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Abstract: In this contribution we present the first example of a microfluidic chip based on BaF2 for Infrared Microspectroscopy (IRMS) of living cells. The advantage in using barium fluoride as platform relies on its high IR transparency, especially in the spectral region below 1300cm−1, where the absorption bands of nucleic acids and carbohydrates are located. Barium fluoride is slightly soluble in water (0.12g/100g water) and it is potentially harmful for living cells. To overcome these problems, here we exploit an approach whose feasibility has been demonstrated previously on CaF2: the surface modification obtained by sputtering a thin Si layer on the surface. The Surface Modified Microfluidic Devices (SM-MD) hence obtained not only solve the BaF2 drawbacks, but also provide a silicon-like substrate fully compatible with standard micro-fabrication processes. These potentialities are here further explored in the direction of chemical or topographical nano-patterning of the silicon-like surface. The silicon thin layer was structured in the shape of 300nm wide grooves (500nm pitch) with a thickness of 35nm by using standard NIL and etching processes; chemical patterning was achieved by exploiting silane chemistry. Finally, we tested the performances of these devices at SISSI beamline@Elettra, collecting IR spectra of single MDA-MB-231 living cells maintained either in physiological solution or complete medium. A comparison of the spectra of a single cell obtained in BaF2 and CaF2 MDs is reported. [Copyright &y& Elsevier]

The main proteases M pro are a group of highly conserved cysteine hydrolases in β-coronaviruses. They have been demonstrated to play an unavoidable role in viral replication, and consequently they have been suggested as key targets for treating coronavirus-caused infectious diseases, mainly from the COVID-19 epidemic. Since the most functional form for M pro enzymatic activity is associated to its homodimer, compounds inhibiting dimerization should also inhibit catalytic activity. We show how PIR-SEIRA (Plasmonic Internal Reflection-Surface Enhanced InfraRed Absorption) spectroscopy can be a noteworthy technique to study proteins subtle structural variations associated to inhibitor binding. Nanoantennas arrays can selectively confine and enhance electromagnetic field via localized plasmonic resonances, thus promoting ultrasensitive detection of biomolecules in close proximity of nanoantenna arrays and enabling the effective investigation of protein monolayers. By adopting this approach, reflection measurements conducted under back illumination of nanoantennas allow to probe anchored protein monolayers, with minimum contribution of environmental buffer molecules. PIR-SEIRA spectroscopy on M pro was carried out by ad hoc designed devices, resonating in the spectral region of Amide I and Amide II bands. We evaluated here the structure of anchored monomers and dimers in different buffered environment and in presence of a newly designed M pro inhibitor. Experimental results show that dimerization is not associated to relevant backbone rearrangements of the protein at secondary structure level, and even if the compound inhibits the dimerization, it is not effective at breaking preformed dimers., 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.)

This study presents a comprehensive multimodal analytical study of an Egyptian ritual Bes-vase, of the 2nd century BCE employing cutting-edge proteomics, metabolomics, genetics techniques, and synchrotron radiation-based Fourier Transformed Infrared microSpectroscopy (SR µ-FTIR) to characterize organic residues of its content. We successfully identified the presence of various functional, bioactive, psychotropic, and medicinal substances, shedding light on the diverse components of a liquid concoction used for ritual practices in Ptolemaic Egypt. Using LC-MS/MS with a new methodological approach, we identified key proteins and metabolites, enabling the identification of botanical sources, confirmed by genetic sequences. Our analyses revealed traces of Peganum harmala, Nimphaea nouchali var. caerulea, and a plant of the Cleome genus, all of which are traditionally proven to have psychotropic and medicinal properties. Additionally, the identification of human fluids suggests their direct involvement in these rituals. Furthermore, metabolomics and SR µ-FTIR analyses also revealed the presence of fermented fruit-based liquid and other ingredients such as honey or royal jelly. The identification of specific chemical compounds, such as alkaloids and flavonoids, provides insight into the psychoactive and therapeutic uses of these in ancient ritual practices. This multidisciplinary study highlights the complexity of ancient cultures and their interactions with psychoactive, medicinal, and bioactive substances. These findings contribute to our understanding of ancient belief systems, cultural practices, and the utilization of natural resources, ultimately enhancing our knowledge of past societies and their connection to the natural world., Competing Interests: Declarations Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Asbestos fiber exposure triggers chronic inflammation and cancer. Asbestos fibers can adsorb different types of proteins. The mechanism of this adsorption, not yet completely understood, has been studied in detail mainly with serum albumin and was shown to induce structural changes in the bound protein. The findings of these works regarded mainly the changes of the protein structure, independently of any relation with asbestos-related diseases. For the first time, we have focused our attention to the consequences of the interaction between asbestos fibers and ferritin, a protein involved in iron metabolism, which is strongly modified in asbestos-related diseases. Even if it is known that ferritin can be adsorbed by asbestos fibers, the results of this interaction for the ferritin secondary structure has not previously been studied. One consequence of asbestos-ferritin interaction, is the formation of the so-called ferruginous/asbestos bodies (ABs). In the AB-coating material, the secondary structure of ferritin is modified, and at present, it is unclear whether or not this modification is a direct consequence of the asbestos interaction. In the present study, chrysotile asbestos, more than other asbestos fiber types tested, was found to rapidly bind holo-ferritin, and the presence of iron seemed to play a key role in this process, since iron-free apo-ferritin was adsorbed at a lower level, and iron-saturated chrysotile lost its ferritin-adsorbing capacity. To directly study the details of ferritin adsorption on asbestos fibers, High Resolution Transmission Electron Microscopy (HR-TEM) was employed together with FTIR microspectroscopy and Infrared nanoscopy, which to the best of our knowledge, have not previously been used for this purpose. Chrysotile-bound apo-ferritin underwent a significant change in secondary structure, showing a shift from a prevalent α-helix to a β-sheet conformation. Conversely, the adsorbed holo-ferritin structure appeared to be only weakly modified. These findings add a new potential mechanism to the toxic activities of asbestos: the fibers can modify the structure, and very likely, the function of adsorbed proteins. This, in relation to ferritin, could be a key mechanism in cell iron homeostasis alteration, typically reported in asbestos-related diseases., Competing Interests: 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., (© 2024 The Authors.)

Guanosine nucleosides and nucleotides have the peculiar ability to self-assemble in water to form supramolecular complex architectures from G-quartets to G-quadruplexes. G-quadruplexes exhibit in turn a large liquid crystalline lyotropic polymorphism, but they eventually cross-link or entangle to form a densely connected 3D network (a molecular hydrogel), able to entrap very large amount of water (up to the 99% v/v). This high water content of the hydrogels enables tunable softness, deformability, self-healing, and quasi-liquid properties, making them ideal candidates for different biotechnological and biomedical applications. In order to fully exploit their possible applications, Attenuated Total Reflection-Fourier Transform InfraRed (ATR-FTIR) spectroscopy was used to unravel the vibrational characteristics of supramolecular guanosine structures. First, the characteristic vibrations of the known quadruplex structure of guanosine 5 ' -monophosphate, potassium salt (GMP/K), were investigated: the identified peaks reflected both the chemical composition of the sample and the formation of quartets, octamers, and quadruplexes. Second, the role of K + and Na + cations in promoting the quadruplex formation was assessed: infrared spectra confirmed that both cations induce the formation of G-quadruplexes and that GMP/K is more stable in the G-quadruplex organization. Finally, ATR-FTIR spectroscopy was used to investigate binary mixtures of guanosine (Gua) and GMP/K or GMP/Na, both systems forming G-hydrogels. The same G-quadruplex-based structure was found in both mixtures, but the proportion of Gua and GMP affected some features, like sugar puckering, guanine vibrations, and base stacking, reflecting the known side-to-side aggregation and bundle formation occurring in these binary systems., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Valentina Notarstefano reports financial support was provided by European Union. Paolo Mariani reports financial support was provided by European Union. The other 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.)

Water under soft nanoconfinement features physical and chemical properties fundamentally different from bulk water; yet, the multitude and specificity of confining systems and geometries mask any of its potentially universal traits. Here, we advance in this quest by resorting to lipidic mesophases as an ideal nanoconfinement system, allowing inspecting the behavior of water under systematic changes in the topological and geometrical properties of the confining medium, without altering the chemical nature of the interfaces. By combining Terahertz absorption spectroscopy experiments and molecular dynamics simulations, we unveil the presence of universal laws governing the physics of nanoconfined water, recapitulating the data collected at varying levels of hydration and nanoconfinement topologies. This geometry-independent universality is evidenced by the existence of master curves characterizing both the structure and dynamics of simulated water as a function of the distance from the lipid-water interface. Based on our theoretical findings, we predict a parameter-free law describing the amount of interfacial water against the structural dimension of the system (i.e., the lattice parameter), which captures both the experimental and numerical results within the same curve, without any fitting. Our results offer insight into the fundamental physics of water under soft nanoconfinement and provide a practical tool for accurately estimating the amount of nonbulk water based on structural experimental data.

Many years ago, asbestos fibers were banned and replaced by synthetic vitreous fibers because of their carcinogenicity. However, the toxicity of the latter fibers is still under debate, especially when it concerns the early fiber interactions with biological cell membranes. Here, we aimed to investigate the effects of a synthetic vitreous fiber named FAV173 on the Xenopus laevis oocyte membrane, the cell model we have already used to characterize the effect of crocidolite asbestos fiber exposure. Using an electrophysiological approach, we found that, similarly to crocidolite asbestos, FAV173 was able to stimulate a chloride outward current evoked by step membrane depolarizations, that was blocked by the potent and specific TMEM16A channel antagonist Ani9. Exposure to FAV173 fibers also altered the oocyte cell membrane microvilli morphology similarly to crocidolite fibers, most likely as a consequence of the TMEM16A protein interaction with actin. However, FAV173 only partially mimicked the crocidolite fibers effects, even at higher fiber suspension concentrations. As expected, the crocidolite fibers' effect was more similar to that induced by the co-treatment with (Fe 3+ + H 2 O 2 ), since the iron content of asbestos fibers is known to trigger reactive oxygen species (ROS) production. Taken together, our findings suggest that FAV173 may be less harmful that crocidolite but not ineffective in altering cell membrane properties.

The study investigated the application of Wastewater-Based Epidemiology (WBE) as a tool for monitoring the SARS-CoV-2 prevalence in a city in northern Italy from October 2021 to May 2023. Based on a previously used deterministic model, this study proposed a variation to account for the population characteristics and virus biodegradation in the sewer network. The model calculated virus loads and corresponding COVID-19 cases over time in different areas of the city and was validated using healthcare data while considering viral mutations, vaccinations, and testing variability. The correlation between the predicted and reported cases was high across the three waves that occurred during the period considered, demonstrating the ability of the model to predict the relevant fluctuations in the number of cases. The population characteristics did not substantially influence the predicted and reported infection rates. Conversely, biodegradation significantly reduced the virus load reaching the wastewater treatment plant, resulting in a 30% reduction in the total virus load produced in the study area. This approach can be applied to compare the virus load values across cities with different population demographics and sewer network structures, improving the comparability of the WBE data for effective surveillance and intervention strategies.

In veterinary medicine, Cannabis has been used to treat pain conditions, inflammation, and seizures. However, little is known about its effect on dogs' behavior. This preliminary research aims to address this knowledge gap by evaluating the effectiveness of cannabidiol (CBD) oil in canine behavioral therapy. Twenty dogs, diagnosed with behavioral disorders and housed in a municipal shelter, participated in a double-blind trial. Ten dogs received CBD oil treatment, while the other ten received a control oil without CBD. Before (T0) and after (T1) the treatment, all the dogs underwent a temperament test to assess their behavior in the presence of four different stimuli: a human stranger, a novel object, a child-like doll, and a conspecific (another dog). Each stimulus was presented individually, and the dogs' behaviors were recorded on video and analyzed. Additionally, hair samples were collected using a shave-reshave technique for cortisol determination through Radio-Immuno-Assay. No behavioral differences were found between the two groups at both T0 and T1. There were no significant differences in the behavioral responses of either group when comparing T0 and T1. However, individual responses to the CBD oil treatment appeared to vary among subjects. A significant increase in hair cortisol levels (p-value <0.05) was observed in the group treated with CBD oil [T0 = 1.60 (1.44-1.93) pg/mg, T1 = 4.81(2.57-6.01) pg/mg]. These findings highlight the importance of individualized treatment when using Cannabis and encourage further research on the use of CBD oil in animal behavioral medicine., Competing Interests: 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., (© 2024 The Authors.)

The divergent supramolecular behavior of a series of tripeptide stereoisomers was elucidated through spectroscopic, microscopic, crystallographic, and computational techniques. Only two epimers were able to effectively self-organize into amphipathic structures, leading to supramolecular hydrogels or crystals, respectively. Despite the similarity between the two peptides' turn conformations, stereoconfiguration led to different abilities to engage in intramolecular hydrogen bonding. Self-assembly further shifted the p K a value of the C-terminal side chain. As a result, across the pH range 4-6, only one epimer predominated sufficiently as a zwitterion to reach the critical molar fraction, allowing gelation. By contrast, the differing p K a values and higher dipole moment of the other epimer favored crystallization. The four stereoisomers were further tested for gold nanoparticle (AuNP) formation, with the supramolecular hydrogel being the key to control and stabilize AuNPs, yielding a nanocomposite that catalyzed the photodegradation of a dye. Importantly, the AuNP formation occurred without the use of reductants other than the peptide, and the redox chemistry was investigated by LC-MS, NMR, and infrared scattering-type near field optical microscopy (IR s-SNOM). This study provides important insights for the rational design of simple peptides as minimalistic and green building blocks for functional nanocomposites.

Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are the major components of long-chain per- and polyfluorinated alkyl substances (PFAS), known for their chemical stability and environmental persistence. Even if PFOA and PFOS have been phased out or are limited in use, they still represent a concern for human and environmental health. Several studies have been per­formed to highlight the toxicological behavior of these chemicals and their mode of action (MoA). Data have suggested a causal association between PFOA or PFOS exposure and carcinogenicity in humans, but the outcomes of epidemiological studies showed some inconsistency. Moreover, the hypothesized MoA based on animal studies is considered not relevant for human cancer. To improve the knowledge on PFAS toxicology and contribute to the weight of evidence for the regu­latory classification of PFAS, we used the BALB/c 3T3 cell transformation assay (CTA), an in vitro model under consideration to be included in an integrated approach to testing and assessment for non-genotoxic carcinogens (NGTxCs). PFOS and PFOA were tested at several concentrations using a validated experimental protocol. Our results demonstrate that PFOA does not induce cell transformation, whereas PFOS exposure induced a concentration-related increase of type III foci. Malignant foci formation was triggered at PFOS concentrations equal to or higher than 50 ppm and was not directly associated with cytotoxicity or proliferation induction. The divergent CTA outcomes suggest that different molecular events could be responsible for the toxicological profiles of PFOS and PFOA, which were not fully captured in our study.

Cryopreservation affects platelets' function, questioning their use for cancer patients. We aimed to investigate the biochemical events that occur over time after thawing to optimize transfusion timing and evaluate the effect of platelet supernatants on tumor cell behavior in vitro . We compared fresh (Fresh-PLT) with Cryopreserved platelets (Cryo-PLT) at 1 h, 3 h and 6 h after thawing. MCF-7 and HL-60 cells were cultured with Fresh- or 1 h Cryo-PLT supernatants to investigate cell proliferation, migration, and PLT-cell adhesion. We noticed a significant impairment of hemostatic activity accompanied by a post-thaw decrease of CD42b + , which identifies the CD62P - -population. FTIR spectroscopy revealed a decrease in the total protein content together with changes in their conformational structure, which identified two sub-groups: 1) Fresh and 1 h Cryo-PLT; 2) 3 h and 6 h cryo-PLT. Extracellular vesicle shedding and phosphatidylserine externalization (PS) increased after thawing. Cryo-PLT supernatants inhibited cell proliferation, impaired MCF-7 cell migration, and reduced ability to adhere to tumor cells. Within the first 3 hours after thawing, irreversible alterations of biomolecular structure occur in Cryo-PLT. Nevertheless, Cryo-PLT should be considered safe for the transfusion of cancer patients because of their insufficient capability to promote cancer cell proliferation, adhesion, or migration.

Infrared scattering-type scanning near-field optical microscopy (IR s-SNOM) and imaging is here exploited together with attenuated total reflection (ATR) IR imaging and scanning electron microscopy (SEM) to depict the chemical composition of fibers in hybrid electrospun meshes. The focus is on a recently developed bio-hybrid material for vascular tissue engineering applications, named Silkothane®, obtained in the form of nanofibrous matrices from the processing of a silk fibroin-polyurethane (SFPU) blend via electrospinning. Morphology and chemistry of single fibers, at both surface and subsurface level, have been successfully characterized with nanoscale resolution, taking advantage of the IR s-SNOM capability to portray the nanoscale depth profile of this modern material working at diverse harmonics of the signal. The applied methodology allowed to describe the superficial characteristics of the mesh up to a depth of about 100 nm, showing that SF and PU do not tend to co-aggregate to form hybrid fibers, at least at the length scale of hundreds of nanometers, and that subdomains other than the fibrillar ones can be present. More generally, in the present contribution, the depth profiling capabilities of IR s-SNOM, so far theoretically predicted and experimentally proven only on model systems, have been corroborated on a real material in its natural conditions with respect to production, opening the room for the exploitation of IR s-SNOM as valuable technique to support the production and the engineering of nanostructured materials by the precise understanding of their chemistry at the interface with the environment.

Research on the effects of engineered nanomaterials (ENMs) on mitochondria, which represent one of the main actors in cell function, highlighted effects on ROS production, gametogenesis and organellar genome replication. Specifically, the mitochondrial effects of cadmium sulfide quantum dots (CdS QDs) exposure can be observed through the variation in enzymatic kinetics at the level of the respiratory chain and also by analyzing modifications of reagent and products in term of the bonds created and disrupted during the reactions through Fourier-transform infrared spectroscopy (FTIR). This study investigated both in intact cells and in isolated mitochondria to observe the response to CdS QDs treatment at the level of electron transport chain in the wild-type yeast Saccharomyces cerevisiae and in the deletion mutant Δtom5 , whose function is implicated in nucleo-mitochondrial protein trafficking. The changes observed in wild type and Δtom5 strains in terms of an increase or decrease in enzymatic activity (ranging between 1 and 2 folds) also differed according to the genetic background of the strains and the respiratory chain functionality during the CdS QDs treatment performed. Results were confirmed by FTIR, where a clear difference between the QD effects in the wild type and in the mutant strain, Δtom5 , was observed. The utilization of these genetic and biochemical approaches is instrumental to clarify the mitochondrial mechanisms implicated in response to these types of ENMs and to the stress response that follows the exposure.

Purpose: Knowledge about pancreatic cancer (PC) biology has been growing rapidly in recent decades. Nevertheless, the survival of PC patients has not greatly improved. The development of a novel methodology suitable for deep investigation of the nature of PC tumors is of great importance. Molecular imaging techniques, such as Fourier transform infrared (FTIR) spectroscopy and Raman hyperspectral mapping (RHM) combined with advanced multivariate data analysis, were useful in studying the biochemical composition of PC tissue., Methods: Here, we evaluated the potential of molecular imaging in differentiating three groups of PC tumors, which originate from different precursor lesions. Specifically, we comprehensively investigated adenocarcinomas (ACs): conventional ductal AC, intraductal papillary mucinous carcinoma, and ampulla of Vater AC. FTIR microspectroscopy and RHM maps of 24 PC tissue slides were obtained, and comprehensive advanced statistical analyses, such as hierarchical clustering and nonnegative matrix factorization, were performed on a total of 211,355 Raman spectra. Additionally, we employed deep learning technology for the same task of PC subtyping to enable automation. The so-called convolutional neural network (CNN) was trained to recognize spectra specific to each PC group and then employed to generate CNN-prediction-based tissue maps. To identify the DNA methylation spectral markers, we used differently methylated, isolated DNA and compared the observed spectral differences with the results obtained from cellular nuclei regions of PC tissues., Results: The results showed significant differences among cancer tissues of the studied PC groups. The main findings are the varying content of β-sheet-rich proteins within the PC cells and alterations in the relative DNA methylation level. Our CNN model efficiently differentiated PC groups with 94% accuracy. The usage of CNN in the classification task did not require Raman spectral data preprocessing and eliminated the need for extensive knowledge of statistical methodologies., Conclusions: Molecular spectroscopy combined with CNN technology is a powerful tool for PC detection and subtyping. The molecular fingerprint of DNA methylation and β-sheet cytoplasmic proteins established by our results is different for the main PC groups and allowed the subtyping of pancreatic tumors, which can improve patient management and increase their survival. Our observations are of key importance in understanding the variability of PC and allow translation of the methodology into clinical practice by utilizing liquid biopsy testing., (© 2023. The Author(s).)

Honey is a natural food appreciated all over the world since antiquity due to its well-recognised beneficial properties. However, it is also considered among the most counterfeited foods. Therefore, analytical methods are currently being developed to allow the verifying of its geographic provenance and its botanical origin. Trace- and ultra-trace elements are usually exploited as chemical descriptors in authentication studies, as they allow the properties declared in the label to be verified. A different matter is to trace a food by means of traceability, that is, to find the link between a food and the soil in which this food originates. For traceability, it has been demonstrated in several studies that the lanthanides are particularly useful to find this link. In the present study, the traceability of the honey chain has been studied by means of ICP-MS and ICP-OES analysis, by comparing the lanthanide distributions of 17 different monofloral honey chains, each one composed of honey, flowers and soil in which such flowers grew. The results show that, while the fingerprint of soil, described by the lanthanide distribution, is transmitted unaltered from soil to flowers, a slight fractionation on the heavier lanthanides (from Dy to Lu) occurs in the passage from flowers to honey.

Core-shell nanoparticles have been extensively researched, particularly as multimodal for medical applications. Scientists are interested in combining the optical properties of nano-plasmonic nanoparticles with the magnetic properties of super-paramagnetic nanoparticles. This combination is very important because it reduces metal toxicity and improves nanoparticle targeting. Tuning the shape and size of the nanoparticles significantly reflects their properties and applications. In previous study, we assessed the SPION@Ag@chitosan core-shell nanocomposite (γFe2O3@Ag@Cs NCs) toxicity both in vitro and preclinically in vivo, using traditional toxicological assessment and biochemical parameters. The results showed that up to100 ug/kg is a safe NP dose as evaluated by pathological and biochemical parameters. The aim of the present study was to gain insight into the effect of γFe2O3@Ag@Cs NC at sub-cytotoxic concentrations (100ug/ml) on the biochemical profile of immune organs (inguinal, axillary, spleen and thymus) by combining the investigation of cytokine secretion to ex vivo FTIR spectroscopy. The four immune organs were treated with 100 ug/kg NC and the time dependence of the effects produced by the treatment was analyzed. The Data shows that the used core-shell NC with the indicate dose have a stimulatory effect on the immune system, as evidenced by an increase in antibody secretion (IgG and IgM), lipid, nucleic acid, and protein synthesis after uptake time which depends on the specific immune organ., 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 © 2022 Elsevier B.V. All rights reserved.)

Multiple sclerosis (MS) and its preclinical models are characterized by marked changes in neuroplasticity, including excitatory/inhibitory imbalance and synaptic dysfunction that are believed to underlie the progressive cognitive impairment (CI), which represents a significant clinical hallmark of the disease. In this study, we investigated several parameters of neuroplasticity in the hippocampus of the experimental autoimmune encephalomyelitis (EAE) SJL/J mouse model, characterized by rostral inflammatory and demyelinating lesions similar to Relapsing-Remitting MS. By combining morphological and molecular analyses, we found that the hippocampus undergoes extensive inflammation in EAE-mice, more pronounced in the CA3 and dentate gyrus (DG) subfields than in the CA1, associated with changes in GABAergic circuitry, as indicated by the increased expression of the interneuron marker Parvalbumin selectively in CA3. By laser-microdissection, we investigated the impact of EAE on the alternative splicing of Arhgef9 , a gene encoding a post-synaptic protein playing an essential role in GABAergic synapses and whose mutations have been related to CI and epilepsy. Our results indicate that EAE induces a specific increase in inclusion of the alternative exon 11a only in the CA3 and DG subfields, in line with the higher local levels of inflammation. Consistently, we found a region-specific downregulation of Sam68, a splicing-factor that represses this splicing event. Collectively, our findings confirm a regionalized distribution of inflammation in the hippocampus of EAE-mice. Moreover, since neuronal circuit rearrangement and dynamic remodeling of structural components of the synapse are key processes that contribute to neuroplasticity, our study suggests potential new molecular players involved in EAE-induced hippocampal dysfunction., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Adinolfi, Di Sante, Rivignani Vaccari, Tredicine, Ria, Bonvissuto, Corvino, Sette and Geloso.)

In this work, polyvinylidene fluoride (PVDF) aerogels with a tailorable phase composition were prepared by following the crystallization-induced gelation principle. A series of PVDF wet gels (5 to 12 wt.%) were prepared from either PVDF−DMF solutions or a mixture of DMF and ethanol as non-solvent. The effects of the non-solvent concentration on the crystalline composition of the PVDF aerogels were thoroughly investigated. It was found that the nucleating role of ethanol can be adjusted to produce low-density PVDF aerogels, whereas the changes in composition by the addition of small amounts of water to the solution promote the stabilization of the valuable β and γ phases. These phases of the aerogels were monitored by FTIR and Raman spectroscopies. Furthermore, the crystallization process was followed by in-time and in situ ATR−FTIR spectroscopy. The obtained aerogels displayed specific surface areas > 150 m2 g−1, with variable particle morphologies that are dependent on the non-solvent composition, as observed by using SEM and Synchrotron Radiation Computed micro-Tomography (SR-μCT).

It is well-known that all the phases of the manufacturing influence the extraordinary aesthetic and acoustic features of Stradivari's instruments. However, these masterpieces still keep some of their secrets hidden by the lack of documentary evidence. In particular, there is not a general consensus on the use of a protein-based ground coating directly spread on the wood surface by the Cremonese Master. The present work demonstrates that infrared scattering-type scanning near-fields optical microscopy (s-SNOM) may provide unprecedented information on very complex cross-sectioned microsamples collected from two of Stradivari's violins, nanoresolved chemical sensitivity being the turning point for detecting minute traces of a specific compound, namely proteins, hidden by the matrix when macro or micro sampling approaches are exploited. This nanoresolved chemical-sensitive technique contributed new and robust evidence to the long-debated question about the use of proteinaceous materials by Stradivari.

Superparamagnetic iron oxide nanoparticles (SPIONs) have great potential for use in medicine, but they may cause side effects due to oxidative stress. In our study, we investigated the effects of silica-coated SPIONs on endothelial cells and whether oleic acid (OA) can protect the cells from their harmful effects. We used viability assays, flow cytometry, infrared spectroscopy, fluorescence microscopy, and transmission electron microscopy. Our results show that silica-coated SPIONs are internalized by endothelial cells, where they increase the amount of reactive oxygen species (ROS) and cause cell death. Exposure to silica-coated SPIONs induced accumulation of lipid droplets (LD) that was not dependent on diacylglycerol acyltransferase (DGAT)-mediated LD biogenesis, suggesting that silica-coated SPIONs suppress LD degradation. Addition of exogenous OA promoted LD biogenesis and reduced SPION-dependent increases in oxidative stress and cell death. However, exogenous OA protected cells from SPION-induced cell damage even in the presence of DGAT inhibitors, implying that LDs are not required for the protective effect of exogenous OA. The molecular phenotype of the cells determined by Fourier transform infrared spectroscopy confirmed the destructive effect of silica-coated SPIONs and the ameliorative role of OA in the case of oxidative stress. Thus, exogenous OA protects endothelial cells from SPION-induced oxidative stress and cell death independent of its incorporation into triglycerides.

Isofuranodiene (IFD) is a sesquiterpene occurring in several plant species, which proved to have multiple anticancer activities. IFD has a lipophilic nature and, hence, a very low water solubility and a poor bioavailability; moreover, it is not stable, undergoing the "Cope rearrangement" to the less active curzerene. The use of appropriate delivery systems can thus be considered as a valid tool to enhance IFD bioavailability, solubility, stability and at the same time also to improve its intracellular uptake and pharmacological activity. Within this frame, monoolein (GMO) nanoparticles loaded with IFD were prepared and their enhanced anticancer activity, compared to pristine IFD, was assessed. In this study, for the first time, an in vitro Fourier Transform Infrared and Raman Microspectroscopy approaches were exploited to evaluate the effects of IFD, alone and loaded in GMO nanoparticles, on MDA-MB 231 breast cancer cell line. The anti-cancer effects of IFD were evidenced by both the spectroscopic techniques and discriminated from the GMO-induced changes in the culture environment; moreover, a synergistic effect of IFD and GMO administration can be envisaged by the experimental results., 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 study investigates the occurrence of anthropogenic fibres inside wild Paracentrotus lividus at a Mediterranean coastal area in 2020. From each sea urchin, the coelomic fluid was directly analysed while digestive tracts and gonads were removed, pre-treated with trypsin (0.3%) and digested with H 2 O 2 (10%) before analysis. A total of 260 fibres and 1 fragment were found in 100 specimens, with an average of 2.6 items/individual. Fibres were more abundant in the digestive system, less in gonads and in the coelomic fluid, respectively. Fourier transform infrared (FTIR) analysis of representative fibres identified 67% natural (cotton-based) and 33% synthetic polymers (polyester) suggesting their origin from textiles, possibly released from laundry sewages. Overall, these results encourage further in-depth investigations on fibres accumulation and potential transfer through the trophic chain up to humans., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Glioblastoma multiforme (GBM) is the most aggressive brain tumor, characterized by short median survival and an almost 100% tumor-related mortality. The standard of care treatment for newly diagnosed GBM includes surgical resection followed by concomitant radiochemotherapy. The prevention of disease progression fails due to the poor therapeutic effect caused by the great molecular heterogeneity of this tumor. Previously, we exploited synchrotron radiation-based soft X-ray tomography and hard X-ray fluorescence for elemental microimaging of the shock-frozen GBM cells. The present study focuses instead on the biochemical profiling of live GBM cells and provides new insight into tumor heterogenicity. We studied bio-macromolecular changes by exploring the live-cell synchrotron-based Fourier transform infrared (SR-FTIR) microspectroscopy in a set of three GBM cell lines, including the patient-derived glioblastoma cell line, before and after riluzole treatment, a medicament with potential anticancer properties. SR-FTIR microspectroscopy shows that GBM live cells of different origins recruit different organic compounds. The riluzole treatment of all GBM cell lines mainly affected carbohydrate metabolism and the DNA structure. Lipid structures and protein secondary conformation are affected as well by the riluzole treatment: cellular proteins assumed cross β-sheet conformation while parallel β-sheet conformation was less represented for all GBM cells. Moreover, we hope that a new live-cell approach for GBM simultaneous treatment and examination can be devised to target cancer cells more specifically, i.e., future therapies can develop more specific treatments according to the specific bio-macromolecular signature of each tumor type.

A main factor hampering life in space is represented by high atomic number nuclei and energy (HZE) ions that constitute about 1% of the galactic cosmic rays. In the frame of the "STARLIFE" project, we accessed the Heavy Ion Medical Accelerator (HIMAC) facility of the National Institute of Radiological Sciences (NIRS) in Chiba, Japan. By means of this facility, the extremophilic species Haloterrigena hispanica and Parageobacillus thermantarcticus were irradiated with high LET ions (i.e., Fe, Ar, and He ions) at doses corresponding to long permanence in the space environment. The survivability of HZE-treated cells depended upon either the storage time and the hydration state during irradiation; indeed, dry samples were shown to be more resistant than hydrated ones. With particular regard to spores of the species P. thermantarcticus , they were the most resistant to irradiation in a water medium: an analysis of the changes in their biochemical fingerprinting during irradiation showed that, below the survivability threshold, the spores undergo to a germination-like process, while for higher doses, inactivation takes place as a consequence of the concomitant release of the core's content and a loss of integrity of the main cellular components. Overall, the results reported here suggest that the selected extremophilic microorganisms could serve as biological model for space simulation and/or real space condition exposure, since they showed good resistance to ionizing radiation exposure and were able to resume cellular growth after long-term storage.

The actual chemical structure of polyfurfuryl alcohol (PFA) is still uncertain in spite of several studies on the topic, variations during the polymerization processes being one reason that must be addressed. The use of a limited set of analytical techniques is often insufficient to provide an exhaustive chemical characterization. Moreover, it is still not possible to exactly determine presence and amount of each specific functional group in the polymeric structure. We employed both Fourier Transform Infrared Spectroscopy (FTIR) and Resonant Raman spectroscopy (RR), corroborated by quantum mechanically aided analysis of the experimental spectra, to infer about the chemical structure of two samples of PFAs, synthetized in different ways and appearing macroscopically different, the first one being a liquid and viscous commercial sample, the second one being a self-prepared solid and rigid sample produced following a thermosetting procedure. The vibrational spectroscopic analysis confirms the presence of differences in their chemical structures. The viscous form of PFA is mainly composed by short polymeric chains, and is characterized by the presence of isolated furfuryl alcohol and furfural residues similar to 5-hydroxymethylfurfural; the thermosetted PFA is formed by more cross-linked structures, characterized by several ketones and alkene double bonds, as well as a significant presence of Diels-Alder structures. In summary, the present study evidences how the use of both FTIR and RR spectroscopy, the latter carried out at several laser excitation wavelengths, indicates an accurate way to spectroscopically investigate complex polymers enabling to satisfactorily infer about their peculiar chemical structure., 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.)

Tannin-furanic rigid foams are bio-based copolymers of tannin plant extract and furfuryl alcohol, promising candidates to replace synthetic insulation foams, as for example polyurethanes and phenolics, in eco-sustainable buildings thanks to their functional properties, such as lightness of the material and fire resistance. Despite their relevance as environmental-friendly alternatives to petroleum derivatives, many aspects of the polymerization chemistry still remain unclear. One of the open issues is on the spatial heterogeneity of the foam, i.e., whether the foam constituents prevalently polymerize in spatially segregated blocks or distribute almost homogenously in the foam volume. To address this matter, here we propose a multiscale FTIR study encompassing 1D FTIR spectroscopy, 2D FTIR imaging and 3D FTIR micro-tomography (FTIR-μCT) on tannin-furanic rigid foams obtained by varying the synthesis parameters in a controlled way. Thanks to the implementation of the acquisition and processing pipeline of FTIR-μCT, we were able for the first time to demonstrate that the polymer formulations influence the spatial organization of the foam at the microscale and, at the same time, prove the reliability of FTIR-μCT data by comparing 2D FTIR images and the projection of the 3D chemical images on the same plane.

Label-free spectromicroscopy methods offer the capability to examine complex cellular phenomena. Electron and X-ray based spectromicroscopy methods, though powerful, have been hard to implement with hydrated objects due to the vacuum incompatibility of the samples and due to the parasitic signals from (or drastic attenuation by) the liquid matrix surrounding the biological object of interest. Similarly, for many techniques that operate at ambient pressure, such as Fourier transform infrared spectromicroscopy (FTIRM), the aqueous environment imposes severe limitations due to the strong absorption of liquid water in the infrared regime. Here we propose a microfabricated multi-compartmental and reusable hydrated sample platform suitable for use with several analytical techniques, which employs the conformal encapsulation of biological specimens by a few layers of atomically thin graphene. Such an electron, X-ray, and infrared transparent, molecularly impermeable and mechanically robust enclosure preserves the hydrated environment around the object for a sufficient time to allow in situ examination of hydrated bio-objects with techniques operating under both ambient and high vacuum conditions. An additional hydration source, provided by hydrogel pads lithographically patterned in the liquid state near/around the specimen and co-encapsulated, has been added to further extend the hydration lifetime. Note that the in-liquid lithographic electron beam-induced gelation procedure allows for addressable capture and immobilization of the biological cells from the solution. Scanning electron microscopy and optical fluorescence microscopy, as well as synchrotron radiation based FTIR and X-ray fluorescence microscopy, have been used to test the applicability of the platform and for its validation with yeast, A549 human carcinoma lung cells and micropatterned gels as biological object phantoms.

Amyloids are proteinaceous deposits considered an underlying pathological hallmark of several degenerative diseases. The mechanism of amyloid formation and its inhibition still represent challenging issues, especially when protein structure cannot be investigated by classical biophysical techniques as for the intrinsically disordered proteins (IDPs). In this view, the need to find an alternative way for providing molecular and structural information regarding IDPs prompted us to set a novel, to our knowledge, approach focused on UV Resonance Raman (UVRR) spectroscopy. To test its applicability, we study the fibrillation of hen-egg white lysozyme (HEWL) and insulin as well as their interaction with resveratrol, employing also intrinsic fluorescence spectroscopy, Fourier transform infrared (FTIR) spectroscopy, and atomic force microscopy (AFM). The increasing of the β-sheet structure content at the end of protein fibrillation probed by FTIR occurs simultaneously with a major solvent exposure of tryptophan (Trp) and tyrosine (Tyr) residues of HEWL and insulin, respectively, as revealed by UVRR and intrinsic fluorescence spectroscopy. However, because the latter technique is successfully used when proteins naturally contain Trp residues, it shows poor performances in the case of insulin, and the information regarding its tertiary structure is exclusively provided by UVRR spectroscopy. The presence of an increased concentration of resveratrol induces mild changes in the secondary structure of both protein fibrils while remodeling HEWL fibril length and promoting the formation of amorphous aggregates in the case of insulin. Although the intrinsic fluorescence spectra of proteins are hidden by resveratrol signal, UVRR Trp and Tyr bands are resonantly enhanced, showing a good sensitivity to the presence of resveratrol and marking a modification in the noncovalent interactions in which they are involved. Our findings demonstrate that UVRR is successfully employed in the study of aggregation-prone proteins and of their interaction with ligands, especially in the case of Trp-lacking proteins., (Copyright © 2021 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Monoolein-based cubic and hexagonal mesophases were investigated as matrices for insulin loading, at low pH, as a function of temperature and in the presence of increasing amounts of oleic acid, as a structural stabilizer for the hexagonal phase. Synchrotron small angle X-ray diffraction, rheological measurements, and attenuated total reflection-Fourier transform infrared spectroscopy were used to study the effects of insulin loading on the lipid mesophases and of the effect of protein confinement in the 2D- and 3D-lipid matrix water channels on its stability and unfolding behavior. We found that insulin encapsulation has only little effects both on the mesophase structures and on the viscoelastic properties of lipid systems, whereas protein confinement affects the response of the secondary structure of insulin to thermal changes in a different manner according to the specific mesophase: in the cubic structure, the unfolding toward an unordered structure is favored, while the prevalence of parallel β-sheets, and nuclei for fibril formation, is observed in hexagonal structures.

In the present study, the cytotoxic effects of 5-azacytidine on primary Oral Squamous Cell Carcinoma cells (OSCCs) from human biopsies, and on Cancer Stem Cells (CSCs) from the same samples, were investigated by an in vitro Fourier Transform InfraRed Microscospectroscopy (FTIRM) approach coupled with multivariate analysis. OSCC is an aggressive tumoral lesion of the epithelium, accounting for ~90% of all oral cancers. It is usually diagnosed in advanced stages, and this causes a poor prognosis with low success rates of surgical, as well as radiation and chemotherapy treatments. OSCC is frequently characterised by recurrence after chemotherapy and by the development of a refractoriness to some employed drugs, which is probably ascribable to the presence of CSCs niches, responsible for cancer growth, chemoresistance and metastasis. The spectral information from FTIRM was correlated with the outcomes of cytotoxicity tests and image-based cytometry, and specific spectral signatures attributable to 5-azacytidine treatment were identified, allowing us to hypothesise the demethylation of DNA and, hence, an increase in the transcriptional activity, together with a conformational transition of DNA, and a triggering of cell death by an apoptosis mechanism. Moreover, a different mechanism of action between OSSC and CSC cells was highlighted, probably due to possible differences between OSCCs and CSCs response.

To understand the fate of plastic in oceans and the interaction with marine organisms, we investigated the incorporation of (bio)polymers and microplastics in selected benthic foraminiferal species by applying FTIR (Fourier Transform Infrared) microscopy. This experimental methodology has been applied to cultured benthic foraminifera Rosalina globularis, and to in situ foraminifera collected in a plastic remain found buried into superficial sediment in the Mediterranean seafloor, Rosalina bradyi, Textularia bocki and Cibicidoides lobatulus. In vitro foraminifera were treated with bis-(2-ethylhexyl) phthalate (DEHP) molecule to explore its internalization in the cytoplasm. Benthic foraminifera are marine microbial eukaryotes, sediment-dwelling, commonly short-lived and with reproductive cycles which play a central role in global biogeochemical cycles of inorganic and organic compounds. Despite the recent advances and investigations into the occurrence, distribution, and abundance of plastics, including microplastics, in marine environments, there remain relevant knowledge gaps, particularly on their effects on the benthic protists. No study, to our knowledge, has documented the molecular scale effect of plastics on foraminifera. Our analyses revealed three possible ways through which plastic-related molecules and plastic debris can enter a biogeochemical cycle and may affect the ecosystems: 1) foraminifera in situ can grow on plastic remains, namely C. lobatulus, R. bradyi and T. bocki, showing signals of oxidative stress and protein aggregation in comparison with R. globularis cultured in negative control; 2) DEHP can be incorporated in the cytoplasm of calcareous foraminifera, as observed in R. globularis; 3) microplastic debris, identified as epoxy resin, can be found in the cytoplasm and the agglutinated shell of T. bocki. We hypothesize that plastic waste and their associated additives may produce modifications related to the biomineralization process in foraminifera. This effect would be added to those induced by ocean acidification with negative consequences on the foraminiferal biogenic carbon (C) storage capacity., 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 Ltd. All rights reserved.)

Surface-Enhanced Infrared Absorption (SEIRA) has been proposed as a valuable tool for protein binding studies, but its performances have been often proven on model proteins undergoing severe secondary structure rearrangements, while ligand binding only marginally involves the protein backbone in the vast majority of the biologically relevant cases. In this study we demonstrate the potential of SEIRA microscopy for highlighting the very subtle secondary structure modifications associated with the binding of Lapatinib, a tyrosine kinase inhibitor (TKI), to epidermal growth factor receptor (EGFR), a well-known driver of tumorigenesis in pathological settings such as lung, breast and brain cancers. By boosting the performances of Mid-IR plasmonic devices based on nanoantennas cross-geometry, accustoming the protein purification protocols, carefully tuning the protein anchoring methodology and optimizing the data analysis, we were able to detect EGFR secondary structure modification associated with few amino acids. A nano-patterned platform with this kind of sensitivity bridges biophysical and structural characterization methods, thus opening new possibilities in studying of proteins of biomedical interest, particularly for drug-screening purposes.

The growing demand for innovative means in biomedical, therapeutic and diagnostic sciences has led to the development of nanomedicine. In this context, naturally occurring tubular nanostructures composed of rolled sheets of alumino-silicates, known as halloysite nanotubes, have found wide application. Halloysite nanotubes indeed have surface properties that favor the selective loading of biomolecules. Here, we present the first, to our knowledge, structural study of DNA-decorated halloysite nanotubes, carried out with nanometric spatially-resolved infrared spectroscopy. Single nanotube absorption measurements indicate a partial covering of halloysite by DNA molecules, which show significant structural modifications taking place upon loading. The present study highlights the constraints for the use of nanostructured clays as DNA carriers and demonstrates the power of super-resolved infrared spectroscopy as an effective and versatile tool for the evaluation of immobilization processes in the context of drug delivery and gene transfer.

A multi-instrumental approach combining highly sensitive Synchrotron Radiation-based techniques was used to provide information on the real composition of a dry black ink powder found in a bronze inkwell of the first century AD. The presence of Pb, Cu and Fe in the powder, revealed by XRF and ICP-OES data, leads to raise several hypotheses on their origin. The inkpot and its lid were also investigated by Hand-Held XRF, revealing a bronze alloy (Cu-Sn) with a certain amount of Fe and Pb. The lid was found to be particularly enriched in lead. XRPD, XAS and FTIR measurements showed a substantial presence of silicates and common clay minerals in the ink along with cerussite and malachite, Pb and Cu bearing-carbonates, respectively. These evidences support the hypothesis of an important contamination of the ink sample by the burial environment (soil) and the presence of degradation products of the bronze inkpot. The combined use of IR, Raman, and GC-MS evidenced that the black ink is mainly composed of amorphous carbon deriving from the combustion of organic material mixed with a natural binding agent, Arabic gum.

A micro-sample detached from a historical bowed string instrument represents a valuable record of the materials used by the great Masters of violin-making art. It allows researchers to collect a wealth of information and to disclose - at least partially - their procedures for finishing and varnishing. In the present work, a set of four cross-sectioned micro-samples - collected from well-preserved bowed string instruments made by Antonio Stradivari and Lorenzo Storioni - are investigated by Synchrotron Radiation (SR) FTIR micro-spectroscopy in reflection mode. SR-FTIR spectra are discussed both as point analysis and as univariate and multivariate chemical maps. The same cross-sections are also investigated by optical microscopy under UV light and SEM-EDX. Moreover, data obtained directly from the musical instruments by a non-invasive approach employing a portable reflection FTIR spectrometer are also considered. FTIR investigation of the cross-sections is a challenging task for such brittle and complex layered micro-samples. Nevertheless, the high intensity of the analytical SR beam used in reflection geometry allowed us to obtain informative FTIR spectra and to fully preserve the integrity of the samples. Both the non-invasive and the micro-invasive reflection FTIR approaches can reveal the materials spread on the wood surface to finish the musical instruments. The fingerprint of Lorenzo Storioni's production around 1790 emerged from the study of the cross-sectioned samples, definitely different from the technique of Stradivari., 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 © 2020 Elsevier B.V. All rights reserved.)

Oral Squamous Cells Carcinoma (OSCC) is characterised by the risk of recurrence and the onset of a refractoriness response to chemotherapy drugs. These phenomena have been recently related to a subpopulation of Cancer Stem Cells (CSCs), which have either an innate or acquired drug resistance, triggered by chemotherapy treatments. In this light, to precisely target chemotherapy regimens, it is essential to improve knowledge on CSCs, with a particular focus on their molecular features. In this work, a subpopulation of CSCs, isolated by tumour sphere formation from primary OSCC cells, were treated with cisplatin for 16, 24 and 48 hours and analysed by infrared absorption and Raman microspectroscopies. CSC spectral data were compared with those obtained in previous work, for primary OSCC cells treated under the same conditions. Routine viability/apoptosis cell-based assays evidenced in CSCs and primary OSCCs, a similar degree of sensitivity to the drug at 24 hours, while a reversion of the conventional monotonic time response exhibited by OSCCs was shown by CSCs at 48 hours. This peculiar time response was also supported by the analysis of IR and Raman data, which pinpointed alterations in the lipid composition and DNA conformation in CSCs. The results obtained suggest that CSCs, although sharing with OSCC cells a similar sensitivity to cisplatin, display the onset of a mechanism of chemoresistance and enrichment of resistant CSCs as a result of drug treatment, shedding new light on the severe issue of refractoriness of some patients to chemotherapy conventionally used for OSCC.

Radiation damage upon soft X-ray exposure is an important issue to be considered in soft X-ray microscopy. The work presented here is part of a more extended study on the topic and focuses on the effects of soft X-rays on paraffin, a common embedding medium for soft-tissues, and on ultralene and Si 3 N 4 windows as sample supports. Our studies suggest that the sample environment indeed plays an important role in the radiation damage process and therefore should be carefully taken into account for the analysis and interpretation of new data. The radiation damage effects were followed over time using a combination of Fourier transform infrared (FTIR) microspectroscopy and X-ray fluorescence (XRF), and it was demonstrated that, for higher doses, an oxidation of both embedding medium and ultralene substrate takes place after the irradiated sample is exposed to air. This oxidation is reflected in a clear increase of C=O and O-H infrared bands and on the XRF oxygen maps, correlated with a decrease of the aliphatic infrared signal. The results also show that the oxidation process may affect quantitative evaluation of light element concentrations.

Cigarette butts are the most common form of litter in the world and their environmental impact is related to both persistence and potential toxic effects for chemical composition. The objective of this study was to assess the acute toxicity (LC 50 -48 h) of human-smoked cigarette butts leachate on 3 cultured genera of benthic foraminifera: the calcareous perforate Rosalina globularis, the calcareous imperforate Quinqueloculina spp., and the agglutinated Textularia agglutinans. The specimens were exposed to 16, 8, 4, 2, and 1 cigarette butts/L concentrations that prove to be acutely toxic to all taxa. Starting from 4 cigarette butts/L, both calcareous genera showed shell decalcification, and death of almost all the individuals, except for the more resistant agglutinated species. These results suggest the potential harmfulness of cigarette butts leachate related to pH reduction and release of toxic substances, in particular nicotine, which leads to physiology alteration and in many cases cellular death., (Copyright © 2020. Published by Elsevier Ltd.)

Cytosine plays a preeminent role in DNA methylation, an epigenetic mechanism that regulates gene expression, the misregulation of which can lead to severe diseases. Several methods are nowadays employed for assessing the global DNA methylation levels, but none of them combines simplicity, high sensitivity, and low operating costs to be translated into clinical applications. Ultraviolet (UV) resonant Raman measurements at excitation wavelengths of 272 nm, 260 nm, 250 nm, and 228 nm have been carried out on isolated deoxynucleoside triphosphates (dNTPs), on a dNTP mixture as well as on genomic DNA (gDNA) samples, commercial from salmon sperm and non-commercial from B16 murine melanoma cell line. The 228 nm excitation wavelength was identified as the most suitable energy for enhancing cytosine signals over the other DNA bases. The UV Raman measurements performed at this excitation wavelength on hyper-methylated and hypo-methylated DNA from Jurkat leukemic T-cell line have revealed significant spectral differences with respect to gDNA isolated from salmon sperm and mouse melanoma B16 cells. This demonstrates how the proper choice of the excitation wavelength, combined with optimized extraction protocols, makes UV Raman spectroscopy a suitable technique for highlighting the chemical modifications undergone by cytosine nucleotides in gDNA upon hyper- and hypo-methylation events., (© 2020 Wiley-VCH GmbH.)