In this study, ultrathin film multilayers of Photosystem II-enriched photosynthetic membranes (BBY) were prepared and immobilized on quartz substrates by means of a Layer by Layer procedure exploiting electrostatic interactions with poly(ethylenimine) as polyelectrolyte. The biomaterials thus obtained were characterized by means of optical techniques and Atomic Force Microscopy, highlighting the fact that the Layer by Layer approach allowed the BBYs to be immobilized with satisfactory results. The activity of these hybrid materials was evaluated by means of optical assays based on the Hill Reaction, indicating that the biosamples, which preserved about 65% of their original activity even ten weeks after preparation, were both stable and active. Furthermore, an investigation of the biochips’ sensitivity to the herbicide terbutryn, as a model analyte, gave interesting results: inhibition of photosynthetic activity was observed at terbutryn concentrations higher than 10-7 M, thus evidencing the potential of such biomaterials in the environmental biosensor field.

In this work, measurements of free acidity, peroxide content, spectrophotometric parameters, chlorophyll content, phenols, sterols, fatty acids and triacylglycerol composition, were carried out on samples of virgin olive oils (VOOs) coming from four different Greek Ionian islands, i.e. Zakynthos, Kefalonia, Lefkada and Kerkyra. An analysis of variance (ANOVA) highlighted statistically significant differences (p<0.01) in the values of 26 analytical parameters among the VOOs produced in the four different geographical regions but a Post-Hoc test showed that no variable was able to distinguish all four origins. Analogously, a Principal Component Analysis (PCA) showed a modest grouping of VOOs according to geographical origin except for Kerkyra samples which were more distinct from others. Applying discriminant function analysis (DFA) a good separation of the four geographical groups was achieved with classification and prediction abilities equal to 97.7% and 95.3%, respectively. Moreover, the analysis of the standardized coefficients showed that the fatty acids and triacylglycerols were the most discriminant variables. This last outcome was confirmed by comparison of the prediction performances obtained applying DFA on four subdatasets containing fatty acids (69.8%), triacylglycerols (76.7%), sterols (62.8%), and remaining parameters (65.1%) together, respectively. As the results showed, the multidisciplinary approach that combines different types of analytical determinations improved the discrimination of geographical origin for Greek virgin olive oils.

The aim of the present study was to characterize and classify olive oils from Western Greece according to cultivar and geographical origin, based on volatile compound composition, by means of Linear Discriminant Analysis. A total of 51 olive oil samples were collected during the harvesting period 2007–2008 from six regions of Western Greece and from six local cultivars. Forty-five of the samples were characterized as extra virgin olive oils. The analysis of volatile compounds was performed by Headspace Solid Phase Microextraction-Gas Chromatography/Mass Spectrometry (HS-SPME-GC/MS). Fifty-three (53) different volatile compounds were tentatively identified and semi-quantified. Using selected volatile compound composition data (selection was based on the application of ANOVA to total volatiles to determine those variables showing substantial differences among samples of different geographical origin/cultivar), the olive oil samples were satisfactorily classified according to geographical origin (87.2%) and cultivar (74%).

MalignantPleuralMesothelioma(MPM)isanaggressiveneoplasm that isveryoftenassociatedwithasbestosexposure.MPMdiagnosisisdifficult, veryoftenrequiringinvasiveapproachessuchasthoracoscopy.Exhaledbreath containshundredsofdifferentvolatileorganiccompounds(VOCs)whichmaybe usedfordiagnosisofvariouspulmonaryandsystemicdisordersincludinglung cancer. Methods: Weenrolled13patientswithanestablisheddiagnosisofMPM,13 healthy controls(HC),and13subjectswithaprofessionalexposuretoasbestos without MPM(EXP).Tedlarbagswereusedtocollecthumanbreath.Samples werecollectedusingasorbent-trapfollowedbythermaldesorptionandanalysis by gaschromatographwithananalyticalmassspectrometer(GC-MS). Results: AnalysisshowedthatthemostabundantVOCs(&gt;15 ng/L)inthein- vestigatedsampleswerecyclopentaneand cyclohexane.Thechemicalprofilewas differentforthesamplesclasses:EXPandMPM,showedalteredlevelsoftoluene, xylene, benzaldehyde,trimethylbenzene, limonene, 2-ethyl-1-hexanol,acetophe- none, cyclopentane.MPMpatientsshowed higherconcentrationofcyclohexane (meanvalue=339.31ng/L)comparedtoEXP(meanvalue=173.06ng/L)and controls(meanvalue=30.68ng/L).ByusingtheTukeyHSDtestitwasfound that cyclohexaneconcentrationforMPMwas significantlydifferentcomparedto HC (p=0.006)whereastheconcentrationforEXPwasnotsignificantlydifferent both comparedtoMPM(p=0.146)andcontrolgroup(p=0.285). Conclusions: BreathAnalysisbyGC-MSmayplayapotentialroleinthenon- invasiveassessmentofMPM.

Malignant pleural mesothelioma (MPM) is an aggressive tumour whose main aetiology is the long-term exposure to asbestos fibres. The diagnostic procedure of MPM is difficult and often requires invasive approaches; therefore, it is clinically important to find accurate markers for MPM by new noninvasive methods that may facilitate the diagnostic process and identify patients at an earlier stage. In the present study, the exhaled breath of 13 patients with histology-established diagnosis of MPM, 13 subjects with long-term certified professional exposure to asbestos (EXP) and 13 healthy subjects without exposure to asbestos (healthy controls, HC) were analysed. An analytical procedure to determine volatile organic compounds by sampling of air on a bed of solid sorbent and thermal desorption GC-MS analysis was developed in order to identify the compounds capable of discriminating among the three groups. The application of univariate (ANOVA) and multivariate statistical treatments (PCA, DFA and CP-ANN) showed that cyclopentane and cyclohexane were the dominant variables able to discriminate among the three groups. In particular, it was found that cyclohexane is the only compound able to differentiate the MPM group from the other two; therefore, it can be a possible marker of MPM. Cyclopentane is the dominant compound in the discrimination between EXP and the other groups (MPM and HC); then, it can be considered a good indicator for long-term asbestos exposure. This result suggests the need to perform frequent and thorough investigations on people exposed to asbestos in order to constantly monitor their state of health or possibly to study the evolution of disease over time. RI de gennaro, gianluigi/B-6237-2013 OI de gennaro, gianluigi/0000-0002-6868-6569

Authentic extravirgin olive oils from 7 different regions (Italy – 3 regions, Greece – 4 regions) have been investigated by 1H Nuclear Magnetic Resonance (NMR) fingerprinting in combination with multivariate statistical analysis. In order to cover the dominating lipid signals as well as signals from compounds of low abundance in the oil, both a simple one pulse experiment and an experiment with multiple saturation of the lipid signals was applied to each sample. Thus, the dynamic range of concentrations covered by the two experiments was of the order of 100,000 allowing for a more comprehensive NMR assessment of the samples. Monte-Carlo embedded cross-validation was used to demonstrate that a combination of principal component analysis, canonical analysis, and classification via nearest class mean can be used to predict the origin of olive oil samples from 1H NMR data. Given the rather limited number of samples tested, correct prediction probabilities of 78% were achieved with region specific correct predictions between 53% and 100%.

A new analytical method for the determination of ochratoxin A (OTA) in red wine has been developed by using a double-extract cleanup and a fluorometric measurement after spectral deconvolution. Wine samples were diluted with a solution containing 1% polyethylene glycol and 5% sodium hydrogencarbonate, filtered, and purified by immunoaffinity and aminopropyl solid-phase column. OTA contents in the purified extract were determined by a spectrofluorometer (excitation wavelength, 330 nm; emission wavelength, 470 nm) after deconvolution of fluorescence spectra. Average recoveries from wine samples spiked with OTA at levels ranging from 0.5 to 3.0 ng/mL were 94.5–105.4% with relative standard deviations (RSD) of <15% (n = 4). The limit of detection (LOD) was 0.2 ng/mL, and the total time of analysis was 30 min. The developed method was tested on 18 red wine samples (naturally contaminated and spiked with OTA at levels ranging from 0.4 to 3.0 ng/mL) and compared with AOAC Official Method 2001.01, based on immunoaffinity column cleanup and HPLC with fluorescence detector. A good correlation (r2 = 0.9765) was observed between OTA levels obtained with the two methods, highlighting the reliability of the proposed method, the main advantage of which is the simple OTA determination by a benchtop fluorometer with evident reductions of cost and time of analysis.

OBJECTIVE: The aim of this study was to determine whether the volatile organic compounds (VOCs) pattern in colorectal cancer (CRC) patients is modified by curative surgery for a potential application in the oncologic follow-up. BACKGROUND: CRC has been proved to induce metabolic derangements detectable by high through-output techniques in exhaled breath showing a specific pattern of VOCs. METHODS: Forty-eight CRC patients and 55 healthy controls (HC) entered the study. Thirty-two patients (M/F: 1.4; mean age 63 years) attended the oncologic follow-up (mean 24 months) and were found disease-free. Breath samples were collected under similar environmental conditions into a Tedlar bags and processed offline by thermal-desorption gas chromatography-mass spectrometry (TD-GC-MS). VOCs were selected by U test to build a Probabilistic Neural Network (PNN) model to set-up a training phase, which was cross-validated using the leave-one out method. RESULTS: A total of 11 VOCs were finally selected for their excellent discriminant performance in identifying disease-free patients in follow-up from CRC patients before surgery, (sensitivity 100%, specificity 97.92%, accuracy 98.75%, and AUC: 1). The same VOCs pattern discriminated follow-up patients from HC, with a sensitivity of 100%, specificity of 90.91%, accuracy of 94.25%, and AUC 0.959. CONCLUSIONS: Exhaled VOCs pattern from CRC patients is modified by cancer removal confirming the tight relationship between tumor metabolism and exhaled VOCs. PNN analysis provides a high discriminatory tool to identify patients disease-free after curative surgery suggesting potential implications in CRC screening and secondary prevention.

Effects of curative surgery of colorectal cancer on exhaled volatile organic compounds (VOCs) after 1 year follow up

In order to investigate the effects of cryomaceration and reductive vinification on chemical and physical indices and on antioxidant compounds of Sauvignon blanc wines, four wine-making procedures were applied: traditional white vinification, skin cryomaceration, vinification in a reductive environment, and a combination of the last two procedures. Significant differences were highlighted by both conventional analyses and NMR spectroscopy. The strongest changes were for organic acid concentrations (tartaric, in particular) and phenolic content. Cryomaceration caused a strong precipitation of tartaric acid, which may be desired if grapes have high acidity values. Cryomaceration protected those fiavans reactive with vanillin from the action of oxidative enzymes. Vinification in a reductive environment, alone or combined with a cryomaceration step, gave wines with the highest solids content and caused a greater extraction of phenolic compounds from skins compared to traditional winemaking or cryomaceration alone, due to SO2 solubilisation. Grape oenological expression can be strongly affected by the application of the investigated wine-making procedures. (C) 2012 Elsevier Ltd. All rights reserved.

Sweet cherries from two Italian regions, Apulia and Emilia Romagna, were analysed using electronic nose (EN) and isotope ratio mass spectrometry (IRMS), with the aim of distinguishing them according to their geographic origin. The data were elaborated by statistical techniques, examining the EN and IRMS datasets both separately and in combination. Preliminary exploratory overviews were performed and then linear discriminant analyses (LDA) were used for classification. Regarding EN, different approaches for variable selection were tested, and the most suitable strategies were highlighted. The LDA classification results were expressed in terms of recognition and prediction abilities and it was found that both EN and IRMS performed well, with IRMS showing better cross-validated prediction ability (91.0%); the EN–IRMS combination gave slightly better results (92.3%). In order to validate the final results, the models were tested using an external set of samples with excellent results.

Although several studies have used bogs in order to reconstruct paleoclimatic conditions and the historical trends of pollutants, scientific literature is still rather controversial about the role of ombrotrophic bogs as reliable record of past environmental changes. Consequently, understanding whether all vegetational and climatic ginformationh are effectively preserved in peat deposits during humification becomes an essential aspect to be tested before using bogs as natural archives. The present work focuses on stable isotopic ratios, i.e., 13C/12C (ƒÂ13C), 15N/14N (ƒÂ15N) and 18O/16O (ƒÂ18O), and is based on the assumption that, if bogs are consistent archives of environmental changes, these types of ginformationh should be recorded also into humic acids (HA), i.e. the fraction of peat more recalcitrant and refractory to degradation. Thus, an 81-cm long peat core, covering the last 2000 years, was collected from the Etang de la Gruere bog (Jura Mountains, Switzerland), cut into 3 cm slices, and HA were isolated from each age dated layer. Stable isotopic ratios (ƒÂ13C, ƒÂ15N and ƒÂ18O) were then determined in bulk peat and corresponding HA samples. An increase in the humification degree and a decrease of the C/N ratio were observed along the profile. The ƒÂ13C of both peat and HA showed a significantly similar trend with depth (p=0.0001), and the same significant correlation was observed for the ƒÂ15N (pb0.0001). Also the ratio between ƒÂ13C and ƒÂ15N closely resembled the C/N ratio observed in the peat and in HA (pb0.0001), thus underlining that the trend of these isotopic ratios is preserved along the studied bog profile. Consequently, our data seem to support both the role of HA as recalcitrant, stable molecules with a long-term residence time, and the potential of ombrotrophic bogs to be used as garchivesh of vegetational changes occurring (at least) in the last 2000 years. Although a certain relationship (pb0.05) between peat and corresponding HA was found also for ƒÂ18O, our data did not allow the solving of the issue of its reliability as paleoenvironmental proxy.

A detailed physico-chemical characterization of the Garganica kid goat meat was carried out on samples (quadriceps femoris muscles) coming from different farms located on the Promontory of Gargano, in Apulia (southern Italy) by determining conventional parameters (moisture, ashes, fat and protein content), stable isotope ratios (15N/14N and 13C/12C), major (Ca, Mg, Na, K), and trace metals (Zn, Mn, Cu, Fe, Cr). Moreover, information on a large number of metabolites were obtained by means of 1H High-Resolution Magic-Angle Spinning Nuclear Magnetic Resonance (1H HR-MAS NMR) spectroscopy. The most interesting findings of this work, for their impact on human nutrition and health, were represented by a considerable protein content (18.9%) and a low fat content (2.98%), characterized by a remarkable average percentage of desirable fatty acids (62.76%) and a good (C18 + C18:1)/C16 average ratio (2.30), that make the Garganica kid goat meat a highly appreciable food product.

In this paper, virgin olive oils (VOOs) coming from three different geographic origins of Apulia, were analysed for free acidity, peroxide value. spectrophotometric indexes, chlorophyll content. sterol, fatty acid, and triacylglycerol compositions. In order to predict the geographical origin of VOOs, different multivariate approaches were applied. By performing principal component analysis (PCA) a modest natural grouping of the VOOs was observed on the basis of their origin, and consequently three supervised techniques, i.e., general discriminant analysis (GDA), partial least squares-discriminant analysis (PLS-DA) and soft independent modelling of class analogy (SIMCA) were used and the results were compared. In particular, the best prediction ability was produced by applying GDA (average prediction ability of 82.5%), even if interesting results were obtained also by applying the other two classification techniques, i.e., 77.2% and 75.5% for PLS-DA and SIMCA, respectively. (C) 2012 Elsevier Ltd. All rights reserved.

T-2 and HT-2 toxins are mycotoxins produced by several Fusarium species that are commonly found in various cereal grains, including oats, barley, wheat and maize. Intake estimates indicate that the presence of these mycotoxins in the diet can be of concern for public health. In this work, the inclusion processes occurring between fluorescent anthracene-derivatives of T-2 and HT-2 toxins and different cyclodextrin (CD) molecules were investigated in aqueous solutions by means of UV–Vis absorption, fluorescence emission and dynamic light scattering. Binding constant values and chemico-physical parameters were calculated. It was found that b-CDs give stronger inclusion reactions with both T-2 and HT-2 derivatives, as stated by important emission intensity increments. Such interactions were found to be fundamentally enthalpy-driven. Among b-CDs, the effect of the methylation at hydroxyl groups was tested: as a result, the di-methyl form of b-CD was found to induce the best fluorescence intensity enhancements.

The European Union provides important guidelines for maintaining the Protected Designation of Origin of numerous food products, as in case of the olive oil, since the origin marks certify a high quality level of food, conferring, as a consequence, prestige and economic value. Therefore, in this context it is extremely important to consider the topic of the characterization of foods based on variety (cultivar) and geographical origin, as this may be used as a criterion for determining authenticity and quality. This chapter constitutes an overview of the results obtained by the authors through studies aiming at the differentiation of the geographic and varietal origin of olive oils, dealing with the classification of Italian olive cultivars based on compositional data and Nuclear Magnetic Resonance determinations, and with the characterization of the geographical origin of Western Greek and Italian virgin olive oils based on instrumental and multivariate statistical analysis.

In this study, non-targeted 1H NMR fingerprinting was used in combination with multivariate statistical techniques for the classification of Italian sweet cherries based on their different geographical origins (Emilia Romagna and Puglia). As classification techniques, Soft Independent Modelling of Class Analogy (SIMCA), Partial Least Squares Discriminant Analysis (PLS-DA), and Linear Discriminant Analysis (LDA) were carried out and the results were compared. For LDA, before performing a refined selection of the number/combination of variables, two different strategies for a preliminary reduction of the variable number were tested. The best average recognition and CV prediction abilities (both 100.0%) were obtained for all the LDA models, although PLS-DA also showed remarkable performances (94.6%). All the statistical models were validated by observing the prediction abilities with respect to an external set of cherry samples. The best result (94.9%) was obtained with LDA by performing a best subset selection procedure on a set of 30 principal components previously selected by a stepwise decorrelation. The metabolites that mostly contributed to the classification performances of such LDA model, were found to be malate, glucose, fructose, glutamine and succinate.

Photosystem II (PSII) proteins from spinach leaves were immobilized onto quartz substrates according to the Layer-by-Layer (LbL) procedure, alternating protein to polyethylenimine (PEI) layers by exploiting electrostatic interactions. The effects of several factors, such as storage conditions, ageing of the PSII-modified substrates, as well as PSII concentration in buffer, on the quality of the prepared multilayers, were investigated by UV-vis Absorption Spectroscopy and Atomic Force Microscopy (AFM). A number of 13 layers was found to be optimal to guarantee intense PSII optical signals with homogeneous morphological distributions of proteins. The multilayers resulted stable if stored in contact with air at 4 degrees C, as observed by UV-vis Absorption spectra recorded after 48 h. The best results in terms of AFM images and electron transfer efficiency (measured by Hill Reaction assays) were gained by using 5.6 x 10(-7) M chlorophyll concentration, obtaining multilayers with the most ordered protein distributions and the highest electron transfer efficiency, i.e. 85 % of an iso-absorbing PSII suspension. The results highlight the possibility to successfully immobilize PSII proteins, without considerable loss of bioactivity, thanks to the mild nature of the electrostatic LbL technique, opening up possibilities of applications in the bioelectrochemical energy conversion and biosensoristic fields.

The pH-related characteristics of 4-thiothymidine and its stability during prolonged exposure, at room temperature, to a neon lamp emitting in the 400–700 nm wavelength range were investigated by different spectroscopic techniques (UV-Vis, FTIR-ATR, 1H-NMR) and by ElectroSpray Ionization Mass Spectrometry (ESI-MS). The evaluation of the nucleoside photostability was performed as a control, with the perspective of studying its reactivity under the same conditions but in the presence of visible lightabsorbing photosensitizers, able to generate reactive oxygen species. The comparison between UV-Vis spectra recorded at different pH values in the 7–12 range suggested the presence of an equilibrium related to the deprotonation of the N3–H group of 4-thiothymidine, with a pKa estimated to be close to 9. Some effects of the deprotonation occurring at alkaline pH were observed also in FTIR-ATR spectra, the main feature being the appearance of a band related to C]N stretching, interpreted with the assumption of a partial double bond character by C2–N3, N3–C4 and C2–O bonds, as a consequence of negative charge delocalization on the pyrimidine ring. As for photostability, UV-Vis, FTIR-ATR and NMR measurements suggested the generation of thymidine as a by-product but only after a prolonged (48 hours) irradiation time, whereas no significant alteration occurred in a shorter time range (1–2 hours), i.e. the one that will be considered in future studies involving the presence of photosensitizers. The nucleoside stability up to 2 hours of irradiation was confirmed by ESI-MS analyses; furthermore, on the other hand, the latter indicated the presence of three additional by-products, besides thymidine, after 48 hours of irradiation. In particular, an hydroxylated form of 4-thiothymidine and two dimeric species, characterized by S–S and S–O covalent bridges between two 4-thiothymidine and a 4-thiothymidine and a thymidine molecule, respectively, were detected.

The interaction of ochratoxin A (OTA) with heptakis-2,6-di-O-methyl-β-cyclodextrin (DIMEB) in aqueous solutions at two different pHs (3.5 and 9.5) was studied by means of spectroscopic, calorimetric and electrochemical techniques. DIMEB affected the physico-chemical properties of OTA. A stronger interaction was observed at pH = 3.5 where the neutral form of OTA prevails. The spectroscopic information indicated that the OTA/DIMEB inclusion process occurred through the insertion of the phenylalanine group into the cyclodextrin (CD) cavity; moreover, voltammetry experiments showed that the isocoumarinic phenolic group was also involved, probably by non-inclusion interactions. At pH = 3.5, binding constants and thermodynamic parameters of the OTA/DIMEB complex were determined by means of the modified Benesi-Hildebrand equation and van’t Hoff plot. A 1:1 stoichiometry for the OTA/DIMEB complex was observed with a binding constant equal to 530 ± 50 L·mol−1 at 25 °C. The calculated thermodynamic parameters indicate that the interaction is highly endothermic and that the complex formation is driven by entropy. The results provide useful information for potential applications of cyclodextrins in the analysis of mycotoxins and in the field of mycotoxin sequestering agents

The discovery of a storeroom full of barley and other cereals (L.9512) in the proto-historic site of Ebla has provided a unique opportunity to study the centralized storage system of the early city-state from a different perspective. Epigraphic evidence available within the site reveals a complex system of taxation which included gathering grain tributes from satellite sites and redistributing semi-finished products such as flour. In this paper, we intend to explore the possibilities of a combined approach to studying the storage system, based on estimated barley grain volumes and d13C-d15N analyses. This approach is used to distinguish between grain from different harvesting sites and to identify any grain cultivated using special agricultural practices (e.g. manuring or irrigation). The basic assumption for this kind of analysis is that the growth-site conditions, natural or anthropogenic, of harvested cereals are reflected in their grain size and d13C-d15N values. Since the remains found in the storeroom were charred, the first task was to evaluate the effect of carbonization on the d13C-d15N and the size of the grains. Thus, the effect of charring was tested on modern samples of Syrian barley landraces. Once it had been ascertained that fresh grains reduced to charred remains retain their original biometric and isotopic traits, the ancient material was examined. Thirteen groups were identified, each characterized by a specific average volume and specific carbon and nitrogen values. The analysis revealed that what had first appeared to be a homogeneous concentration of grain was in fact an assemblage of barley harvested from different sites.

A method is described for the total phosphine detection in cereal caryopses, comprising the steps of inserting a sample of cereal caryopses inside a container (1) equipped with hermetic closure (2); adding to the sample an aqueous solution of H 2 SO 4 with a v/v concentration in the range of 5-20%, with obtainment of an aqueous dispersion, and hermetically closing the first container (1); subjecting the aqueous dispersion contained in the first container (1) to the action of microwaves for a time not greater than 3 minutes; drawing a predetermined volume of gas overlying the aqueous dispersion and detecting any phosphine that may be present by means of colorimetric and/or spectrophotometric methods, preferably by bringing it into contact with a predetermined volume of an aqueous solution of AgNO 3 of known molarity, inside a second container (3) with hermetic closure and visually analysing the obtained colour and/or spectrophotometrically measuring the absorbance at 400 nm of the aqueous solution. Equipment for the execution of the aforesaid method is described