José R. Fernandes

New technique to mechanically induce long-period fibre gratings

A full characterization of reversible long-period fibre gratings produced through the wind of a string around a fibre/ grooved tube set is presented. This technique enables a good control over the gratings isolation loss-peaks and has high repeatability. The tunability of the resonant wavelengths through the etching of the fibres cladding, prior to perform the gratings, have been investigated. These mechanically induced long-period fibre gratings have shown high sensitivity to loads.

Evanescent wave DNA-aptamer biosensor based on long period gratings for the specific recognition of E. coli outer membrane proteins

An evanescent wave fiber optic sensor for detection of Escherichia coli (E. coli) outer membranes proteins (EcOMPs) using long period gratings (LPGs) as a refractometric platform is presented. The sensing probes were attained by the functionalization of LPGs inscribed in single mode fiber using two different methods of immobilization; electrostatic assembly and covalent binding. The resulting label-free configuration enabled the specific recognition of EcOMPs in water by monitoring the resonance wavelength shift due to refractive index changes induced by binding events. The sensors displayed linear responses in the range of 0.1 nM to 10 nM EcOMPs with sensitivities of -0.1563±0.005 nm decade(-1) [EcOMP, M] (electrostatic method) and -0.1597±0.004 nm decade(-1) [EcOMP, M] (covalent method). The devices could be regenerated (under low pH conditions) with a deviation less than 0.1% for at least three subsequent detection events. The sensors were also applied to spiked environmental water samples.

Treatment of crystallized-fruit wastewater by UV-A LED photo-Fenton and coagulation-flocculation.

This work reports the treatment of crystallized-fruit effluents, characterized by a very low biodegradability (BOD5/COD <0.19), through the application of a UV-A LED photo-Fenton process. Firstly, a Box-Behnken design of Response Surface Methodology was applied to achieve the optimal conditions for the UV-A LED photo-Fenton process, trying to maximize the efficiency by saving chemicals and time. Under the optimal conditions ([H2O2] = 5459 mg/L; [Fe(3+)] = 286 mg/L; time >180 min), a COD removal of 45, 64 and 74% was achieved after 360 min, using an irradiance of 23, 70 and 85 W/m(2) respectively. Then a combination of UV-A LED photo-Fenton with coagulation-flocculation-decantation attained a higher COD removal (80%), as well as almost total removal of turbidity (99%) and total suspended solids (95%). Subsequent biodegradability of treated effluents increased, allowing the application of a biological treatment step after the photochemical/CFD with 85 W/m(2).

Biosensor for label-free DNA quantification based on functionalized LPGs.

A label-free fiber optic biosensor based on a long period grating (LPG) and a basic optical interrogation scheme using off the shelf components is used for the detection of in-situ DNA hybridization. A new methodology is proposed for the determination of the spectral position of the LPG mode resonance. The experimental limit of detection obtained for the DNA was 62±2nM and the limit of quantification was 209±7nM. The sample specificity was experimentally demonstrated using DNA targets with different base mismatches relatively to the probe and was found that the system has a single base mismatch selectivity.

High Resolution Melting (HRM) applied to wine authenticity

Wine authenticity methods are in increasing demand mainly in Denomination of Origin designations. The DNA-based methodologies are a reliable means of tracking food/wine varietal composition. The main aim of this work was the study of High Resolution Melting (HRM) application as a screening method for must and wine authenticity. Three sample types (leaf, must and wine) were used to validate the three developed HRM assays (Vv1-705bp; Vv2-375bp; and Vv3-119bp). The Vv1 HRM assay was only successful when applied to leaf and must samples. The Vv2 HRM assay successfully amplified all sample types, allowing genotype discrimination based on melting temperature values. The smallest amplicon, Vv3, produced a coincident melting curve shape in all sample types (leaf and wine) with corresponding genotypes. This study presents sensitive, rapid and efficient HRM assays applied for the first time to wine samples suitable for wine authenticity purposes.

Pulsed laser deposition of SiO2 - P2O5 - CaO - MgO glass coatings on titanium substrates

Thin films of bioactive glass-ceramic have been deposited on titanium substrates by the Pulsed Laser Deposition (PLD) technique under different experimental conditions. The effect of parameters such as deposition pressure and temperature of heat treatments was studied. The microstructure and the crystalline phases of the coatings were characterized using SEM, EDX and XRD analysis; the phases present were titanium oxides, calcium magnesium silicates and phosphates. The adhesion of the as-deposited films has been examined by scratch tests. The interfacial adhesion of the coatings was better when the deposition was performed at low pressure. Samples were immersed in simulated body fluid (SBF), and a calcium-phosphate precipitate was observed on the surface of less crystallized samples, suggesting that there is some relationship between surface reactivity and crystallinity.

Simple Method for Crystallizing Ceramic Thin Films Using Platinum Bottom Electrodes as Resistive Heating Elements

A simple method for crystallizing amorphous thin films was developed using platinum bottom electrodes as heating elements. A current was applied to chromel wires in contact with the platinum and the temperature was measured using a type K thermocouple. A proportional feedback algorithm was used for controlling the process. The optimum parameters for the temperature control were determined and applied to the crystallization of PZT thin films. The results obtained for a heat treatment at 650°C with a 15°C/s heating and cooling rate showed a pure perovskite phase; the ferroelectric properties were comparable with those from conventional heat treatment methods.

Applying high-resolution melting (HRM) technology to olive oil and wine authenticity

Olive oil and wine production have a worldwide economic impact. Their market reliability is under great concern because of the increasing number of fraud and adulteration attempts. The need for a traceability system in all its extension is crucial particularly for the cases of olive oils and wines with certified labels, in which only a limited number of olives and grapevine varieties, respectively, are allowed in a restricted well-defined geographical area. Molecular markers have been vastly applied to the food sector, and in particular High-Resolution DNA Melting technology has been successfully applied for olive oil and wine authentication, as part of the traceability system. In this review, the applications of HRM and their usefulness for this sector considering, Safety, Security and Authenticity will be reviewed. A broad overview of the HRM technique will be presented, focusing on the aspects that are crucial for its success, in particular the new generation of fluorescent dsDNA dyes used for amplicon detection and quantification, and the data analysis. A brief outlook on the olive oil and wine authenticity procedures, based on new DNA technology advances, and in which way this may influence the future establishment of a traceability system will be discussed.

Synthesis, spectroscopic characterization and biological evaluation of unsymmetrical aminosquarylium cyanine dyes

New unsymmetrical aminosquarylium cyanine dyes were synthesized and their potential as photosensitizers evaluated. New dyes, derived from benzothiazole and quinoline, were prepared by nucleophilic substitution of the corresponding O-methylated, the key intermediate that was obtained by methylation with CF3SO3CH3 of the related zwitterionic unsymmetrical dye, with ammonia and methylamine, respectively. All three news dyes herein described displayed intense and narrow bands in the Vis/NIR region (693-714nm) and their singlet oxygen formation quantum yields ranged from 0.03 to 0.05. In vitro toxicity, in Caco-2 and HepG2 cells, indicated that dark toxicity was absent for concentrations up to 5µM (for the less active dye) or up to 1µM (for the two more active dyes). The three dyes present potential as photosensitizers, differing in irradiation conditions and period of incubation in the presence of irradiated dye. The less active dye needs a longer irradiation period to exhibit phototoxicity which is only evident after longer period of contact with cells (24h). However, the remaining two more active dyes produce higher phototoxicity, even at shorter incubation periods (1h), with shorter irradiation time (7min). Although in different extents, these dyes show promising in vitro results as photosensitizers.

Alternative SNP detection platforms, HRM and biosensors, for varietal identification in Vitis vinifera L. using F3H and LDOX genes

The wine sector requires quick and reliable methods for Vitis vinifera L. varietal identification. The number of V. vinifera varieties is estimated in about 5,000 worldwide. Single Nucleotide Polymorphisms (SNPs) represent the most basic and abundant form of genetic sequence variation, being adequate for varietal discrimination. The aim of this work was to develop DNA-based assays suitable to detect SNP variation in V. vinifera, allowing varietal discrimination. Genotyping by sequencing allowed the detection of eleven SNPs on two genes of the anthocyanin pathway, the flavanone 3-hydroxylase (F3H, EC: 1.14.11.9), and the leucoanthocyanidin dioxygenase (LDOX, EC 1.14.11.19; synonym anthocyanidin synthase, ANS) in twenty V. vinifera varieties. Three High Resolution Melting (HRM) assays were designed based on the sequencing information, discriminating five of the 20 varieties: Alicante Bouschet, Donzelinho Tinto, Merlot, Moscatel Galego and Tinta Roriz. Sanger sequencing of the HRM assay products confirmed the HRM profiles. Three probes, with different lengths and sequences, were used as bio-recognition elements in an optical biosensor platform based on a long period grating (LPG) fiber optic sensor. The label free platform detected a difference of a single SNP using genomic DNA samples. The two different platforms were successfully applied for grapevine varietal identification.