Microplastics ingestion and endocrine disrupting chemicals (EDCs) by breeding seabirds in the east tropical Atlantic: Associations with trophic and foraging proxies (δ15N and δ13C).

In this study we found that endocrine disrupting chemicals (EDCs) were omnipresent in a tropical seabird community comprising diverse ecological guilds and distinct foraging and trophic preferences. Because EDCs tend to bioaccumulate within the food web and microplastics can absorb and release harmful chemical compounds, our findings draw attention to the potential threats to wildlife. Thus, the goal of this study was to investigate the role of plastic ingestion, trophic and foraging patterns (δ15N and δ13C) of five tropical seabird species breeding in sympatry, on the exposure to EDCs, namely Polybrominated diphenyl ethers (PBDEs), methoxylated polybrominated diphenyl ethers (MeO-PBDEs) and personal care products (PCPs, e.g., musk fragrances and UV-filters). Results indicated that microplastics occurrence and EDCs detection frequency varied among species. Microplastics occurrence was higher in species with dual and coastal foraging strategies. Preen oil had higher levels of MeO-PBDEs and PCPs, while serum had higher levels of PBDEs. In brown boobies, the correlation between microplastics and ∑PBDEs levels was significant, suggesting that microplastics ingestion is a key PBDEs route. Trophic position (δ15N) plays a key role in PBDEs accumulation, particularly in Bulwer's petrel, which occupies a high trophic position and had more specialized feeding ecology than the other species. MeO-PBDEs were linked to foraging habitat (δ13C), although the link to foraging locations deserves further investigation. Overall, our findings not only fill key gaps in our understanding of seabirds' exposure to microplastics and EDCs, but also provide an essential baseline for future research and monitoring efforts. These findings have broader implications for the marine wildlife conservation and pollution management in sensitive environments, such as the tropical regions off West Africa.

Anthropogenic debris ingestion in a tropical seabird community: Insights from taxonomy and foraging distribution.

Oceans have been considered as an unlimited supply of goods and services, but resource extraction and waste disposal became ubiquitous and have been damaging the health of marine ecosystems. Finding suitable sentinel species of the human impacts on the oceans is thus imperative, since they may work as early warnings of disruptive situations. In this study, we investigated how taxonomy and foraging distribution influenced the occurrence of anthropogenic debris among five seabird species inhabiting the tropical Atlantic region. Occurrence of anthropogenic debris was assessed using faeces of breeding individuals as a proxy of ingestion. A total of 268 particles were extracted from all samples. The categories "fragments" and "fibres", as well as the colour "blue", were the most prevalent characteristics across species. There was a high diversity of polymers from cellulosic particles to synthetic plastics (Anthropogenic Cellulosic 26.9 %; Polyester 7.7 %; Varnish 5.8 %; Polypropylene 1.9 %). Species with a more coastal foraging strategy exhibited higher occurrence and number of anthropogenic debris when compared to species foraging comparably more in pelagic areas. This suggests that anthropogenic debris are more prevalent in coastal foraging areas, where human activities occur in higher number and frequency (e.g., fisheries) and sources of freshwater input from inland are at close distance. These results provide more evidence to the growing perception on the ubiquity and diversity of anthropogenic debris in the marine environment, and further support the usefulness of using seabirds as bio-indicators of anthropogenic pollution in both neritic and oceanic regions.

Differentiating present-day from ancient bones by vibrational spectroscopy upon acetic acid treatment.

Acetic acid treatment for an accurate differentiation between ancient and recent human bones was assessed using Raman and FTIR-ATR spectroscopies. Each set of skeletal samples was analysed by these techniques, prior and after chemical washing, in order to determine the variations in bone´s chemical composition and crystallinity. Bone samples were collected from several independent sources: recent bones burned under controlled experimental conditions or cremated, and archaeological (XVII century and Iron Age). The effect of acetic acid, expected to impact mostly on carbonates, was clearly evidenced in the spectra of all samples, particularly in FTIR-ATR, mainly through the bands typical of A- and B-carbonates. Furthermore, as seen for crematoria and archaeological samples, acetic acid was found to remove contaminants such as calcium hydroxide. Overall, acetic acid treatment can be an effective method for removing carbonates (exogenous but possibly also endogenous) and external contaminants from bone. However, these effects are dependent on the skeletal conditions (e.g. post-mortem interval and burning settings). In addition, this chemical washing was shown to be insufficient for an unequivocal discrimination between recent and archaeological skeletal remains. Based on the measured IR indexes, only cremated bones could be clearly distinguished.

Water dynamics in human cancer and non-cancer tissues.

Normal-to-malignant transformation is a poorly understood process associated with cellular biomechanical properties. These are strongly dependent on the dynamical behaviour of water, known to play a fundamental role in normal cellular activity and in the maintenance of the three-dimensional architecture of the tissue and the functional state of biopolymers. In this study, quasi-elastic neutron scattering was used to probe the dynamical behaviour of water in human cancer specimens and their respective surrounding normal tissue from breast and tongue, as an innovative approach for identifying particular features of malignancy. This methodology has been successfully used by the authors in human cells and was the first study of human tissues by neutron scattering techniques. A larger flexibility was observed for breast versus tongue tissues. Additionally, different dynamics were found for malignant and non-malignant specimens, depending on the tissue: higher plasticity for breast invasive cancer versus the normal, and an opposite effect for tongue. The data were interpreted in the light of two different water populations within the samples: one displaying bulk-like dynamics (extracellular and intracellular/cytoplasmic) and another with constrained flexibility (extracellular/interstitial and intracellular/hydration layers).

The impact of moderate heating on human bones: an infrared and neutron spectroscopy study.

This study aims to analyse human bones exposed to low/medium temperatures (200-650°C) under experimentally controlled conditions, both oxidizing and reducing, using complementary optical and neutron vibrational spectroscopy techniques. Clear differences were observed between the aerobically and anaerobically heated bones. The organic constituents disappeared at lower temperatures for the former (ca 300°C), while they lingered for higher temperatures in anaerobic environments (ca 450-550°C). Unsaturated non-graphitizing carbon species (chars) were detected mainly for anaerobically heated samples, and cyanamide formation occurred only at 650°C in reducing settings. Overall, the main changes were observed from 300 to 400°C in anaerobic conditions and from 450 to 500°C in aerobic environments. The present results enabled the identification of specific spectroscopic biomarkers of the effect of moderate temperatures (less than or equal to 650°C) on human bone, thus contributing to a better characterization of forensic and archaeological skeletal remains subject to heating under distinct environmental settings. In particular, these data may provide information regarding cannibalism or ancient bone boiling and defleshing rituals.

Profiling of human burned bones: oxidising versus reducing conditions.

Complementary optical and neutron-based vibrational spectroscopy techniques (Infrared, Raman and inelastic neutron scattering) were applied to the study of human bones (femur and humerus) burned simultaneously under either aerobic or anaerobic conditions, in a wide range of temperatures (400 to 1000 °C). This is the first INS study of human skeletal remains heated in an oxygen-deprived atmosphere. Clear differences were observed between both types of samples, namely the absence of hydroxyapatite's OH vibrational bands in bone burned anaerobically (in unsealed containers), coupled to the presence of cyanamide (NCNH2) and portlandite (Ca(OH)2) in these reductive conditions. These results are expected to allow a better understanding of the heat effect on bone´s constituents in distinct environmental settings, thus contributing for an accurate characterisation of both forensic and archaeological human skeletal remains found in distinct scenarios regarding oxygen availability.

Role of intracellular water in the normal-to-cancer transition in human cells-insights from quasi-elastic neutron scattering.

The transition from normal to malignant state in human cells is still a poorly understood process. Changes in the dynamical activity of intracellular water between healthy and cancerous human cells were probed as an innovative approach for unveiling particular features of malignancy and identifying specific reporters of cancer. Androgen-unresponsive prostate and triple-negative breast carcinomas were studied as well as osteosarcoma, using the technique of quasi-elastic neutron scattering. The cancerous cells showed a considerably higher plasticity relative to their healthy counterparts, this being more significant for the mammary adenocarcinoma. Also, the data evidence that the prostate cancer cells display the highest plasticity when compared to triple-negative mammary cancer and osteosarcoma, the latter being remarkably less flexible. Furthermore, the results suggest differences between the flexibility of different types of intracellular water molecules in normal and cancerous cells, as well as the number of molecules involved in the different modes of motion. The dynamics of hydration water molecules remain virtually unaffected when going from healthy to cancer cells, while cytoplasmic water (particularly the rotational motions) undergoes significant changes upon normal-to-cancer transition. The results obtained along this study can potentially help to understand the variations in cellular dynamics underlying carcinogenesis and tumor metastasis, with an emphasis on intracellular water.

First analysis of ancient burned human skeletal remains probed by neutron and optical vibrational spectroscopy.

Burned skeletal remains are abundant in archaeological and paleontological sites, the result of fire or of ancient funerary practices. In the burning process, the bone matrix suffers structural and dimensional changes that interfere with the reliability of available osteometric methods. Recent studies showed that these macroscopic changes are accompanied by microscopic variations are reflected in vibrational spectra. An innovative integrated approach to the study of archaeological combusted skeletal remains is reported here, where the application of complementary vibrational spectroscopic techniques-INS (inelastic neutron scattering), FTIR (Fourier transform infrared), and micro-Raman-enables access to the complete vibrational profile and constitutes the first application of neutron spectroscopy to ancient bones. Comparison with data from modern human bones that were subjected to controlled burning allowed identification of specific heating conditions. This pioneering study provides archaeologists and anthropologists with relevant information on past civilizations, including regarding funerary, burial, and cooking practices and environmental settings.

Human bone probed by neutron diffraction: the burning process.

The first neutron diffraction study of human burned bone is reported, aiming at a comprehensive elucidation of the heat-induced bone diagenesis process. Chemical and crystallinity changes were probed in different types of bone (femur, humerus and tibia) upon heating to different maximum temperatures (from 400 to 1000 °C, under aerobic conditions). Fourier transform infrared spectroscopy has provided valuable complementary information. Noticeable crystallographic and domain size variations were detected, mainly between 700 and 900 °C, the high temperature interval (>700 °C) corresponding to an organized, highly symmetric inorganic bone matrix, virtually devoid of carbonates and organic constituents, while the lower range (<700 °C) revealed a considerably lower crystallinity associated with the presence of carbonates, lipids and collagen. This work contributes to a better understanding of heat-induced changes in bone and is therefore relevant for archaeology, biomaterials and forensic science.

Heat-induced Bone Diagenesis Probed by Vibrational Spectroscopy.

Complementary vibrational spectroscopic techniques - infrared, Raman and inelastic neutron scattering (INS) - were applied to the study of human bone burned under controlled conditions (400 to 1000 °C). This is an innovative way of tackling bone diagenesis upon burning, aiming at a quantitative evaluation of heat-induced dimensional changes allowing a reliable estimation of pre-burning skeletal dimensions. INS results allowed the concomitant observation of the hydroxyl libration (OHlibration), hydroxyl stretching (ν(OH)) and (OHlibration + ν(OH)) combination modes, leading to an unambiguous assignment of these INS features to bioapatite and confirming hydroxylation of bone's inorganic matrix. The OHlib, ν(OH) and ν4(PO43-) bands were identified as spectral biomarkers, which displayed clear quantitative relationships with temperature revealing heat-induced changes in bone's H-bonding pattern during the burning process. These results will enable the routine use of FTIR-ATR (Fourier Transform Infrared-Attenuated Total Reflectance) for the analysis of burned skeletal remains, which will be of the utmost significance in forensic, bioanthropological and archaeological contexts.

Novel platinum-based anticancer drug: a complete vibrational study.

The introduction of cisplatin to oncology, in the 1970s, marked the onset of the search for novel and improved metal-based anticancer drugs. Polynuclear PtII and PdII complexes with linear alkylamines as bridging ligands are a class of potential antineoplastic agents that have shown promising cytotoxicity against low-prognosis human cancers, such as metastatic breast adenocarcinoma and osteosarcoma. The present study reports an analysis of [µ-N,N'-bis(3-aminopropyl)butane-1,4-diamine-κ4N,N':N'',N''']bis[dichloridoplatinum(II)], [Pt2Cl4(C10H26N4)], denoted Pt2Spm (Spm is spermine), by vibrational spectroscopy coupled to theoretical calculations. Within the latter, the Density Functional Theory (DFT - mPW1PW/6-31G*) and Effective Core Potential (ECP - LANL2DZ) approaches were used, in order to ensure the most accurate representation of the molecule and achieve a maximum agreement with the experimental data. The solid-state geometry of Pt2Spm corresponds to Ci symmetry, displaying 132 vibrational modes. A complete assignment of the experimental vibrational profile of the system was attained through the combined application of complementary Raman, FT-IR and Inelastic Neutron Scattering (INS) techniques. INS allowed an unequivocal identification of the CH2 and NH2 rocking modes, not clearly detected by the optical techniques, while Raman measurements led to a clear discrimination of the Pt-N stretching frequencies from the two distinct Pt-N moieties within the chelate. The metal-to-metal distances calculated for the molecule under study were found to allow the establishment of effective inter- and intrastrand crosslinks with DNA. These results will hopefully help to clarify the mode of action of the compound, at the molecular level, contributing to the development of improved cisplatin-like chemotherapeutic drugs having a higher efficacy and specificity coupled to lower acquired resistance and deleterious side effects.

Crystal clear: Vibrational spectroscopy reveals intrabone, intraskeleton, and interskeleton variation in human bones.

OBJECTIVES: Vibrational spectroscopy is a valuable tool for the study of burned skeletal remains. Nonetheless, most investigations have been focused on a limited number of samples as well as on faunal bones rather than human bones. Conclusions based on those investigations may lack representativeness, namely about the intrabone, intra- and interskeleton variability of several chemometric indices. We aimed to investigate this issue on a large sample of human bones. MATERIAL AND METHODS: Powder samples were collected from 168 bones from four human skeletons. The sampling targeted 47 long bones, 72 short bones, and 49 tarsal bones as well as different bone regions in a total of 638 powder samples. Bones were experimentally burned in an electric muffle furnace for two hours to maximum temperatures ranging from 400°C to 1000°C. Another 623 burned samples were then collected totaling 1261 samples subjected to FTIR-ATR analysis. The CI, BPI, C/C, and OH/P indices were calculated. RESULTS: An important intrabone, intra- and interskeleton variation was observed, especially for the BPI. The CI, C/C, and OH/P indices revealed much less variation so site-specific sampling may not be as critical in these cases. Clear differences between our results and those from previous investigations were observed, namely on the temperature increment evolution of the CI and C/C indices. DISCUSSION: The relatively large heterogeneity, especially at the intrabone level, is possibly the consequence of microstructural bone differences. The dissimilarities observed between our investigation and other published studies are probably due to the fact that the samples used here came from human rather than faunal bones. Also, our samples were buried previously to the experimental burning so this may also partly explain our contrasting results, since previous research was mostly performed on fresh bone. Future inferences based on vibrational spectroscopy analyses should take into account the possible effect of all these sources.

Biomaterials from human bone - probing organic fraction removal by chemical and enzymatic methods.

Two different deproteination and defatting processes of human bone were investigated, by combined infrared and neutron techniques: a previously reported hydrazine extraction and a newly developed multi-enzymatic treatment. Complementary Fourier transform infrared total attenuated reflectance and inelastic neutron scattering spectroscopies were applied, allowing access to all vibrational modes of the samples. The effectiveness of the different experimental protocols for removing the organic constituents of bone (lipids and protein) was probed, as well as their effect on bone's structural and crystallinity features. The results thus gathered are expected to have an impact on bioanthropological, archaeological and medical sciences, namely regarding the development of novel biocompatible materials for orthopaedic xenografts.

Intracellular water - an overlooked drug target? Cisplatin impact in cancer cells probed by neutrons.

The first neutron scattering study on human nucleated cells is reported, addressing the subject of solvent-slaving to a drug by probing intracellular water upon drug exposure. Inelastic and quasi-elastic neutron scattering spectroscopy with isotope labelling was applied for monitoring interfacial water response to the anticancer drug cisplatin, in the low prognosis human metastatic breast cancer cells MDA-MB-231. Optical vibrational data were also obtained for lyophilised cells. Concentration-dependent dynamical changes evidencing a progressive mobility reduction were unveiled between untreated and cisplatin-exposed samples, concurrent with variations in the native organisation of water molecules within the intracellular medium as a consequence of drug action. The results thus obtained yielded a clear picture of the intracellular water response to cisplatin and constitute the first reported experimental proof of a drug impact on the cytomatrix by neutron techniques. This is an innovative way of tackling a drug's pharmacodynamics, searching for alternative targets of drug action.

Chemotherapeutic response to cisplatin-like drugs in human breast cancer cells probed by vibrational microspectroscopy.

Studies of drug-cell interactions in cancer model systems are essential in the preclinical stage of rational drug design, which relies on a thorough understanding of the mechanisms underlying cytotoxic activity and biological effects, at a molecular level. This study aimed at applying complementary vibrational spectroscopy methods to evaluate the cellular impact of two Pt(ii) and Pd(ii) dinuclear chelates with spermine (Pt2Spm and Pd2Spm), using cisplatin (cis-Pt(NH3)2Cl2) as a reference compound. Their effects on cellular metabolism were monitored in a human triple-negative metastatic breast cancer cell line (MDA-MB-231) by Raman and synchrotron-radiation infrared microspectroscopies, for different drug concentrations (2-8 µM) at 48 h exposure. Multivariate data analysis was applied (unsupervised PCA), unveiling drug- and concentration-dependent effects: apart from discrimination between control and drug-treated cells, a clear separation was obtained for the different agents studied - mononuclear vs. polynuclear, and Pt(ii) vs. Pd(ii). Spectral biomarkers of drug action were identified, as well as the cellular response to the chemotherapeutic insult. The main effect of the tested compounds was found to be on DNA, lipids and proteins, the Pd(ii) agent having a more significant impact on proteins while its Pt(ii) homologue affected the cellular lipid content at lower concentrations, which suggests the occurrence of distinct and unconventional pathways of cytotoxicity for these dinuclear polyamine complexes. Raman and FTIR microspectroscopies were confirmed as powerful non-invasive techniques to obtain unique spectral signatures of the biochemical impact and physiological reaction of cells to anticancer agents.

Conformational study and reassessment of the vibrational assignments for Norspermidine.

The present study presents and discusses the conformational preferences of Norspermidine (NSpd). The effects of varying the dielectric constant on the conformational preferences are discussed, with a view to infer which conformation will correspond to the most stable in the pure condensed liquid phase. Within the same context, a set of NSpd-NH3 molecular adducts were simulated in order to determine the relevance of intermolecular hydrogen bonding on the overall stability and relative positioning of the respective vibrational frequencies. The calculations presently performed allowed a reassessment of the vibrational assignments for NSpd. A full assignment of the NSpd vibrational spectra is presented, with special emphasis being given to the vibrational modes that proved to be most affected by hydrogen bonding. The various inconsistencies of a prior study found in the literature were identified and rectified.

Determination of preferred conformations of ibuprofen in chloroform by 2D NOE spectroscopy.

Solution of an anti-inflammatory drug ibuprofen ((RS)-2-(4-isobutylphenyl) propionic acid) in chloroform was studied by nuclear magnetic resonance spectroscopy. A set of 2D NOESY spectra was analyzed in order to obtain atom-atom distances. Since ibuprofen is known to exist as an ensemble of different conformations, these distances are averaged over the ensemble. To compare experimental and calculated distances, three models of averaging were concerned. Our data allowed to determine the dominant conformers of ibuprofen dissolved in chloroform. The population of conformers in the saturated solution leads to a certain crystal morphology formed within the nucleation process. Observed and calculated (13)C chemical shifts (at the DFT/B3LYP/6-311+G(2d,p) level) were in good agreement.

Gamma scintigraphy in the analysis of ketoprofen behaviour from matrix tablets.

The aim of this work was to study in vitro and in vivo the behaviour of matrix tablets (quick and extended release) containing ketoprofen (KTP) as a model drug and cellulose ether polymers, using gamma scintigraphy. The matrix tablets were prepared by the direct compression method and labelled by incorporating a drop of technetium ((99m)Tc). It was spectrophotometricaly confirmed that the radioisotope inclusion did not modify the kinetics of KTP release. In vitro studies were carried out for the tablets using the paddle method of the USP 35/NF30. The images were processed by defining regions of interest over the tablet (99m)Tc and the percentage of remaining activity/time curves were generated for both formulations. In vitro gamma scintigraphy studies showed significant differences (p<0.05) between both formulations. Identical results were obtained from the in vivo studies. In vivo tests were performed on five healthy volunteers. The scintigraphy images were acquired during 2.5 and 7.5h for quick and extended release formulations, respectively. The position of the extended release formulation tablet along the gastrointestinal tract (GIT) was assessed. The described results demonstrate the in vitro/in vivo correlation for the drug release profile and exhibit the importance of gamma scintigraphy for the drug location through the GIT.

A conformational study of hydroxyflavones by vibrational spectroscopy coupled to DFT calculations.

The conformational preferences of a series of hydroxyflavones were studied by Raman and FTIR spectroscopies, coupled to Density Functional Theory calculations. Special attention was paid to the effect of hydroxyl substitution, due to its importance on the biological activity of these compounds. Their conformational preferences were found to be determined mainly by the orientation of the hydroxylic groups at C(7) and within the catechol moiety, leading to the occurrence of distinct conformers in the solid state. A complete assignment of the experimental spectra was carried out for these molecules, in the light of their most stable conformers and the corresponding predicted vibrational pattern.

In vitro release of ketoprofen from hydrophilic matrix tablets containing cellulose polymer mixtures.

The effect of cellulose ether polymer mixtures, containing both hydroxypropylcellulose (HPC) and hydroxypropylmethylcellulose (HPMC K15M or K100M), on ketoprofen (KTP) release from matrix tablets was investigated. In order to evaluate the compatibility between the matrix components, Raman spectroscopy, scanning electron microscopy (SEM), and X-ray powder diffraction (XRPD) experiments were performed. The results evidence the absence of significant intermolecular interactions that could eventually lead to an incompatibility between the drug and the different excipients. Formulations containing mixtures of polymers with both low and high viscosity grades were prepared by a direct compression method, by varying the polymer/polymer (w/w) ratio while keeping the drug amount incorporated in the solid dispersion constant (200 mg). The hardness values of different matrices were found within the range 113.8 to 154.9 N. HPLC analysis showed a drug content recovery between 99.3 and 102.1%, indicating that no KTP degradation occurred during the preparation process. All formulations attained a high hydration degree after the first hour, which is essential to allow the gel layer formation prior to tablet dissolution. Independent-model dissolution parameters such as t(10%) and t(50%) dissolution times, dissolution efficiency (DE), mean dissolution time (MDT), and area under curve (AUC) were calculated for all formulations. Zero-order, first-order, Higuchi, and Korsmeyer-Peppas kinetic models were employed to interpret the dissolution profiles: a predominantly Fickian diffusion release mechanism was obtained - with Korsmeyer-Peppas exponent values ranging from 0.216 to 0.555. The incorporation of HPC was thus found to play an essential role as a release modifier from HPMC containing tablets.