Whey valorisation: A complete and novel technology development for dairy industry starter culture production

Whey is the major by-product of the dairy industry, produced in large quantities and usually disposed off causing major environmental pollution, due to its high organic load that makes treatment cost prohibitive. This paper comprises a contribution on the valorisation of this high polluting liquid waste of the dairy industry, based on research for the production of novel dairy starter cultures using whey as raw material. Starter cultures are used for cheese ripening in order to: (i) accelerate ripening, (ii) improve quality and (iii) increase shelf-life. The developed technology involves biomass production from whey followed by thermal drying of cultures. Specifically, Kluyveromyces marxianus, Lactobacillus bulgaricus and kefir yeasts were thermally dried, and their efficiency in lactose and milk whey fermentations was studied. The most suitable culture regarding its technological properties was kefir, which was used for cheese ripening in freeze-dried and thermally dried form. Besides the reduction of production cost, which is an essential requirement for the food industry, the use of thermally dried kefir displayed several other advantages such as acceleration of ripening, increase of shelf-life, and improvement of hard-type cheese quality.     

Do long-term changes in sea surface temperature at the breeding areas affect the breeding dates and reproduction performance of Mediterranean loggerhead turtles? Implications for climate change

Global climate change is likely to have an important influence on the phenology, behaviour and population dynamics of many species. We investigate climatic related changes in the breeding phenology of Mediterranean loggerhead marine turtles Caretta caretta over a 19 year period and the potential relationship between these changes and reproductive success and performance. We found that the studied population has experienced fluctuating sea surface temperatures (SST) with an increasing trend during the last century. With increasing spring SST there is a trend towards earlier nesting. However, there is no significant relationship between SST and nesting season, defined as the duration between the first recorded emergence and the last nest laid. Our analyses indicate that marine turtles display phenological changes, and thus maintain favorable thermal conditions at the nesting sites. Furthermore, increasing spring SST was correlated with decreasing clutch size and increasing hatching success that resulted in an apparent lack of correlation between SST and hatchling production. This apparent independence might be misleading since it only holds for a limited range of SST values. Thus, if we estimate the effect of climate change on loggerhead population growth as neutral, based on the apparent independence between SST and total number of hatchlings, we will be underestimating the population extinction risk.     

Synthesis of bicyclic molecular scaffolds (BTAa): an investigation towards new selective MMP-12 inhibitors

Starting from 3-aza-6,8-dioxa-bicyclo[3.2.1]octane scaffold (BTAa) a virtual library of molecules was generated and screened in silico against the crystal structure of the Human Macrophage Metalloelastase (MMP-12). The molecules obtaining high score were synthesized and the affinity for the catalytic domain of MMP-12 was experimentally proved by NMR experiments. A BTAa scaffold 20 having a N-hydroxyurea group in position 3 and a p-phenylbenzylcarboxy amide in position 7 showed a fair inhibition potency (IC50 = 149 microM) for MMP-12 and some selectivity towards five different MMPs. These results, taken together with the X-ray structure of the adduct between MMP-12, the inhibitor 20 and the acetohydroxamic acid (AHA), suggest that bicyclic scaffold derivatives may be exploited for the design of new selective matrix metalloproteinase inhibitors (MMPIs).     

A chimeric satellite transgene sequence is inefficiently targeted by viroid-induced DNA methylation in tobacco

In plants, transgenes containing Potato spindle tuber viroid (PSTVd) cDNA sequences were efficient targets of PSTVd infection-mediated RNA-directed DNA methylation. Here, we demonstrate that in PSTVd-infected tobacco plants, a 134 bp PSTVd fragment (PSTVd-134) did not become densely methylated when it was inserted into a chimeric Satellite tobacco mosaic virus (STMV) construct. Only about 4–5% of all cytosines (Cs) of the PSTVd-134 were methylated when flanked by satellite sequences. In the same plants, C methylation was approximately 92% when the PSTVd-134 was in a PSTVd full length sequence context and roughly 33% when flanked at its 3′ end by a 19 bp PSTVd and at its 5′ end by a short viroid-unrelated sequence. In addition, PSTVd small interfering RNAs (siRNAs) produced from the replicating viroid failed to target PSTVd-134-containing chimeric STMV RNA for degradation. Satellite RNAs appear to have adopted secondary structures that protect them against RNA interference (RNAi)—mediated degradation. Protection can be extended to short non-satellite sequences residing in satellite RNAs, rendering them poor targets for nuclear and cytoplasmic RNAi induced in trans.     

Production of human osteoclasts in a three-dimensional bone marrow culture system

Osteoclasts, which are derived from hemopoietic stem cells, play important roles in bone remodeling and resorption. Osteoclast development is critically dependent on the bone microenvironment. We have developed a novel human ex vivo bone marrow model that mimics bone marrow both structurally and functionally by providing artificial scaffolding, thus obtaining a three-dimensional growth configuration with high cell density and intimate physical contact between hemopoietic and stromal cells. In this study, we utilized the 3-D culture system to produce multinucleated cells (MNCs) from human bone marrow cells cultured in the presence of Vitamin D₃ and the absence of hydrocortisone and any exogenous growth factors. These multinucleated cells had the phenotypic and functional characteristics of osteoclasts as determined by their morphology, expression of tartarate resistant acid phosphatase (TRAP), and the ability to resorb bone. Scoring of the resorption pits revealed a dose response to Vitamin D₃ stimulation with 10⁻⁷ M being the optimal concentration. Furthermore, addition of parathyroid hormone (10⁻⁸ M) resulted in an up-to three-fold enhancement of bone resorption. These findings suggest that the 3-D culture system represent a physiologically relevant model to study osteoclastogenesis and elucidate the molecular and cellular signals associated with this process.     

Application of the OECD 301F respirometric test for the biodegradability assessment of various potential endocrine disrupting chemicals

The biodegradability of several potential endocrine disrupting compounds, namely 4-n-nonylphenol (4-n-NP), nonylphenol monoethoxylate (NP1EO), nonylphenol diethoxylate (NP2EO), bisphenol A (BPA), triclosan (TCS), di-(2-ethylhexyl)-phthalate (DEHP), perfluorooctanoate (PFOA) and perfluorononanoate (PFNA) was evaluated in this study, using OECD method 301F (manometric respirometry test) and activated sludge as inoculum. According to the results, 4-n-NP and BPA meet the strict definition of ready biodegradability and they are not expected to be persistent during the activated sludge process. Partial biodegradation was observed for DEHP (58.7+/-5.7%, n=3), TCS (52.1+/-8.5%, n=3) and NP1EO (25.9+/-8.1%, n=3), indicating their possible biodegradation in wastewater treatment systems, while no biodegradation was observed for NP2EO, PFOA and PFNA. Experiments in the co-presence of a readily biodegradable compound showed the absence of co-metabolic phenomena during 4-n-NP, BPA and TCS biodegradation. Using first order kinetics to describe biodegradation of the target compounds, half-lives of 4.3+/-0.6, 1.3+/-0.2, 1.8+/-0.5, 6.9+/-2.6 days were calculated for 4-n-NP, BPA, TCS and DEHP, respectively. Toxicity tests using marine bacterium Vibrio fischeri showed that biodegradation of 4-n-NP, NP1EO, BPA and TCS is a simultaneous detoxification process, while possible abiotic or biotic transformations of NP2EO, DEHP, PFOA and PFNA during respirometric test resulted to significant increase of their toxicities.     

Comparative study of the formation of oxidative damage marker 8-hydroxy-2'-deoxyguanosine (8-OHdG) adduct from the nucleoside 2'-deoxyguanosine by transition metals and suspensions of particulate matter in relation to metal content and redox reactivity

An association between exposure to ambient particulate matter (PM) and increased incidence of mortality and morbidity due to lung cancer and cardiovascular diseases has been demonstrated by recent epidemiological studies. Reactive oxygen species (ROS), especially hydroxyl radicals, generated by PM, have been suggested by many studies as an important factor in the oxidative damage of DNA by PM. The purpose of this study was to characterize quantitatively hydroxyl radical generation by various transition metals in the presence of H₂O₂ in aqueous buffer solution (pH 7.4) and hydroxylation of 2'-deoxyguanosine (dG) to 8-hydroxy-2'-deoxyguanosine (8-OHdG) under similar conditions. The order of metals' redox reactivity and hydroxyl radical production was Fe(II), V(IV), Cu(I), Cr(III), Ni(II), Co(II), Pb(II), Cd(II). Then, we investigated the generation of hydroxyl radicals in the presence of H₂O₂ by various airborne PM samples, such as total suspended particulate (TSP), PM₁₀, PM_2.5 (PM with aerodynamic diameter 10 and 2.5 μm), diesel exhaust particles (DEP), gasoline exhaust particles (GEP) and woodsmoke soot under the same conditions. When suspensions of PMs were incubated with H₂O₂ and dG at pH 7.4, all particles induced hydroxylation of dG and formation of 8-OHdG in a dose-dependent increase. Our findings demonstrated that PM's hydroxyl radical (HO√) generating ability and subsequent dG hydroxylation is associated with the concentration of water-soluble metals, especially Fe and V and other redox or ionizable transition metals and not their total metal content, or insoluble metal oxides, via a Fenton-driven reaction of H₂O₂ with metals. Additionally, we observed, by Electron paramagnetic resonance (EPR), that PM suspensions in the presence of H₂O₂ generated radical species with dG, which were spin-trapped by 2-methyl-2-nitroso-propane (MNP).     

Retinoid receptor cross-talk in respiratory epithelium cancer chemoprevention

The rationale for using retinoids in the prevention of respiratory epithelium cancers is based on their ability to coordinately regulate differentiation, proliferation and apoptosis. The complex retinoid signaling pathways and their cross-reactions are modulated by multiple mechanisms that are gradually being elucidated. It is possible that significant molecular changes take place during the very early stages of respiratory epithelial carcinogenesis, which enable cancer cells to escape apoptosis and result in unimpeded proliferation. Here, we propose that a "switch on/off" model dictates the cross-talk between retinoid receptors and other signal transducing pathways during respiratory epithelium carcinogenesis. This model might contribute to the development of novel selective retinoids and their clinical evaluation in combinatorial chemopreventive strategies.