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The modern two-phase technology of the Leopard series for olive oil industrial extraction developed by the Pieralisi Group (Pieralisi Maip S.p.A., Jesi, AN, Italy) generates large quantities of a novel by-product (olive paste - OP) constituted of the partially defatted wet drupe pulp without any traces of the kernel. OP is rich in lipophilic and hydrophilic molecules able to exert beneficial effects on human health, including triterpenic acids, phytosterols, tocopherols, carotenoids, polyphenols, minerals and fibers [1]. In the perspective of by-product valorization through a modern biorefinery approach, it is worth noting that dried OP contains more than 10% oil characterized by a well-balanced lipid profile, rich in mono and polyunsaturated fatty acids, and a very good oxidative stability, due to the high concentration of fat-soluble antioxidants. This makes OP oil particularly suitable as functional ingredient for food/feed industry, as well as for the formulation of nutraceutical, cosmeceutical and pharmaceutical products. Supercritical carbon dioxide (SC-CO2) is a green and environmentally friendly technology for the effective extraction of high-value natural molecules. It is gaining a foothold in industrial production of solvent-free vegetable oils and has been also applied to concentrate oil products of lipophilic micronutrients (e.g. vitamin E in soybean and olive oils) [2]. Nevertheless, the industrial application of SC-CO2 technology requires a careful optimization of the operative parameters to make the process efficient from both an economic and a productive point of view. In this work, a response surface methodology approach, based on the Box-Behnken Design, was used to determine the optimal parameters of pressure, temperature and time to simultaneously maximize oil production from the freeze-dried OP of cultivar Leccino and concentrate the most abundant lipophilic bioactives. The experimental data were fitted to a second-order polynomial equation by multiple regression analysis, and examined using appropriate statistical methods. The 3-D response surface plots derived from the mathematical models were applied to determine the optimal extraction parameters, which resulted: temperature 70 °C, pressure 35.5 MPa and time 62 min. Under these conditions, the oil experimental yield was 14.0±0.7%, in close agreement with the predicted value. Compared to the freeze-dried OP, SC-CO2 extracted oil was much concentrated in phytosterols (~13 fold), tocopherols (~6 fold) and squalene (~8 fold); total carotenoid concentration remained almost unchanged, while triterpenic acids, being polar, were substantially reduced (- 79.5%). However, their total content was much higher than in extra virgin olive oil. The biological effects produced by OP oil intake were examined on 3 groups of healthy BALB/c mice (n=5/group) orally administered (for 4 weeks) with 20 or 40 mg/die of OP oil. Untreated animals were used as control. OP oil supplementation had no significant effects neither on body weight, nor on the activity of most liver cytoprotective enzymes (glutathione-disulfide reductase, glutathione S-transferase and glucose-6-phosphate dehydrogenase) with the exception of NAD(P)H:Quinone Oxidoreductase 1, whose activity was significantly increased (P

Le malattie mitocondriali, conosciute anche come mitocondriopatie, sono un gruppo eterogeneo di patologie ereditarie che, pur pre- sentando una notevole variabilità clinica relativamente all’età di insorgenza, il tipo di evoluzione ed i tessuti coinvolti, sono caratterizzate dalla presenza di alterazioni nel funzionamento dei mitocondri. L’alterato funzionamento dei mitocondri determina, nei tessuti interessati, la comparsa di un deficit energetico, in quanto questi organelli sono responsabili della sintesi di ATP mediante il processo della fosforilazione ossidativa (OXPHOS). Ad essere maggiormente colpiti sono quindi i tessuti che presentano una maggiore richiesta energetica, come il muscolo ed il cervello. Per questo motivo, le malattie mitocondriali sono spesso definite “encefalomiopatie”.

Negli ultimi anni, l’aumento della prevalenza dell’obesità è stato correlato a un decremento del potenziale riproduttivo nell’uomo. Sempre più evidenze scientifiche suggeriscono che la supplementazione della dieta con acidi grassi abbia un impatto sulla qualità e la funzionalità degli spermatozoi. In particolare, è stato osservato che diete ricche di SFA e povere di PUFA compromettono la qualità spermatica, mentre la supplementazione con PUFA ne determina un miglioramento. A tal proposito, utilizzando modelli animali nei quali è stato possibile indurre una condizione di obesità mediante la somministrazione di diete iperlipidiche, recenti studi hanno dimostrato che la somministrazione di MUFA è in grado di contrastare gli effetti negativi indotti da una dieta ad alto contenuto in grassi, aumentando la motilità dei gameti, riducendo lo stress ossidativo e migliorando l’efficienza del processo di respirazione mitocondriale. La somministrazione di PUFA, soprattutto quelli della serie n-3, invece, ha determinato un miglioramento ancora più marcato della qualità spermatica, associato ad un aumento della concentrazione e della motilità degli spermatozoi, ad un incremento delle attività degli enzimi mitocondriali coinvolti nel metabolismo energetico degli spermatozoi e a una riduzione del danno ossidativo. Questi risultati contribuiscono, da un alto, a chiarire i meccanismi molecolari mediante i quali MUFA e PUFA agiscono su alterazioni metaboliche causate dalla somministrazione di una dieta ricca di grassi, che spesso caratterizza le abitudini alimentari delle popolazioni occidentali. D’altro canto, essi suggeriscono che gli acidi grassi introdotti con la dieta sono in grado di influenzare positivamente la qualità e la funzionalità degli spermatozoi, fornendo un nuovo e interessante approccio per il trattamento dell’infertilità maschile legata all’obesità.

Up to now the content of the organic halides in sediments was measured as extractable organic halides compounds by different methods including microcoulometric titration one. We describe a procedure for microcoulometric determination of total organic halide pollutants by using a direct combustion of the sediments into the microcoulometric system furnace. An accelerated removal of the inorganic chlorides by reaction with potassium nitrate using ultrasonic radiation was attained. The procedure was validated for analysis of bottom sediments from natural and anthropogenic sources. The repeatability RSD = 8.5%, the expanded relative uncertainty U (n = 7, P = 95%, k = 2) = 6.4%, the reproducibility RSD was within the range 10.0–7.3%, the average recovery R = 97.6% and method LOD = 11 mg kg−1 Cl.

In the Mediterranean Sea, the biological invasion by the green algae Caulerpa racemosa var. cylindracea is maybe the most striking marine sea-bottom landscape change of the last decades. In this paper, we have synthesized the results of 5 years of studies about the C. cylindracea effects on fish of ecological and economic importance, such as the white sea bream Diplodus sargus. We found that secondary metabolite of C. cylindracea has the potential to enter in trophic chains and accumulate in fish tissues. Ecotoxicological analyses made on tissues of the D. sargus specimens showed alterations of some of main antioxidant systems, such as catalase, glutathione peroxidases and total glutathione, and a significant induction of P450 biotransformation system. We found also that the C. cylindracea based-diet can alter muscle fatty acids composition in the white sea bream by reducing the percentage of polyunsaturated fatty acids (PUFA) of the n-3 and n-6 series, such as eicosapentaenoic (EPA, C20:5), docosahexaenoic (DHA, C22:6) and arachidonic acids (AA, C20:4). On the whole, this work sheds light on an unexplored and critical aspect of biological invasions with implication on the health of both humans and the environment.

The precise and accurate determination of water in petroleum products—gasoline, diesel oil and aromatic hydrocarbons—is of significant importance for their normal functional and operational characteristics for economics as well as in meeting international requirements. The standard method ISO 12937 for the measurement of water by the coulometric Karl Fischer titration method was subject to within-laboratory assessment. Thereby it was established that the analytical procedure could not always be relied upon, especially when low levels of water were determined. The aim of this work is to assess each stage of the test procedure depending on the particular problems defined by the samples, using high-reactive Karl Fischer reagents and coulometric cells with and without a diaphragm, and to indicate the major factors causing the uncertainty. A reduced combined uncertainty was achieved by modifying the sample preparation step, the administration of the moisture in the coulometric cell, and sample injection manipulation. Experiments showed that the cell without the diaphragm ensures lower and uniform uncertainty of the measurements in the range of 0.01 to 10 mg H2O in comparison to the cell with a diaphragm and therefore is more appropriate for the determination of low water levels in the petroleum products. A procedure that utilized a mixture of the reagent and toluene (10:6) was able to resolve the problem of two phases of formation and reduced conductivity of the reagent in direct titration of diesel oils. The results suggest that the modified procedure ensures relative expanded uncertainty equal to or less than 2.0% (n=5, confidence interval close to 95%) and 99.4% recovery for petroleum products investigated in repeatability conditions.

Motivation: A basic problem of structural biochemistry studies is to which extent a mutation will affect the stability, and then the function of the protein. From this point of view, an important aspect regards the function of metabolite mitochondrial carriers, in relation to the role that such proteins cover in some mitochondrial pathologies. Sequence studies have shown that the PX(D/E)XX(K/R) signature is characteristic of all mitochondrial carriers, and possibly involved in the transition from the open to closed states, corresponding to the active/inactive state of the carrier. In this study our approach is to combine predictions and experimental validation adopting as a test case the dicarboxylate carrier (DIC) of S. cerevisiae. Because DIC structure is unknown, we integrated a routinely expert dependent strategy in a automatic tool based on a Grid infrastructure to facilitate the generation of carrier models, including site directed mutagenesis. Methods: Transport activity of mutagenized proteins was measured in vitro in reconstituted systems. In parallel in silico experiments protein stability was predicted by using I-Mutant3, available at http://gpcr2.biocomp.unibo.it/cgi/predictors/I-Mutant3.0/IMutant3.0. cgi. DIC structure was computed by homology modelling using a service that integrates several software and data involved in a Grid infrastructure, available at https://sara.unile.it/cgi-bin/bioinfo/enter. Results: Protein stability changes upon mutations can be both predicted and/or tested in vitro, by monitoring the function of mutagenized proteins in reconstituted systems. We found that mutations in the DIC carrier signature reduced or blocked nearly completely the protein transport activity, in agreement with the functional role of the carrier motif. Then the question is whether the observed effect on the activity can be also related to a change on protein stability upon mutation. This was tackled by computational methods. According to our predictions protein destabilization would correlate with the observed loss of protein activity. Our data therefore corroborate the finding that single point mutations may hamper protein stability when placed in functionally relevant structural position, and further add to the possibility of predicting a priori whether the mutation destabilize the protein.

In the Mediterranean Sea, the biological invasion by the green algae Caulerpa racemosa var. cylindracea is maybe the most striking marine sea-bottom landscape change of the last decades. In this paper, we have synthesized the results of 5 years of studies about the C. cylindracea effects on fish of ecological and economic importance, such as the white sea bream Diplodus sargus. We found that a secondary metabolite of C. cylindracea has the potential to enter in trophic chains and accumulate in fish tissues. Ecotoxicological analyses made on tissues of the D. sargus specimens showed alterations of some of main antioxidant systems, such as catalase, glutathione peroxidases and total glutathione, and a significant induction of P450 biotransformation system. We found also that the C. cylindracea based-diet can alter muscle fatty acids composition in the white sea bream by reducing the percentage of polyunsaturated fatty acids (PUFA) of the n-3 and n-6 series, such as eicosapentaenoic (EPA, C20:5), docosahexaenoic (DHA, C22:6) and arachidonic acids (AA, C20:4). On the whole, this work sheds light on an unexplored and critical aspect of biological invasions with implication on the health of both humans and the environment.

The International Laboratory of BioInformatics (LIBI - www.libi.it) aims at setting up of an advanced Bioinformatics and Computational Biology Laboratory, focusing on the central activities of basic and applied research in modern Biology and Biotechnologies. The main activities include the construction and the maintenance of general genomic, proteomic and transcriptomic databases (e.g. ENSEMBL) as well as specialized databases developed by LIBI partners (e.g. MitoRes, UTRdb, UTRSite, ASPicDB, CSTdb, etc.); the design and implementation of new algorithms and software for the analysis of genomes and their expression products and for the prediction of the structure of proteins; the development of a Grid Problem Solving Environment in order to exploit the distributed computational infrastructure.