First In Vivo Insights on the Effects of Tempol-Methoxycinnamate, a New UV Filter, as Alternative to Octyl Methoxycinnamate, on Zebrafish Early Development.

The demand for organic UV filters as active components in sunscreen products has rapidly risen over the last century, as people have gradually realized the hazards of overexposure to UV radiation. Their extensive usage has resulted in their ubiquitous presence in different aquatic matrices, representing a potential threat to living organisms. In this context, the need to replace classic UV filters such as octyl methoxycinnamate (OMC), one of the most popular UV filters reported to be a potential pollutant of aquatic ecosystems, with more environmentally friendly ones has emerged. In this study, using zebrafish, the first in vivo results regarding the effect of exposure to tempol-methoxycinnamate (TMC), a derivative of OMC, are reported. A comparative study between TMC and OMC was performed, analyzing embryos exposed to similar TMC and OMC concentrations, focusing on morphological and molecular changes. While both compounds seemed not to affect hatching and embryogenesis, OMC exposure caused an increase in endoplasmic reticulum (ER) stress response genes, according to increased eif2ak3, ddit3, nrf2, and nkap mRNA levels and in oxidative stress genes, as observed from modulation of the sod1, sod2, gpr, and trx mRNA levels. On the contrary, exposure to TMC led to reduced toxicity, probably due to the presence of the nitroxide group in the compound's molecular structure responsible for antioxidant activity. In addition, both UV filters were docked with estrogen and androgen receptors where they acted differently, in agreement with the molecular analysis that showed a hormone-like activity for OMC but not for TMC. Overall, the results indicate the suitability of TMC as an alternative, environmentally safer UV filter.

Characterisation of Bee Pollen from the Marche Region (Italy) According to the Botanical and Geographical Origin with Analysis of Antioxidant Activity and Colour, Using a Chemometric Approach.

Attempts have often been made to isolate and characterise monofloral pollens to correlate nutritional with botanical properties. Nevertheless, pollen harvested in a particular area that can have a high biodiversity could have healthier properties. In addition, the analysis of the pollen's botanical composition can be important for characterising the typical flora of a specific geographical area. On this basis, various pollens collected in different locations of the Marche region (Italy) and in different harvesting periods were analyzed for botanical composition and antioxidant (total phenolic content, ABTS, DPPH and ORAC tests), granulometry and colour (CIE L*a*b*) properties to evaluate the biodiversity of pollen sources within a particular geographical area and to correlate this to the nutraceutical characteristics. Antioxidant activity results showed values generally higher than those of monofloral pollens harvested in the same areas but manually separated according to colour, shape and size. This suggests that even the floral species present in low percentages may have an influence on the nutraceutical properties of these products. The multivariate statistical elaboration of the obtained results permitted the separation of samples containing a prevalent botanical species and the grouping of all the samples into separate clusters corresponding to different areas of Marche.

Cubic and Hexagonal Mesophases for Protein Encapsulation: Structural Effects of Insulin Confinement.

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 ß-sheets, and nuclei for fibril formation, is observed in hexagonal structures.

Morphological, Physicochemical and FTIR Spectroscopic Properties of Bee Pollen Loads from Different Botanical Origin.

Bee pollen loads generally have a homogeneous and monospecific pollen content and assume a typical form and color, due to the typical bee foraging habits, thus having a typical composition related to the botanical origin. The present study aims to characterize bee pollen loads belonging to different botanical species using morphological, spectroscopic and color properties and to find relationships between these variables. IR spectra analysis allowed to have a reliable picture of the components present in the different samples; color and granulometry permits a visual identification of pollen load belonging to different species. Multivariate analysis enabled differentiation among the botanical origin of most of the bee pollen samples, grouping them according to the family and the genus and confirming the possibility to use IR and color measurements for the evaluative analysis and classification of bee pollen samples, to promote the consumption of this bee product as functional food.

Lyotropic Liquid-Crystalline Nanosystems as Drug Delivery Agents for 5-Fluorouracil: Structure and Cytotoxicity.

Lyotropic cubic liquid-crystalline systems have received increasing attention due to their unique microstructural and physicochemical properties as efficient nanocarriers for drug delivery. We report the preparation and characterization of bulk phases and cubosome dispersions of phytantriol loaded with the anticancer drug 5-fluorouracil, in neutral and anionic forms. In both cases, a Pn3m cubic phase was observed. The phytantriol phase behavior can be influenced by the addition of ionic agents, and, to this purpose, a positively charged lipid, such as N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride salt (DOTAP), was included in the studied formulations. It was found to induce a variation of the spontaneous membrane curvature of the phytantriol lipid bilayer, generating a transition from the Pn3m to the Im3m cubic phase. When 5-fluorouracil, in its anionic form (5-FUs), was encapsulated in these latter systems, a further transition to the HII hexagonal phase was observed as a consequence of the formation of a complex phytantriol/DOTAP/5-FUs. The physicochemical characterization was performed with various complementary techniques including synchrotron small-angle X-ray scattering, dynamic light scattering, and attenuated total reflection Fourier transform infrared and UV resonance Raman spectroscopies. Encapsulation of 5-fluorouracil in the corresponding nanodispersions was evaluated, and their in vitro cytotoxicity was assessed in MDA-MB-231 cell line. Phytantriol cubosomes containing 5-fluorouracil showed a higher toxicity compared with the bare drug solution, and hence they represent potential nanocarriers in the delivery of 5-fluorouracil for cancer therapy.

Consanguinity and late fertility: spatial analysis reveals positive association patterns.

The role of consanguinity on human complex traits is an important and controversial issue. In this work we focused on the Sardinian population and examined the effect of consanguineous unions on late female fertility. During the last century the island has been characterized by a high incidence of marriages between relatives, favoured by socio economic conditions and geographical isolation, and by high fertility despite a widespread tendency to delay reproduction. Through spatial analysis techniques, we explored the geographical heterogeneity of consanguinity and late fertility, and identified in Central-Eastern Sardinia a common area with an excess of both traits, where the traits are positively associated. We found that their association did not significantly affect women's fertility in the area, despite the expected negative role of both traits. Intriguingly, this critical zone corresponds well to areas reported by previous studies as being peculiar for a high frequency of centenarians and for lower risk in pregnancy outcome. The proposed approach can be generally exploited to identify target populations on which socioeconomic, biodemographic and genetic data can be collected at the individual level, and deeper analyses carried out to disentangle the determinants of complex biological traits and to investigate their association.

Influence of steeping conditions (time, temperature, and particle size) on antioxidant properties and sensory attributes of some white and green teas.

The influence of commonly used steeping times and temperatures, as well as leaf size on the antioxidant activity and sensory attributes of tea were studied. Five unblended white and green tea samples from China and Malawi, infused in hot (70 °C and 90 °C; 7 min) or cold water (room temperature: 15, 30, 60, or 120 min) either as whole leaves or as milled, were analyzed. Total phenolic and flavonoid contents as well as antioxidant power (ABTS assay) were measured. The results show that the maximum extraction efficiency occurs with cold water for 120 min and with hot water at 90 °C and that only in the case of teas from whole, large leaves, the extraction was greater in cold than in hot infusions. Moreover, tea infusions prepared from milled leaves have the greatest antioxidant activity. In the sensory evaluation of some of the tea infusions, white teas were perceived more fragrant than green ones and were judged as the most favorite by the majority of the judges, especially for the brew prepared in cold water from whole leaves; all infusions obtained from the milled leaves in fact have a more bitter and astringent taste.

Reactions of nitric oxide and nitrogen dioxide with coenzyme Q: involvement of the isoprenic chain.

The formation of a di-tert-alkyl nitroxide has been observed by Electron Spin Resonance during the exposure of coenzyme CoQ(10), in both the oxidized and reduced forms, to nitrogen dioxide (˙NO(2)) or to nitric oxide (˙NO) in the presence of oxygen. The same kind of nitroxide has been observed also with CoQ(1), CoQ(3) or with 1-phenyl-3-methyl-2-butene, chosen as model compounds. In all cases, the formation of the nitroxide may be justified only by admitting the involvement of the isoprenic chain of the coenzymes and in particular the addition of ˙NO(2) to the double bond. A mechanism which accounts for the formation of the nitroxide as well as the other compounds observed in the reactions is proposed and confirmed by a spectroscopic investigation (FT-IR, (1)H NMR, X-ray analysis) and by ESI-MS.

The influence of mannose-based esters on the mesophase behaviour of lyotropic liquid crystalline nanosystems as drug delivery vectors.

Lyotropic Liquid Crystalline (LLC) nanoparticles represent an emerging class of smart, biocompatible, and biodegradable systems for the delivery of drugs. Among these, structures with complex 3D architectures such as cubosomes are of particular interest. These are non- lamellar assemblies having hydrophobic and hydrophilic portions able to carry drugs of different nature. They can further be modulated including suitable additives to control the release of the active payload, and to promote an active targeting. Starting from monoolein (GMO) cubic phase, different concentrations of mannose-based esters were added, and the eventual structural modifications were monitored to ascertain the effects of the presence of glycolipids. Moreover, the structural properties of these nanosystems loaded with Dexamethasone (DEX), a very well-known anti-inflammatory steroid, were also studied. Experiments were carried out by synchrotron Small Angle X-ray Scattering (SAXS), Raman Microspectroscopy (RMS) and Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR) measurements. The drug delivery potential (i.e. entrapment efficiency and release properties) of the obtained nanoparticles was evaluated. Finally, in vitro cytocompatibility and anti-inflammatory activity studies of the prepared formulations were carried out. Inclusion of mannose-based surfactants up to 10 mol% influenced the structural parameters of Im3m cubic phase and swollen cubic phases were obtained with the different glycolipids with lattice parameters significantly higher than GMO. A complete cytocompatibility and an increased DEX activity were observed, thus suggesting the possibility to use GMO/glycolipids nanoparticles to formulate innovative drug delivery systems.

Spatial analysis of the aptitude to late maternity on the island of Sardinia.

This study examines local heterogeneity in the aptitude of Sardinian mothers towards late reproduction, and explores its temporal persistence and association with both post-reproductive longevity and propensity to consanguineous marriage. Data on women's fertility from 1961 and birth records for 1980-1996 from Vital Statistics were analysed by means of the following indicators: the incidence of old mothers at last childbirth, female mortality (1980-2001) at 80 years of age and over and the proportion of consanguineous marriages (1930-1969). A variable kernel-smoothing method was used to create interpretable representations of the true spatial structure of the indicators, and to highlight areas of higher than expected intensity. In particular, an area of reproductive and post-reproductive longevity was identified where the traits combine with a higher tendency to relatedness. Intriguingly, this area corresponds approximately to the geographically and historically well defined central-eastern zone, which was the refuge of Sardinians during past invasions, and overlaps the Ogliastra region, which has been widely studied for its genetic homogeneity.

Radical-scavenging activity, protective effect against lipid peroxidation and mineral contents of monofloral Cuban honeys.

Several monofloral Cuban honeys were analyzed to determine their free radical-scavenging activity and from this the total antioxidant content was estimated. The protective effect against lipid peroxidation in an in vitro model of rat liver homogenates was evaluated and, lastly, the mineral content of the honeys, which can be related to the maintenance of intracellular oxidative balance, was determined. The scavenging capacities against hydroxyl and superoxide radicals were determined using the spin-trapping technique and the hypoxanthine/xanthine oxidase assay, respectively. Lipid peroxidation was evaluated through the production of TBARS and hydroperoxides. All honeys tested showed potential antioxidant activity with Linen vine displaying the highest scavenging capacity towards the DPPH, hydroxyl and superoxide radicals, while the least efficient was Christmas vine honey. Honeys also inhibited, in a concentration-dependent mode, lipid peroxidation in rat liver homogenates, with Linen vine resulting the best while the least effective was Christmas vine honey. The ability to scavenge free radicals and protect against lipid peroxidation may contribute to the ability of certain Cuban honeys to help in preventing/reducing some inflammatory diseases in which oxidative stress is involved. A total of eight minerals were identified and quantified as follows: cadmium, chromium, copper, nickel, iron, manganese, lead, and zinc. Minerals found in higher concentrations were iron, zinc and manganese.

Nanoscale Structure of Langmuir-Blodgett Film of Bent-Core Molecules.

Bent-core mesogens (BCMs) are a class of thermotropic liquid crystals featuring several unconventional properties. However, the interpretation and technological exploitation of their unique behavior have been hampered by the difficulty of controlling their anchoring at surfaces. To tackle this issue, we report the nanoscale structural characterization of BCM films prepared using the Langmuir-Blodgett technique. Even though BCMs are quite different from typical amphiphilic molecules, we demonstrate that stable molecular films form over water, which can then be transferred onto silicon substrates. The combination of Brewster angle microscopy, atomic force microscopy, and X-ray reflectivity measurements shows that the molecules, once transferred onto a solid substrate, form a bilayer structure with a bottom layer of flat molecules and an upper layer of upright molecules. These results suggest that Langmuir-Blodgett films of BCMs can provide a useful means to control the alignment of this class of liquid crystals.

A vibrational in vitro approach to evaluate the potential of monoolein nanoparticles as isofuranodiene carrier in MDA-MB 231 breast cancer cell line: New insights from Infrared and Raman microspectroscopies.

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.

Synthesis and thermal stability of benzoxazine nitroxides.

A new class of stable nitroxides (aminoxyls) having a 1,4-benzoxazine structure were synthesized and the corresponding thermal stability tested. All derivatives were stable in the entire range of temperatures employed, except those having a benzyl or a tert-butyl group at the ß-position with respect to the aminoxyl function, which underwent radical fragmentation. Such a behavior allowed a kinetic study, carried out by means of EPR spectroscopy, to determine the corresponding rate constants and activation parameters (E(a)). Appropriate DFT calculations were performed for all nitroxides including also the thermally stable ones, in order to study the geometries of the fragmentation transition States as well as to compute the corresponding bond dissociation enthalpies (BDH), useful for further modeling purposes. The data obtained were interpreted on the basis of the relative stability of the leaving radical, according to the corresponding E(a) and BDH, whereas in the case of tert-butyl derivatives steric hindrance should play a determinant role, as evidenced by a comparison of the geometric, kinetic, and thermodynamic parameters upon the whole series.

Antioxidant Activity Level, Bioactive Compounds, Colour and Spectroscopic Analysis (UV-Vis and FT-IR) of Flavoured Drinks Made with Wine and Sour Cherries (Prunuscerasus Var. austera).

In recent years, the increase in consumer interest towards simpler and authentic lifestyles has led to an explosive growth in the production and business of typical agri-food products and, among these, of wines and its derived beverages. With the aim of promoting a typical Italian beverage, the so-called "Vino di visciole" or "Visner", listed in the national table of traditional agri-food products, the antioxidant and colour properties of fifteen samples from different provinces of the Marche region and obtained with different recipes were analysed. The "in vitro" total antioxidant activity (TAA) determined using ABTS assays, total phenolic content (TPC), total flavonoid content (TFC), total anthocyanins content (TAC), and colour (Somers assay) were measured. In addition, a spectroscopic FT-IR and UV-Vis analysis was carried out to analyse samples with multivariate techniques. The results showed that the production area, the recipe, and the type of cherries used to make the alcoholic beverage do not influence the antioxidant properties and the phytochemical contents of the samples. The multivariate treatment of the spectroscopic features (mainly UV-Vis) rather allowed the differentiation of samples with high antioxidant activity using easy and low-cost instrumental techniques that require little time and can be employed in routine analysis.

The reaction of spiro[indoline-naphthopyrans] with nitrogen oxides. Not a case of biradical double trapping.

The reactions of six differently substituted photochromic spiro[indoline-naphtopyrans] with .NO or .NO(2) under normal daylight conditions have been investigated by means of EPR spectroscopy along with those of three structurally related spiro[indoline-benzopyrans]. The spectra due to cyclic oxynitroxides originating from double trapping of biradicals by .NO were observed with the three latter derivatives, this finding being in agreement with previous results. Similar signals were also observed with the six former compounds, but in this case they were responsible for just a minor component of the spectra, the main spectral signals being due to hitherto unreported paramagnetic species that on the basis of their spectral parameters are identified as iminoxy radicals. DFT calculations at the B3LYP/6-31G* level carried out on a variety of radicals support this assignment.

3-Benzyl-3-hy-droxy-2-phenyl-3H-indole 1-oxide.

The asymmetric unit of the title compound, C(21)H(17)NO(2), contains two crystallographically independent mol-ecules of similar geometry. The indole ring systems form dihedral angles of 8.30 (5) and 9.58 (5)° with the attached phenyl rings, and 56.96 (5) and 57.68 (5)° with the aromatic rings of the respective benzyl groups. The mol-ecular conformations are stabilized by intra-molecular C-H⋯O hydrogen bonds. In the crystal structure, centrosymmetrically related pairs of mol-ecules are linked into dimers through pairs of inter-molecular O-H⋯O hydrogen bonds, generating 12-membered rings with R(2) (2)(12) motifs. The dimers are further linked into a three-dimensional network by C-H⋯O inter-actions.

Synchrotron Characterization of Hexagonal and Cubic Lipidic Phases Loaded with Azolate/Phosphane Gold(I) Compounds: A New Approach to the Uploading of Gold(I)-Based Drugs.

Gold(I) phosphane compounds have recently attracted a renewed interest as potential new protagonists in cancer therapy. A class of phosphane gold(I) complexes containing azolate ligands has been successfully tested against several cancer cell lines and, in particular, against basal-like breast (BLB) cancer, a form characterized by strongly severe diagnosis and short life lapse after classic chemotherapy. Even though the anticancer activity of gold(I) phosphane compounds is thoroughly ascertained, no study has been devoted to the possibility of their delivery in nanovectors. Herein, nonlamellar lyotropic liquid crystalline lipid nanosystems, a promising class of smart materials, have been used to encapsulate gold(I) azolate/phosphane complexes. In particular, ((triphenylphosphine)-gold(I)-(4,5-dichloroimidazolyl-1H-1yl)) (C-I) and ((triphenylphosphine)-gold(I)-(4,5-dicyanoimidazolyl-1H-1yl)) (C-II) have been encapsulated in three different lipid matrices: monoolein (GMO), phytantriol (PHYT) and dioleoyl-phosphatidylethanolamine (DOPE). An integrated experimental approach involving X-ray diffraction and UV resonant Raman (UVRR) spectroscopy, based on synchrotron light and attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, has been employed to establish the effects of drug encapsulation on the structure and phase behavior of the host mesophases. The results indicate that gold(I) complexes C-I and C-II are successfully encapsulated in the three lipid matrices as evidenced by the drug-induced phase transitions or by the changes in the mesophase lattice parameters observed in X-ray diffraction experiments and by the spectral changes occurring in UV resonant Raman spectra upon loading the lipid matrices with C-I and C-II.

Nanostructured liquid crystalline particles as delivery vectors for isofuranodiene: Characterization and in-vitro anticancer activity.

Isofuranodiene is an oxygenated sesquiterpene containing a furan ring isolated from the essential oil of Smyrnium olusatrum L. (Apiaceae) owning notable anticancer activity. Despite its biological potential, the high lipophilicity along with a relatively low stability due to Cope rearrangement giving rise to a less active compound, make the perspective of its therapeutical use unlikely. On this basis, in the present work we evaluated bulk and dispersed non lamellar liquid crystalline phases as effective delivery vectors for isofuranodiene, and capable of preserving its structure and enhancing the biological activity. Small-angle X-ray scattering, dynamic light scattering, and UV resonance Raman spectroscopy were used to characterize the nanosystems in an integrated experimental approach. Encapsulation of isofuranodiene in the lipid matrix resulted in a transition from a cubic Im3m to a reversed hexagonal phase because of the highly lipophilic character of the drug, as obtained in SAXS measurements, and in significant shifts in the components of the Raman spectrum of isofuranodiene. The anticancer activity of isofuranodiene-loaded lipidic nanoparticles was assessed on MDA-MB 231 cell line by MTT assay and was found to be higher than that of pristine isofuranodiene.

Investigation of human pancreatic cancer tissues by Fourier Transform Infrared Hyperspectral Imaging.

Fourier-transform infrared hyperspectral imaging (FTIR-HSI) provides hyperspectral images containing both morphological and chemical information. It is widely applied in the biomedical field to detect tumor lesions, even at the early stage, by identifying specific spectral biomarkers. Pancreatic neoplasms present different prognoses and are not always easily classified by conventional analyses. In this study, tissue samples with diagnosis of pancreatic ductal adenocarcinoma and pancreatic neuroendocrine tumor were analyzed by FTIR-HSI and the spectral data compared with those from healthy and dysplastic samples. Multivariate/univariate approaches were complemented to hyperspectral images, and definite spectral markers of the different lesions identified. The malignant lesions were recognizable both from healthy/dysplastic pancreatic tissues (high values of phospholipids and triglycerides with shorter, more branched and less unsaturated alkyl chains) and between each other (different amounts of total lipids, phosphates and carbohydrates). These findings highlight different metabolic pathways characterizing the different samples, well detectable by FTIR-HSI.