XRF analysis searching for fingerprint elemental profile in south-eastern Sicily tomatoes.

The implementation of analytical techniques able to certify food quality and origin in a fast and non-destructive way is becoming a widespread need in the agri-food sector. Among the physical non-destructive techniques, X-ray fluorescence (XRF) spectrometry is often used to analyze the elemental composition of biological samples. In this study, X-ray fluorescence (XRF) elemental profiles were measured on tomato samples belonging to different geographical areas in Sicily (Italy). The purpose of this investigation was aiming to establish a protocol for in-situ measurement and analysis able to provide quality assessment and traceability of PGI agri-food products, specifically sustaining health safety and self qualifying bio-chemical signature. In detail, sampling was performed in one of the most tomato productive area of south-eastern Sicily (Pachino district), characterised by a relative higher amount of Organic Carbon and Cation Exchange Capacity, and compared with samples from other growing areas of Sicily, falling in Ragusa province and Mt. Etna region. Experimental data were analyzed in the framework of multivariate analysis by using principal component analysis and further validated by discriminant analysis. The results show the presence of specific elemental signatures associated to several characterizing elements. This methodology establishes the possibility to disentangle a clear fingerprint pattern associated to the geographical origin of an agri-food product.

Applications of Delayed Luminescence for tomato fruit quality assessment across varied Sicilian cultivation zones.

The food industry places significant emphasis on ensuring quality and traceability as key components of a healthy diet. To cater to consumer demands, researchers have prioritized the development of analytical techniques that can rapidly and non-invasively provide data on quality parameters. In this study, we propose to use the Delayed Luminescence (DL), an ultra-weak and photo-induced emission of optical photons, as a tool for a rapid evaluation of quality profile associated with fruit ripening, in support of traditional analysis methods. Delayed Luminescence measurements have been performed on cherry tomatoes, with and without the PGI "Pomodoro di Pachino" certification, harvested from two different growing areas of south-eastern Sicily (Italy). Then, DL emissions were correlated with soluble solid content and titratable acidity values, which are known to affect the flavor, the commerciality and the maturity degree of tomato fruits. In addition, we evaluated the changes in the DL parameters with respect to the geographical origin of the cherry tomatoes, with the aim of testing the possibility of applying the technique for identification purposes. The signals of Delayed Luminescence appeared to be good indicators of the macromolecular structure of the biological system, revealing structural changes related to the content of total soluble solids present in the juice of tomatoes analyzed, and they appeared unsuitable for authenticating vegetable crops, since the differences in the photon yields emitted by tomato Lots were not related to territory of origin. Thus, our results suggest that DL can be used as a nondestructive indicator of important parameters linked to tomato fruit quality.

Effect of Astaxanthin on Tissue Transglutaminase and Cytoskeletal Protein Expression in Amyloid-Beta Stressed Olfactory Ensheathing Cells: Molecular and Delayed Luminescence Studies.

Astaxanthin, a natural compound of Haematococcus pluvialis, possesses antioxidant, anti-inflammatory, anti-tumor and immunomodulatory activities. It also represents a potential therapeutic in Alzheimer's disease (AD), that is related to oxidative stress and agglomeration of proteins such as amyloid-beta (Aß). Aß is a neurotoxic protein and a substrate of tissue transglutaminase (TG2), an ubiquitary protein involved in AD. Herein, the effect of astaxanthin pretreatment on olfactory ensheathing cells (OECs) exposed to Aß(1-42) or by Aß(25-35) or Aß(35-25), and on TG2 expression were assessed. Vimentin, GFAP, nestin, cyclin D1 and caspase-3 were evaluated. ROS levels and the percentage of cell viability were also detected. In parallel, delayed luminescence (DL) was used to monitor mitochondrial status. ASTA reduced TG2, GFAP and vimentin overexpression, inhibiting cyclin D1 levels and apoptotic pathway activation which induced an increase in the nestin levels. In addition, significant changes in DL intensities were particularly observed in OECs exposed to Aß toxic fragment (25-35), that completely disappear when OECs were pre-incubated in astaxantin. Therefore, we suggest that ASTA pre-treatment might represent an innovative mechanism to contrast TG2 overexpression in AD.

Synergic pro-apoptotic effects of Ferulic Acid and nanostructured lipid carrier in glioblastoma cells assessed through molecular and Delayed Luminescence studies.

Herein, we assessed the effect of Ferulic Acid (FA), a natural antioxidant with anti-cancer effect, on the human glioblastoma cells through molecular and Delayed Luminescence (DL) studies. DL, a phenomenon of ultra-week emission of optical photons, was used to monitor mitochondrial assessment. The effect of FA loaded in nanostructured lipid carriers (NLCs) was also assessed. To validate NLCs as a drug delivery system for glioblastoma treatment, particular attention was focused on their effect. We found that free FA induced a significant decrease in c-Myc and Bcl-2 expression levels accompanied by the apoptotic pathway activation. Blank NLCs, even if they did not induce cytotoxicity and caspase-3 cleavage, decreased Bcl-2, ERK1/2, c-Myc expression levels activating PARP-1 cleavage. The changes in DL intensity and kinetics highlighted a possible effect of nanoparticle matrix on mitochondria, through the involvement of the NADH pool and ROS production that, in turn, activates ERK1/2 pathways. All the effects on protein expression levels and on the activation of apoptotic pathway appeared more evident when the cells were exposed to FA loaded in NLCs. We demonstrated that the observed effects are due to a synergic pro-apoptotic influence exerted by FA, whose bio-availability increases in the glioblastoma cells, and NLCs formulation.

Dynamic changes in cytoskeleton proteins of olfactory ensheathing cells induced by radiofrequency electromagnetic fields.

Several evidences have suggested the ability of radiofrequency electromagnetic fields to influence biological systems, even if the action mechanisms are not well understood. There are few data on the effect of radiofrequency electromagnetic fields on self-renewal of neural progenitor cells. A particular glial type that shows characteristics of stem cells is olfactory ensheathing cells (OECs). Herein, we assessed the non-thermal effects induced on OECs through radiofrequency electromagnetic fields changing the envelope of the electromagnetic wave. Primary OEC cultures were exposed to continuous or amplitude-modulated 900 MHz electromagnetic fields, in the far-field condition and at different exposure times (10, 15, 20 min). The expression of OEC markers (S-100 and nestin), cytoskeletal proteins (GFAP and vimentin), apoptotic pathway activation by caspase-3 cleavage and cell viability were evaluated. Our results highlight that 20 min of exposure to continuous or amplitude-modulated 900 MHz electromagnetic fields induced a different and significant decrease in cell viability. In addition, according to the electromagnetic field waveform, diverse dynamic changes in the expression of the analysed markers in OECs and activation of the apoptotic pathway were observed. The data suggest that radiofrequency electromagnetic fields might play different and important roles in the self-renewal of OEC stem cells, which are involved in nervous system repair.

Impact of structure on the delayed luminescence of d-Glucose-based polymer chains.

Glucose is a natural chemical compound and is one of the most abundant organic molecules in nature. Plants and algae are able to produce it from water and carbon dioxide during photosynthesis, using energy of photons coming from the sun. It is very important in life processes because, in energy metabolism, Glucose is the most important source of energy in all organisms. As energy reservoir it is partially stored as a polymer, in plants mainly as starch and amylopectin and in animals as glycogen. Moreover it is used as cellulose, the most abundant carbohydrate, to strengthen plants and algae cell walls. In this paper we study the Delayed Luminescence from Glucose and its polymers, Amylose and Cellulose, composed by chains of glucose connected by different bond, as well as Glucose water solution, in order to acquire new knowledge on the mechanisms responsible for this phenomenon and check the possibility to give in-depth analysis of possible collective states present in Glucose-based structures. The phenomenon of DL in biological systems is not a byproduct, as one can naively expect. Instead, it is a property and necessity of the condensed matter, which can be also used as a tool to study the latter. It is a manifestation of the physical and biochemical processes in the system, on one hand side, and, on the other hand side, of its structural properties, in particular, of the presence and type of crystal-like structure, resulting in specific energy spectrum and electron transitions, as will be presented below. We show that the quantum yield and time trends of the Delayed Luminescence depend on the structure of systems under study. Significant differences in Delayed Luminescence parameters from cellulose before and after imbibition have been observed, indicating that Delayed Luminescence could be used to discriminate between various structures and follow the formation or demolition of them. The experimental results qualitatively agree with the soliton mechanism of the Delayed luminescence.

Non-destructive evaluation of watermelon seeds germination by using Delayed Luminescence.

The evaluation of optical properties of biological samples is gaining increasing interests both in scientific and commercial fields concerning agriculture and food processing. The optical techniques can indeed be able to provide information on quality assessment in a fast and non-destructive way. This feature makes them suitable for automatic management of control processes. In this paper, we propose to use the Delayed Luminescence, a ultra-weak and photo-induced emission of optical photons, as a tool for a rapid evaluation of germination performance, the principal index reflecting seed quality. Two lots of 'Mirella' F1 watermelon dried seeds, of 96 seeds each, were considered. The seeds were analyzed in the conditions as provided by seed/breeding company. Characteristics of Delayed Luminescence emission from each single seed were correlated to the different germination levels as assessed by International Seed Testing Procedures. Parametric differences in the two lots were determined, based on the relaxation kinetics of some spectral components. A control test, with the aim to construct a calibration model, was conceived and successfully tested. Time decays of Delayed Luminescence spectral components at central wavelengths 450 nm, 550 nm and 650 nm, corresponding to spectrum region where natural biomarkers as NADH, flavins and lipopigments, protoporphyrin and ROS respectively emit, have been evaluated. The results show that such time decays are strictly connected to the biological state of the system under analysis and allow also proposing Delayed Luminescence measurement as a quick, cheap and non-destructive test for seed viability analysis.

Exploring the behaviour of water in glycerol solutions by using delayed luminescence.

The crucial role of water in the engine of life have encouraged many researchers in studying, both theoretically and experimentally, the possible "structure" of water. Many properties of water have been related to the interplay between two distinct and interconverting structural species, namely the low-density water (LDW) and the high-density water (HDW). Supported by the results obtained with other aqueous solutions, this paper deals with the possibility of using the ultra-weak delayed luminescence (DL) to investigate water structuring in a mixture with glycerol, characterized only by hydrogen bonds between the various molecules. Spectral and temporal characteristics of DL decays give information on the two components of the mixture, by evidencing the contribution of water at glycerol concentrations close to the values used in cryopreservation. DL results have shown a correlation with LDW clusters size as determined by other researchers on the basis of neutron diffraction experiments and computational modelling, as reported in Literature.

Effects of Ion Irradiation on Seedlings Growth Monitored by Ultraweak Delayed Luminescence.

The optical technique based on the measurement of delayed luminescence emitted from the biological samples has demonstrated its ability to provide valid and predictive information on the functional status of various biological systems. We want to extend this technique to study the effect of ionizing radiation on biological systems. In particular we are interested in the action of ion beams, used for therapeutic purposes or to increase the biological diversity. In general, the assessment of the damage that radiation produces both in the target objects and in the surrounding tissues, requires considerable time because is based on biochemical analysis or on the examination of the evolution of the irradiated systems. The delayed luminescence technique could help to simplify this investigation. We have so started our studies performing irradiations of some relatively simple vegetable models. In this paper we report results obtained from mung bean (Vigna radiata) seeds submitted to a 12C ion beam at the energy of 62 MeV/nucleon. The dry seeds were irradiated at doses from 50 to 7000 Gy. The photoinduced delayed luminescence of each seed before and after ion irradiation was measured. The growth of seedlings after irradiation was compared with that of untreated seeds. A growth reduction on increasing the dose was registered. The results show strong correlations between the ion irradiation dose, seeds growth and delayed luminescence intensity. In particular, the delayed luminescence intensity is correlated by a logistic function to the seedlings elongation and, after performing a suitable measurement campaign based on blind tests, it could become a tool able to predict the growth of seeds after ion irradiation. Moreover these results demonstrate that measurements of delayed luminescence could be used as a fast and non-invasive technique to check the effects of ion beams on relatively simple biological systems.

Delayed luminescence to monitor programmed cell death induced by berberine on thyroid cancer cells.

Correlation between apoptosis and UVA-induced ultraweak photon emission delayed luminescence (DL) from tumor thyroid cell lines was investigated. In particular, the effects of berberine, an alkaloid that has been reported to have anticancer activities, on two cancer cell lines were studied. The FTC-133 and 8305C cell lines, as representative of follicular and anaplastic thyroid human cancer, respectively, were chosen. The results show that berberine is able to arrest cell cycle and activate apoptotic pathway as shown in both cell lines by deoxyribonucleic acid fragmentation, caspase-3 cleavage, p53 and p27 protein overexpression. In parallel, changes in DL spectral components after berberine treatment support the hypothesis that DL from human cells originates mainly from mitochondria, since berberine acts especially at the mitochondrial level. The decrease of DL blue component for both cell lines could be related to the decrease of intra-mitochondrial nicotinamide adenine dinucleotide and may be a hallmark of induced apoptosis. In contrast, the response in the red spectral range is different for the two cell lines and may be ascribed to a different iron homeostasis.

Ultra-weak delayed luminescence in cancer research: a review of the results by the ARETUSA equipment.

The study of the photoinduced ultraweak photon emission in the optical wavelength range, namely the Delayed Luminescence, from human cells and tissues has an increasingly growing interest in view of its possible application in optical biopsy. Due to the low level, dedicated experimental set-up are necessary to reveal such photoluminescence signal. The paper reviews the results obtained in the field of cancer research, by using the experimental equipment for fast ultraweak luminescence analysis ARETUSA developed at the National Southern Laboratories of the National Nuclear Physics Institute (LNS-INFN), in Catania, Italy. Delayed Luminescence signals from normal and cancer cells are compared and the relationship between Delayed Luminescence and apoptosis is investigated.

Mitochondrial respiratory complex I probed by delayed luminescence spectroscopy.

The role of mitochondrial complex I in ultraweak photon-induced delayed photon emission [delayed luminescence (DL)] of human leukemia Jurkat T cells was probed by using complex I targeting agents like rotenone, menadione, and quercetin. Rotenone, a complex I-specific inhibitor, dose-dependently increased the mitochondrial level of reduced nicotinamide adenine dinucleotide (NADH), decreased clonogenic survival, and induced apoptosis. A strong correlation was found between the mitochondrial levels of NADH and oxidized flavin mononucleotide (FMNox) in rotenone-, menadione- and quercetin-treated cells. Rotenone enhanced DL dose-dependently, whereas quercetin and menadione inhibited DL as well as NADH or FMNox. Collectively, the data suggest that DL of Jurkat cells originates mainly from mitochondrial complex I, which functions predominantly as a dimer and less frequently as a tetramer. In individual monomers, both pairs of pyridine nucleotide (NADH/reduced nicotinamide adenine dinucleotide phosphate) sites and flavin (FMN-a/FMN-b) sites appear to bind cooperatively their specific ligands. Enhancement of delayed red-light emission by rotenone suggests that the mean time for one-electron reduction of ubiquinone or FMN-a by the terminal Fe/S center (N2) is 20 or 284 µs, respectively. All these findings suggest that DL spectroscopy could be used as a reliable, sensitive, and robust technique to probe electron flow within complex I in situ.

Detailed analysis of apoptosis and delayed luminescence of human leukemia Jurkat T cells after proton irradiation and treatments with oxidant agents and flavonoids.

Following previous work, we investigated in more detail the relationship between apoptosis and delayed luminescence (DL) in human leukemia Jurkat T cells under a wide variety of treatments. We used menadione and hydrogen peroxide to induce oxidative stress and two flavonoids, quercetin, and epigallocatechin gallate, applied alone or in combination with menadione or H(2)O(2). 62 MeV proton beams were used to irradiate cells under a uniform dose of 2 or 10 Gy, respectively. We assessed apoptosis, cell cycle distributions, and DL. Menadione, H(2)O(2) and quercetin were potent inducers of apoptosis and DL inhibitors. Quercetin decreased clonogenic survival and the NAD(P)H level in a dose-dependent manner. Proton irradiation with 2 Gy but not 10 Gy increased the apoptotic rate. However, both doses induced a substantial G(2)/M arrest. Quercetin reduced apoptosis and prolonged the G(2)/M arrest induced by radiation. DL spectroscopy indicated that proton irradiation disrupted the electron flow within Complex I of the mitochondrial respiratory chain, thus explaining the massive necrosis induced by 10 Gy of protons and also suggested an equivalent action of menadione and quercetin at the level of the Fe/S center N2, which may be mediated by their binding to a common site within Complex I, probably the rotenone-binding site.

Delayed luminescence: a novel technique to obtain new insights into water structure.

Fully understanding the structure of water is a crucial point in biophysics because this liquid is essential in the operation of the engines of life. Many of its amazing anomalies seem to be tailored to support biological processes and, during about a century, several models have been developed to describe the water structuring. In particular, a theory assumes that water is a mixture of domains constituted by two distinct and inter-converting structural species, the low-density water (LDW) and the high-density water (HDW). According to this theory, by using some particular solutes or changing the water temperature, it should be possible to modify the equilibrium between the two species, changing in this way the water behavior in specific biological processes, as in governing the shape and stability of the structures of proteins. In this work, we assess the possibility of obtaining information on the structures induced in water by specific salts or by temperature by measuring the delayed luminescence (DL) of some salt solutions and of water in the super-cooled regime. Previous works have demonstrated that the delayed luminescence of a system is correlated with its dynamic ordered structures. The results show significant DL signals only when the formation of LDW domains is expected. The measurement reveals a similar activation energy for the domains both in aqueous salt solutions and super-cooled water. It is worth noting that the time trend of DL signals suggests the existence of structures unusually long-lasting in time, up to the microsecond range.

Effects of menadione, hydrogen peroxide, and quercetin on apoptosis and delayed luminescence of human leukemia Jurkat T-cells.

Menadione (MD) is an effective cytotoxic drug able to produce intracellularly large amounts of superoxide anion. Quercetin (QC), a widely distributed bioflavonoid, can exert both antioxidant and pro-oxidant effects and is known to specifically inhibit cell proliferation and induce apoptosis in different cancer cell types. We have investigated the relation between delayed luminescence (DL) induced by UV-laser excitation and the effects of MD, hydrogen peroxide, and QC on apoptosis and cell cycle in human leukemia Jurkat T-cells. Treatments with 500 µM H2O2 and 250 µM MD for 20 min produced 66.0 ± 4.9 and 46.4 ± 8.6% apoptotic cell fractions, respectively. Long-term (24 h) pre-exposure to 5 µM, but not 0.5 µM QC enhanced apoptosis induced by MD, whereas short-term (1 h) pre-incubation with 10 µM QC offered 50% protection against H2O2-induced apoptosis, but potentiated apoptosis induced by MD. Since physiological levels of QC in the blood are normally less than 10 µM, these data can provide relevant information regarding the benefits of flavonoid-combined treatments of leukemia. All the three drugs exerted significant effects on DL. Our data are consistent with (1) the involvement of Complex I of the mitochondrial respiratory chain as an important source of delayed light emission on the 10 µs-10 ms scale, (2) the ability of superoxide anions to quench DL on the 100 µs-10 ms scale, probably via inhibition of reverse electron transfer at the Fe/S centers in Complex I, and (3) the relative insensitivity of DL to intracellular OH• and H2O2 levels.

Generalization of DT equations for time dependent sources.

New equations for paralyzable, non paralyzable and hybrid DT models, valid for any time dependent sources are presented. We show how such new equations include the equations already used for constant rate sources, and how it's is possible to correct DT losses in the case of time dependent sources. Montecarlo simulations were performed to compare the equations behavior with the three DT models. Excellent accordance between equations predictions and Montecarlo simulation was found. We also obtain good results in the experimental validation of the new hybrid DT equation. Passive quenched SPAD device was chosen as a device affected by hybrid DT losses and active quenched SPAD with 50 ns DT was used as DT losses free device.

Time-resolved spectral measurements of delayed luminescence from a single soybean seed: effects of thermal damage and correlation with germination performance.

Delayed luminescence from a single dry soybean seed was investigated in both spectral and time domains, under different excitation wavelengths. Emission spectra were collected, under 337 nm laser excitation, from native and artificially deteriorated seeds and the time-dependence of different spectral components was analyzed in detail. The single seed viability was evaluated through observation of germination properties after imbibition and compared with different parameters related to the luminescence kinetics. The significant correlation found between single seed delayed luminescence parameters and germination capability strongly validates the connection of this phenomenon with the functional state of the system and suggests the development of a non-invasive technique for seed quality determination.

Laser-ultraviolet-A induced ultra weak photon emission in human skin cells: A biophotonic comparison between keratinocytes and fibroblasts.

Photons participate in many atomic and molecular interactions and processes. Recent biophysical research has discovered an ultraweak radiation in biological tissues. It is now recognized that plants, animal and human cells emit this very weak biophotonic emission which can be readily measured with a sensitive photomultiplier system. UVA laser induced biophotonic emission of cultured cells was used in this report with the intention to detect biophysical changes between young and adult fibroblasts as well as between fibroblasts and keratinocytes. With suspension densities ranging from 1-8 x 106 cells/ml, it was evident that an increase of the UVA-laser-light induced photon emission intensity could be observed in young as well as adult fibroblastic cells. By the use of this method to determine ultraweak light emission, photons in cell suspensions in low volumes (100 microl) could be detected, in contrast to previous procedures using quantities up to 10 ml. Moreover, the analysis has been further refined by turning off the photomultiplier system electronically during irradiation leading to the first measurements of induced light emission in the cells after less than 10 micros instead of more than 100 milliseconds. These significant changes lead to an improvement factor up to 106 in comparison to classical detection procedures. In addition, different skin cells as fibroblasts and keratinocytes stemming from the same donor were measured using this new highly sensitive method in order to find new biophysical insight of light pathways. This is important in view to develop new strategies in biophotonics especially for use in alternative therapies.

Corresponding measurements of delayed luminescence and impedance spectroscopy on acupuncture points.

Measurements of impedance spectroscopy and delayed luminescence have been performed on the acupuncture points PC4 and PC8 and other two control points of ten volunteers. The results show that there is a highly significant difference between the imaginary parts of the impedance of the acupunctural points and that of the control points. The same difference has not be observed in the values of the total number of counts of delayed luminescence. However a relationship has been detected between the imaginary part of the impedance of PC8 point and the total number of delayed luminescence counts, similar to that one found before for collagen, and it has been seen that the temporal dynamics of the two phenomena measured on one control points are similar. In particular, these final results confirm the close connection between the delayed luminescence and the dielectric properties of biological tissues.

Single Photon Avalanche Diodes: Towards the Large Bidimensional Arrays.

Single photon detection is one of the most challenging goals of photonics. In recent years, the study of ultra-fast and/or low-intensity phenomena has received renewed attention from the academic and industrial communities. Intense research activity has been focused on bio-imaging applications, bio-luminescence, bio-scattering methods, and, more in general, on several applications requiring high speed operation and high timing resolution. In this paper we present design and characterization of bi-dimensional arrays of a next generation of single photon avalanche diodes (SPADs). Single photon sensitivity, dark noise, afterpulsing and timing resolution of the single SPAD have been examined in several experimental conditions. Moreover, the effects arising from their integration and the readout mode have also been deeply investigated.