Development of therapeutic contact lenses using a supercritical solvent impregnation method

We present some selected results indicating the feasibility of preparing therapeutic finished ophthalmic articles, namely commercially available soft contact lenses, using a supercritical solvent impregnation (SSI) technique. Several commercial soft contact lenses were tested and, among these, four lenses were selected for more complete studies: Nelfilcon A (FocusDailies^®, CIBA Vision), Omafilcon A (Proclear^® Compatibles, CooperVision), Methafilcon A (Frequency^® 55, CooperVision) and Hilafilcon B (SofLens^® 59 Comfort, Bausch & Lomb). Supercritical carbon dioxide (scCO₂) was the chosen supercritical fluid and two ophthalmic drugs were tested: flurbiprofen (a NSAID, hydrophobic) and timolol maleate (an anti-glaucoma drug, hydrophilic). The effects of operational pressure, of impregnation duration and of the addition of a cosolvent (ethanol) were studied on the overall drug loading yields. Depending on the experiment, we employed pressures from 9 up to 16 MPa and impregnation times from 30 up to 180 min. Temperature was kept constant and equal to 313 K. The employed depressurization rates were kept low and between 0.1 and 0.2 MPa/min.Results are discussed in terms of the employed operational conditions and taking in consideration all the possible interactions between supercritical fluids, drugs, cosolvents and the polymers which compose the employed hydrogel contact lenses. In vitro drug release experiments were carried out in order to evaluate the resulting drug release profiles. Obtained results were also compared with drug-loaded contact lenses obtained by conventional drug “soaking” in aqueous solutions. Results also proved that SSI can be considered as a viable, efficient and safe alternative for the impregnation of drugs, including those of hydrophobic character or presenting low aqueous solubility, into commercial soft contact lenses. SSI proved to be a “tunable” process since the variation of the employed operational conditions indicated that it is possible to control the amount of impregnated drug. In the end, the ophthalmic articles were recovered undamaged and without the presence of harmful solvent residues. This method also permits to process already prepared commercial contact lenses, without interfering with their manufacture methods and, after processing, store them for future use.     

Therapeutic Strategies for IVD Regeneration through Hyaluronan/SDF-1-Based Hydrogel and Intravenous Administration of MSCs

Intervertebral disc (IVD) degeneration involves a complex cascade of events, including degradation of the native extracellular matrix, loss of water content, and decreased cell numbers. Cell recruitment strategies for the IVD have been increasingly explored, aiming to recruit either endogenous or transplanted cells. This study evaluates the IVD therapeutic potential of a chemoattractant delivery system (HAPSDF5) that combines a hyaluronan-based thermoreversible hydrogel (HAP) and the chemokine stromal cell derived factor-1 (SDF-1). HAPSDF5 was injected into the IVD and was combined with an intravenous injection of mesenchymal stem/stromal cells (MSCs) in a pre-clinical in vivo IVD lesion model. The local and systemic effects were evaluated two weeks after treatment. The hydrogel by itself (HAP) did not elicit any adverse effect, showing potential to be administrated by intradiscal injection. HAPSDF5 induced higher cell numbers, but no evidence of IVD regeneration was observed. MSCs systemic injection seemed to exert a role in IVD regeneration to some extent through a paracrine effect, but no synergies were observed when HAPSDF5 was combined with MSCs. Overall, this study shows that although the injection of chemoattractant hydrogels and MSC recruitment are feasible approaches for IVD, IVD regeneration using this strategy needs to be further explored before successful clinical translation.     

Unravelling the origin of Vitis vinifera L. Verdelho

Background and Aims

The history of Vitis vinifera L. cv. Verdelho has been studied, however, its geographic origin remains unknown. Although not widespread in mainland Portugal, Verdelho has gained a reputation as a Madeira wine, which has led most authors to attribute its origin to the Madeira Islands. The objective of this work was to determine the origin of Verdelho.

Methods and Results

An integrated strategy was followed, combining the molecular analysis of 28 nuclear and three chloroplast DNA microsatellites with an historical literature search. Historical records of the presence of Verdelho in the Azores Islands were found and these preceded those of the Madeira Islands by about 200 years. Moreover, the genetic analysis supports Verdelho as the progenitor of the Azores cultivars, Arinto dos Açores and Terrantez do Pico, and as the progeny of cv. Savagnin.

Conclusions

Verdelho is a key cultivar in the Azorean genetic diversity pool, sharing first‐degree relationships with the two most important autochthonous grape cultivars, and reinforcing the historical data of its ancestry in the region.

Significance of the Study

The results presented here, both historical and genetic, shed new light on the origin of Verdelho, and suggest that the Azores Islands is the site where this cultivar was likely to have originated.     

Advances and perspectives in using microalgae to produce biodiesel

Carbon-neutral renewable liquid biofuels are needed to displace petroleum-derived transport fuels in the near future – which contribute to global warming and are of a limited availability. A promising alternative is conveyed by microalgae, the oil content of which may exceed 80% (w/w_DW) – as compared with 5% of the best agricultural oil crops. However, current implementation of microalga-based systems has been economically constrained by their still poor volumetric efficiencies – which lead to excessively high costs, as compared with petrofuel prices. Technological improvements of such processes are thus critical – and this will require a multiple approach, both on the biocatalyst and bioreactor levels. Several bottlenecks indeed exist at present that preclude the full industrial exploitation of microalgal cells: the number of species that have been subjected to successful genetic transformation is scarce, which hampers a global understanding (and thus a rational design) of novel blue-biotechnological processes; the mechanisms that control regulation of gene expression are not fully elucidated, as required before effective bioprocesses based on microalgae can be scaled-up; and new molecular biology tools are needed to standardize genetic modifications in microalgae – including efficient nuclear transformation, availability of promoter or selectable marker genes, and stable expression of transgenes. On the other hand, a number of pending technological issues are also present: the relatively low microalga intrinsic lipid productivity; the maximum cell concentration attainable; the efficiency of harvest and sequential recovery of bulk lipids; and the possibility of by-product upgrade. This review briefly covers the state of the art regarding microalgae toward production of biofuels, both from the point of view of the microalgal cell itself and of the supporting bioreactor; and discusses, in a critical manner, current limitations and promising perspectives in this field.     

The competitive adsorption of human proteins onto natural-based biomaterials

This study aims to further the understanding of nanoscale structures relevant for cellular recognition on contact and interaction with natural-based materials. The correlation between surface characteristics and protein adsorption from unitary and complex protein systems was investigated with respect to altering the bulk chemistry of the substrate material. Polymeric blends of starch and cellulose acetate, polycaprolactone (SPCL) and ethylene vinyl alcohol (SEVA-C) were used. Different proteins, bovine serum albumin, human serum albumin (HSA) and human fibronectin (HFN), were selected for this study. The construction of adsorption isotherms is an important starting point towards characterizing the interactions between surfaces and proteins. In this study, albumin adsorption isotherms fit the Freundlich model and were correlated with the chemistry and morphology of surfaces. In addition, protein distribution, quantification and competition were measured using fluorimetry and visualized by confocal microscopy. The analysis of unitary systems demonstrated that the adsorption of HSA was generally lower than that of HFN. In the latter case, SPCL and SEVA-C blends reached adsorption values of 97 and 89 per cent, respectively. In studying the co-adsorption of proteins, an increase in both HSA and HFN on SEVA-C surfaces was observed. SPCL showed no substantial increase in the adsorption of the proteins in competitive conditions. The similarity of these materials with other polysaccharide-based materials increases the relevance of the presented results. This study provides valuable information for the development of strategies towards the control of protein orientation and functionality as the availability of cell signalling epitopes for a broader family of materials that continue to be a significant component of this field of research.     

Infrared nanosecond laser effects on the formation of copper nanoparticles

For the first time copper nanoparticles were formed inside Cu ion-doped glass after infrared irradiation by a nanosecond laser and sucessive annealing at 700 °C for 1 h. The surface plasmon resonance was observed in the absorption spectra of copper nanoparticles. Their spectral position is in good agreement with the values predicted by Mie theory for the copper nanoparticles. The possible mechanisms responsible by the observed results are discussed.     

High Pdr12 levels in spoilage yeast (Saccharomyces cerevisiae) correlate directly with sorbic acid levels in the culture medium but are not sufficient to provide cells with acquired resistance to the food preservative

Sorbic acid is a commonly used food preservative against yeast and fungal food spoilage. Understanding its effect on the molecular physiology of yeast cells will allow the food industry to develop knowledge-based strategies to make more optimal use of its preservative action. Here we show that the yeast membrane protein Pdr12, previously shown to be prominently involved in sorbic acid resistance development in laboratory strains, was strongly induced by the presence of sorbic acid in the culture medium in Saccharomyces strains isolated from spoiled foods. Induction of Pdr12 expression was seen both under laboratory conditions and upon growth in a commercial soft drink. Induction was rapid and maintained for the duration of the stress. No Pdr12-like protein induction was seen in Zygosaccharomyces bailii or Zygosaccharomyces lentus, two well-known beverages spoilage organisms. Finally, unexpectedly, our studies showed for the first time that pre-inducing Pdr12p to maximal levels by subjecting cells to a mild sorbic acid stress did not lead to cells with an acquired resistance. Neither more rapid growth in the presence of the acid nor growth at higher sorbic acid concentrations at a given environmental pH was observed. Thus we have shown that while important in resistance development against sorbic acid, by itself induction of the pump is not sufficient to acquire resistance to the preservative.     

The effect of recirculating aquaculture systems on the concentrations of heavy metals in culture water and tissues of Nile tilapia Oreochromis niloticus

To date, farming fish in recirculating aquaculture systems (RAS) is one of the most environmentally friendly ways of producing fish. However, with the trend towards intensification, and consequently decrease in water exchange rates, these systems may accumulate substances, such as heavy metals, in the water and fish. Inductively-coupled plasma-optical emission spectrometry (ICP-OES) and inductively coupled plasma-mass spectroscope (ICP-MS) were used to determine Al, As, Cd, Cr, Cu, Fe, Mn, Ni, Pb and Zn, in the water and fish (Nile tilapia Oreochromis niloticus). Three RAS were used, differing in daily water exchange rates (30, 70 and 1500 l/kg feed/d). The concentrations of As, Fe, Mn, Ni and Zn in the water increased with decreasing water exchange rates, suggesting an accumulation of heavy metals as more water was re-used. Such accumulation in the water was, however, not translated into accumulation in the liver and muscle. Accumulation of heavy metals was always higher in the liver than in the muscle; however, As reached 1.61 mg/kg wet weight in the muscle of fish farmed in RAS-70 l/kg feed/d. However, these levels are considerably lower than permissible safety levels for human consumption.