometry imaging (MSI) of cholesterol has been under-reported compared to other lipids due to the difficulty in ionizing the sterol molecule. In the present work, we have employed an on-tissue enzyme-assisted derivatization strategy to improve detection of cholesterol in brain tissue sections. We report distribution and levels of cholesterol across specific structures of the mouse brain, in a model of Niemann-Pick type C1 disease, and during brain development. MSI revealed that in the adult mouse, cholesterol is the highest in the pons and medulla and how its distribution changes during development. Cholesterol was significantly reduced in the corpus callosum and other brain regions in the Npc1 null mouse, confirming hypomyelination at the molecular level. Our study demonstrates the potential of MSI to the study of sterols in neuroscience.

MSPolyCalc: A web-based App for polymer mass spectrometry data interpretation. The case study of a pharmaceutical excipient.

RATIONALE: In contrast to biological polymers, synthetic macromolecules consist of distributions of sizes, chemical compositions, functionalities and eventually architectures. The mass spectrum of a synthetic polymer may exhibit a tremendous number of signals. The availability of suitable IT tools to support interpretation is key. METHODS: A web-based tool is presented: MSPolyCalc. It offers a set of functionalities, including the calculation of polymer distributions, molecular formulae and a match evaluation for peak assignment based on both mass and spectral accuracy (similarity score). The software was successfully tested with mass spectra exhibiting resolutions ranging from 10,000 to 240,000. RESULTS: The molecular characterization of a synthetic poly(ethylene glycol)-based excipient was achieved. MSPolyCalc allowed the discrimination of six polymer compositions of variable relative abundance. Secondary ionization adducts with very low intensity consisting of matrix-analyte clusters were also successfully identified. CONCLUSIONS: MSPolyCalc offers assisted data interpretation to target the needs of polymer chemists. It facilitates structure characterization, ionization adduct identification, and end-group determination together with visual result reporting.

Reactivity of Nickel(II) Porphyrins in oCVD Processes-Polymerisation, Intramolecular Cyclisation and Chlorination.

Oxidative chemical vapour deposition of (5,15-diphenylporphyrinato)nickel(II) (NiDPP) with iron(III) chloride as oxidant yielded a conjugated poly(metalloporphyrin) as a highly coloured thin film, which is potentially useful for optoelectronic applications. This study clarified the reactive sites of the porphyrin monomer NiDPP by HRMS, UV/Vis/NIR spectroscopy, cyclic voltammetry and EPR spectroscopy in combination with quantum chemical calculations. Unsubstituted meso positions are essential for successful polymerisation, as demonstrated by varying the porphyrin meso substituent pattern from di- to tri- and tetraphenyl substitution. DFT calculations support the proposed radical oxidative coupling mechanism and explain the regioselectivity of the C-C coupling processes. Depositing the conjugated polymer on glass slides and on thermoplastic transparent polyethylene naphthalate demonstrated the suitability of the porphyrin material for flexible optoelectronic devices.

Conductive Fused Porphyrin Tapes on Sensitive Substrates by a Chemical Vapor Deposition Approach.

Oxidative polymerization of nickel(II) 5,15-diphenyl porphyrin and nickel(II) 5,15-bis(di-3,5-tert-butylphenyl) porphyrin by oxidative chemical vapor deposition (oCVD) yields multiply fused porphyrin oligomers in thin film form. The oCVD technique enables one-step formation, deposition, and p-doping of conjugated poly(porphyrins) coatings without solvents or post-treatments. The decisive reactions and side reactions during the oCVD process are shown by high-resolution mass spectrometry. Owing to the highly conjugated structure of the fused tapes, the thin films exhibit an electrical conductivity of 3.6×10-2  S cm-1 and strong absorption in the visible to near-infrared spectral region. The formation of smooth conjugated poly(porphyrins) coatings, even on sensitive substrates, is demonstrated by deposition and patterning on glass, silicon, and paper. Formation of conductive poly(porphyrins) thin films could enable the design of new optoelectronic devices using the oCVD approach.

Plasma-Deposited Nanocapsules Containing Coatings for Drug Delivery Applications.

Coatings consisting in gentamicin-containing nanocapsules have been synthetized by means of an aerosol-assisted atmospheric pressure plasma deposition process. The influence of different parameters affecting the process has been extensively investigated by means of a morphological and chemical characterization of the coatings. Scanning electron microscopy highlighted the presence of nanocapsules whose size and abundance depend on power input and deposition time. A detailed analysis carried out with matrix-assisted laser desorption ionization coupled to high-resolution mass spectrometry allowed to detect and identify the presence of gentamicin embedded in the coatings and its rearrangement, as a result of the interaction with the plasma. The release of gentamicin in water has been monitored by means of UV-vis fluorescence spectroscopy, and its biological activity has been evaluated as well by the disk diffusion assay against Staphylococcus aureus and Pseudomonas aeruginosa. It is confirmed that the antibacterial activity of gentamicin is preserved in the plasma-deposited coatings. Preliminary cytocompatibility investigations indicated that eukaryotic cells well tolerate the release of gentamicin from the coatings.

Determination and imaging of metabolites from Vitis vinifera leaves by laser desorption/ionisation time-of-flight mass spectrometry.

Analysis of grapevine phytoalexins at the surface of Vitis vinifera leaves has been achieved by laser desorption/ionisation time-of-flight mass spectrometry (LDI-ToFMS) without matrix deposition. This simple and rapid sampling method was successfully applied to map small organic compounds at the surface of grapevine leaves. It was also demonstrated that the laser wavelength is a highly critical parameter. Both 266 and 337 nm laser wavelengths were used but the 266 nm wavelength gave increased spatial resolution and better sensitivity for the detection of the targeted metabolites (resveratrol and linked stilbene compounds). Mass spectrometry imaging of grapevine Cabernet Sauvignon leaves revealed specific locations with respect to Plasmopara viticola pathogen infection or light illumination.

Post-source decay of alkylated and functionalized polycyclic aromatic compounds.

Post-source decay (PSD) is a valuable tool for providing structural information from large molecules by time-of-flight mass spectrometry (TOFMS). We used PSD to obtain this type of data from small molecules in the laser desorption/ionization (LDI) study of diesel engine exhaust particles. As the original nitrogen laser (lambda = 337 nm, E = 3.5 eV/photon) of our TOF mass spectrometer does not yield sufficient energy to ionize polycyclic aromatic hydrocarbons (PAHs), a second laser with a shorter wavelength has been coupled to the instrument. The fourth harmonic of a Nd:YAG laser (lambda = 266 nm, 4.6 eV/photon) has been chosen to achieve two-photon single-step desorption/ionization of PAHs. The PSD fragmentation of functionalized, alkylated and sulfur PAHs is discussed. Diesel engine exhaust particles are also studied as an example of a real complex sample. This technique is presented herein as a way to identify small molecules in environmental samples. Information provided by LDI-PSD-TOFMS can be a way to distinguish pollutants with very close molecular weights even if the resolving power of a TOF mass spectrometer is not sufficient.

Isolation and characterization of a gene cluster involved in PAH degradation in Mycobacterium sp. strain SNP11: expression in Mycobacterium smegmatis mc(2)155.

Mycobacterium sp. strain SNP11 is able to grow with pyrene, fluoranthene, phenanthrene and fluorene the sole carbon and energy sources. A probe based on the previously described gene pdoA2, which encodes the alpha subunit of a PAH ring-hydroxylating dioxygenase in Mycobacterium sp. strain 6PY1 [S. Krivobok et al., Identification of pyrene-induced proteins in Mycobacterium sp. strain 6PY1: evidence for two ring-hydroxylating dioxygenases, J. Bacteriol. 185(13) (2003) 3828-3841], was used to isolate a 14kb DNA fragment from strain SNP11. Twelve putative open reading frames (ORFs), divided into two groups by a promoter intergenic region, were detected in this DNA sequence. The first gene cluster, located upstream of the promoter region, showed low but significant deduced amino acid sequence homologies with enzymes involved in aromatic degradation. The second gene cluster, under control of the promoter, contained pdoA2 (designated phdA in this study) and several other ORFs with deduced amino acid sequences closely related to enzymes involved in the phenanthrene-degrading pathway of Nocardioides sp. strain KP7. Gene expression analysis in Mycobacterium smegmatis mc(2)155 revealed broad substrate specificity of the ring-hydroxylating dioxygenase, since transformant cells containing phdAB strongly oxidized fluoranthene, phenanthrene, anthracene, fluorine and dibenzofuran. Laser desorption/ionization time-of-flight mass spectrometry (LDI-ToF MS) analyses of culture media after PAH degradation by M. smegmatis transformants also revealed that the second gene cluster, located downstream of the promoter, takes an active share in initial phenanthrene and anthracene degradation by allowing transformation of these two PAHs in aromatic ring-cleaved metabolites.