Estudi sobre el ritme de restauració de les activitats extractives de Catalunya

L’objecte d’aquest projecte és determinar si el ritme restaurador de les activitats extractives és el mateix que el de l’extracció del mineral. Per avaluar-ho s’ha analitzat la dinàmica de les explotacions mineres que exploten els quatre tipus principals de recursos a Catalunya, en el període de 10 anys (1993-2003), amb l’ajuda de Sistemes d’Informació Geogràfica (SIG). Per dur a terme aquest estudi s’ha emprat bases cartogràfiques diferents: els Mapes de cobertes del Sòl de Catalunya (MCSC) i el de les activitats extractives (EXTCATA). A partir de l’anàlisi dels resultats, es determina que si bé, en general, el procés de restauració dels espais afectats per les activitats extractives està sent molt important des de l’entrada en vigor de la Llei 12/1981, no es pot afirmar que aquests treballs es realitzin de forma compaginada amb els d’explotació.

Estudi pilot del progrés de la restauració a les activitats extractives de Catalunya

L’objecte d’aquest estudi és determinar el ritme restaurador i caracteritzar la tipologia de les restauracions d’algunes activitats extractives de Catalunya. S’ha analitzat l’evolució dels treballs de restauració en una selecció d’activitats extractives de Catalunya i s’ha estimat la qualitat de les restauracions efectuades en base a la observació de la vegetació desenvolupada, el control de l’erosió i la integració paisatgística. Per dur a terme aquest estudi s’ha creat una base de dades en format de qüestionari, combinada amb un sistema d’informació geogràfica, que s’ha assajat en un treball de camp amb una mostra limitada d’activitats extractives. A partir de la interpretació dels resultats, es determina que actualment la gestió incorpora la necessitat de restaurar, però com a element subordinat a la producció. Això produeix una manca de planificació que incideix directament en la qualitat de les restauracions dutes a terme. La gestió de les pedreres encara ha de situar el concepte de restauració integrada com a eix central del desenvolupament i planificació de l’activitat extractiva.

The implementation of liquid chromatography tandem mass spectrometry for the official control of lipophilic toxins in seafood: Single-laboratory validation under four chromatographic conditions

We performed a comprehensive study to assess the fit for purpose of four chromatographic conditions for the determination of six groups of marine lipophilic toxins (okadaic acid and dinophysistoxins, pectenotoxins, azaspiracids, yessotoxins, gymnodimine and spirolides) by LC-MS/MS to select the most suitable conditions as stated by the European Union Reference Laboratory for Marine Biotoxins (EURLMB). For every case, the elution gradient has been optimized to achieve a total run-time cycle of 12 min. We performed a single-laboratory validation for the analysis of three relevant matrices for the seafood aquaculture industry (mussels, pacific oysters and clams), and for sea urchins for which no data about lipophilic toxins have been reported before. Moreover, we have compared the method performance under alkaline conditions using two quantification strategies: the external standard calibration (EXS) and the matrix-matched standard calibration (MMS). Alkaline conditions were the only scenario that allowed detection windows with polarity switching in a 3200 QTrap mass spectrometer, thus the analysis of all toxins can be accomplished in a single run, increasing sample throughput. The limits of quantification under alkaline conditions met the validation requirements established by the EURLMB for all toxins and matrices, while the remaining conditions failed in some cases. The accuracy of the method and the matrix effects where generally dependent on the mobile phases and the seafood species. The MMS had a moderate positive impact on method accuracy for crude extracts, but it showed poor trueness for seafood species other than mussels when analyzing hydrolyzed extracts. Alkaline conditions with EXS and recovery correction for OA were selected as the most proper conditions in the context of our laboratory. This comparative study can help other laboratories to choose the best conditions for the implementation of LC-MS/MS according to their own necessities.

Prevalence and persistence of gymnodimines in clams from the Gulf of Gabes (Tunisia) studied by mouse bioassay and LC-MS/MS

In this work we studied the toxicity in clams from the Gulf of Gabes, Tunisia (Southern Mediterranean). Samples from two stations (M2 and S6) were collected monthly from January 2009 to September 2010, and analyzed by the official control method of mousse bioassay (MBA) for lipophilic toxins. All samples were also analyzed with the LC-MS/MS method for the determination of lipophilic toxins, namely: okadaic acid group, pectenotoxins, yessotoxins and azaspiracids, spirolides and gymnodimines (GYMs). The results showed prevalence of GYMs since it was the only toxin group identified in these samples with a maximum of 2,136 μg GYM -A kg-1 (February 2009 at M2). Furthermore, GYMs showed persistence in the area, with only one blank sample below the limit of detection. Interestingly, this blank sample was found in June 2009 after an important toxic episode which supports the recent findings regarding the high detoxification capability of clams, much faster than that reported for oysters. In comparison, good agreement was found among MBA, the LD50 value of 80-100 μg kg-1 reported for GYM- A, and quantitative results provided by LC-MS/MS. On the contrary to that previously reported for Tunisian clams, we unambiguously identified and quantified by LC-MS/MS the isomers GYM- B/C in most samples. Phytoplankton identification and enumeration of Karenia selliformis usually showed higher densities at site M2 than S6 as expected bearing in mind toxin results, although additional results would be required to improve the correlation between K. selliformis densities and quantitative results of toxins. The prevalence and persistence of GYMs in this area at high levels strongly encourages the evaluation of the chronic toxic effects of GYMs. This is especially important taking into account that relatively large quantities of GYMs can be released into the market due to the replacement of the official control method from mouse bioassay to the LC-MS/MS for lipophilic toxins (Regulation (EU) No 15/2011), and the lack of Regulation for this group of toxins.

L'altra cara del progrés. Estudi multidimensional sobre les conseqüències de les activitats d'explotació intensiva sobre l'ecosistema del manglar i les activitats econòmiques tradicionals a la comunitat de Cumbe, Brasil

La localitat de Cumbe és una comunitat tradicional que es troba al tram baix del riu Jaguaribe situat a l’estat de Ceará (Brasil) i que ha viscut centenars d’anys a partir de les pràctiques extractives que es porten a terme a l’ecosistema del manglar. Fa 20 anys va aparèixer a la regió una nova economia basada en l’explotació intensiva. L’aqüicultura destinada a la cria de gamba està causant danys severs a l’ecosistema del manglar i perjudicant greument la font d’ingressos de moltes famílies, amenaçant la continuïtat de l’economia local i deteriorant la qualitat de vida de les persones que s’hi dediquen. A partir de l’observació personal i la immersió en aquesta societat s’ha detectat que l’economia tradicional està infravalorada pel govern i per les persones que donen suport a l’aqüicultura. Per aquest motiu s’han creat una sèrie d’indicadors, a partir de les activitats extractives del cranc, que posin en valor aquesta economia tradicional envers la nova economia emergent.

Conjugation of genetically-engineered protein phosphatases to magnetic particles for okadaic acid detection

This work presents the functional characterisation of a protein phosphatase 2A (PP2A) catalytic subunit obtained by genetic engineering and its conjugation to magnetic particles (MPs) via metal coordination chemistry for the subsequent development of assays for diarrheic lipophilic marine toxins. Colorimetric assays with free enzyme have allowed the determination of the best enzyme activity stabiliser, which is glycerol at 10%. They have also demonstrated that the recombinant enzyme can be as sensitive towards okadaic acid (OA) (LOD=2.3μg/L) and dinophysistoxin-1 (DTX-1) (LOD=15.2μg/L) as a commercial PP2A and, moreover, it has a higher operational stability, which makes possible to perform the protein phosphatase inhibition assay (PPIA) with a lower enzyme amount. Once conjugated to MPs, the PP2A catalytic subunit still retains its enzyme activity and it can also be inhibited by OA (LOD=30.1μg/L).

Study of the variability in chemical composition of bark layers of Quercus Suber L. from different production areas

Cork is the bark of the cork oak tree (Quercus suber L), a renewable and biodegradable raw bioresource concentrated mainly in the Mediterranean region. Development of its potential uses as a biosorbent will require the investigation of its chemical composition; such information can be of help to understand its interactions with organic pollutants. The present study investigates the summative chemical composition of three bark layers (back, cork, and belly) of five Spanish cork samples and one cork sample from Portugal. Suberin was the main component in all the samples (21.1 to 53.1%), followed by lignin (14.8 to 31%), holocellulose (2.3 to 33.6%), extractives (7.3 to 20.4%), and ash (0.4 to 3.3%). The Kruskal-Wallis test was used to determine whether the variations in chemical composition with respect to the production area and bark layers were significant. The results indicate that, with respect to the bark layer, significant differences were found only for suberin and holocellulose contents: they were higher in the belly and cork than in the back. Based on the results presented, cork is a material with a lot of potential because of its heterogeneity in chemical composition

Precursor uptake assays and metabolic analyses in isolated tomato fruit chromoplasts

Background Carotenoids are the most widespread group of pigments found in nature. In addition to their role in the physiology of the plant, carotenoids also have nutritional relevance as their incorporation in the human diet provides health benefits. In non-photosynthetic tissues, carotenoids are synthesized and stored in specialized plastids called chromoplasts. At present very little is known about the origin of the metabolic precursors and cofactors required to sustain the high rate of carotenoid biosynthesis in these plastids. Recent proteomic data have revealed a number of biochemical and metabolic processes potentially operating in fruit chromoplasts. However, considering that chloroplast to chromoplast differentiation is a very rapid process during fruit ripening, there is the possibility that some of the proteins identified in the proteomic analysis could represent remnants no longer having a functional role in chromoplasts. Therefore, experimental validation is necessary to prove whether these predicted processes are actually operative in chromoplasts. Results A method has been established for high-yield purification of tomato fruit chromoplasts suitable for metabolic studies. Radiolabeled precursors were efficiently incorporated and further metabolized in isolated chromoplast. Analysis of labeled lipophilic compounds has revealed that lipid biosynthesis is a very efficient process in chromoplasts, while the relatively low incorporation levels found in carotenoids suggest that lipid production may represent a competing pathway for carotenoid biosynthesis. Malate and pyruvate are efficiently converted into acetyl-CoA, in agreement with the active operation of the malic enzyme and the pyruvate dehydrogenase complex in the chromoplast. Our results have also shown that isolated chromoplasts can actively sustain anabolic processes without the exogenous supply of ATP, thus suggesting that these organelles may generate this energetic cofactor in an autonomous way. Conclusions We have set up a method for high yield purification of intact tomato fruit chromoplasts suitable for precursor uptake assays and metabolic analyses. Using targeted radiolabeled precursors we have been able to unravel novel biochemical and metabolic aspects related with carotenoid and lipid biosynthesis in tomato fruit chromoplasts. The reported chromoplast system could represent a valuable platform to address the validation and characterization of functional processes predicted from recent transcriptomic and proteomic data.

In rats, oral oleoyl-DHEA is rapidly hydrolysed and converted to DHEA-sulphate

Background: Dehydroepiandrosterone (DHEA) released by adrenal glands may be converted to androgens and estrogens mainly in the gonadal, adipose, mammary, hepatic and nervous tissue. DHEA is also a key neurosteroid and has antiglucocorticoid activity. DHEA has been used for the treatment of a number of diseases, including obesity; its pharmacological effects depend on large oral doses, which effect rapidly wanes in part because of its short half-life in plasma. Since steroid hormone esters circulate for longer periods, we have studied here whether the administration of DHEA oleoyl ester may extend its pharmacologic availability by keeping high circulating levels. Results: Tritium-labelled oleoyl-DHEA was given to Wistar male and female rats by gastric tube. The kinetics of appearance of the label in plasma was unrelated to sex; the pattern being largely coincident with the levels of DHEA-sulfate only in females, and after 2 h undistinguishable from the results obtained using labelled DHEA gavages; in the short term, practically no lipophilic DHEA label was found in plasma. After 24 h only a small fraction of the label remained in the rat organs, with a different sex-related distribution pattern coincident for oleoyl- and free- DHEA gavages. The rapid conversion of oleoyl-DHEA into circulating DHEA-sulfate was investigated using stomach, liver and intestine homogenates; which hydrolysed oleoyl-DHEA optimally near pH 8. Duodenum and ileum contained the highest esterase activities. Pure hog pancreas cholesterol-esterase broke down oleoyl-DHEA at rates similar to those of oleoyl-cholesterol. The intestinal and liver esterases were differently activated by taurocholate and showed different pH-activity patterns than cholesterol esterase, suggesting that oleoyl-DHEA can be hydrolysed by a number of esterases in the lumen (e.g. cholesterol-esterase), in the intestinal wall and the liver. Conclusion: The esterase activities found may condition the pharmacological availability (and depot effect) of orally administered steroid hormone fatty acid esters such as oleoyl-DHEA. The oral administration of oleoyl-DHEA in order to extend DHEA plasma availability has not been proved effective, since the ester is rapidly hydrolysed, probably in the intestine itself, and mainly converted to DHEA-sulfate at least in females.

TrkB and TrkC signaling are required for maturation and synaptogenesis of hippocampal connections

Recent studies have suggested a role for neurotrophins in the growth and refinement of neural connections, in dendritic growth, and in activity-dependent adult plasticity. To unravel the role of endogenous neurotrophins in the development of neural connections in the CNS, we studied the ontogeny of hippocampal afferents intrkB (¿/¿) and trkC (¿/¿) mice. Injections of lipophilic tracers in the entorhinal cortex and hippocampus of newborn mutant mice showed that the ingrowth of entorhinal and commissural/associational afferents to the hippocampus was not affected by these mutations. Similarly, injections of biocytin in postnatal mutant mice (P10¿P16) did not reveal major differences in the topographic patterns of hippocampal connections. In contrast, quantification of biocytin-filled axons showed that commissural and entorhinal afferents have a reduced number of axon collaterals (21¿49%) and decreased densities of axonal varicosities (8¿17%) in both trkB (¿/¿) and trkC (¿/¿) mice. In addition, electron microscopic analyses showed thattrkB (¿/¿) and trkC (¿/¿) mice have lower densities of synaptic contacts and important structural alterations of presynaptic boutons, such as decreased density of synaptic vesicles. Finally, immunocytochemical studies revealed a reduced expression of the synaptic-associated proteins responsible for synaptic vesicle exocytosis and neurotransmitter release (v-SNAREs and t-SNAREs), especially in trkB (¿/¿) mice. We conclude that neither trkB nor trkC genes are essential for the ingrowth or layer-specific targeting of hippocampal connections, although the lack of these receptors results in reduced axonal arborization and synaptic density, which indicates a role for TrkB and TrkC receptors in the developmental regulation of synaptic inputs in the CNS in vivo. The data also suggest that the genes encoding for synaptic proteins may be targets of TrkB and TrkC signaling pathways.

A simple halide-to-anion exchange method for heteroaromatic salts and ionic liquids

A broad and simple method permitted halide ions in quaternary heteroaromatic and ammonium salts to be exchanged for a variety of anions using an anion exchange resin (A− form) in non-aqueous media. The anion loading of the AER (OH− form) was examined using two different anion sources, acids or ammonium salts, and changing the polarity of the solvents. The AER (A− form) method in organic solvents was then applied to several quaternary heteroaromatic salts and ILs, and the anion exchange proceeded in excellent to quantitative yields, concomitantly removing halide impurities. Relying on the hydrophobicity of the targeted ion pair for the counteranion swap, organic solvents with variable polarity were used, such as CH3OH, CH3CN and the dipolar nonhydroxylic solvent mixture CH3CN:CH2Cl2 (3:7) and the anion exchange was equally successful with both lipophilic cations and anions.

A simple halide-to-anion exchange method for heteroaromatic salts and ionic liquids

A broad and simple method permitted halide ions in quaternary heteroaromatic and ammonium salts to be exchanged for a variety of anions using an anion exchange resin (A− form) in non-aqueous media. The anion loading of the AER (OH− form) was examined using two different anion sources, acids or ammonium salts, and changing the polarity of the solvents. The AER (A− form) method in organic solvents was then applied to several quaternary heteroaromatic salts and ILs, and the anion exchange proceeded in excellent to quantitative yields, concomitantly removing halide impurities. Relying on the hydrophobicity of the targeted ion pair for the counteranion swap, organic solvents with variable polarity were used, such as CH3OH, CH3CN and the dipolar nonhydroxylic solvent mixture CH3CN:CH2Cl2 (3:7) and the anion exchange was equally successful with both lipophilic cations and anions.

Inhibition of nociceptive responses after systemic administration of amidated kyotorphin

BACKGROUND AND PURPOSE Kyotorphin (KTP; L-Tyr-L-Arg), an endogenous neuropeptide, is potently analgesic when delivered directly to the central nervous system. Its weak analgesic effects after systemic administration have been explained by inability to cross the blood-brain barrier (BBB) and detract from the possible clinical use of KTP as an analgesic. In this study, we aimed to increase the lipophilicity of KTP by amidation and to evaluate the analgesic efficacy of a new KTP derivative (KTP-amide - KTP-NH 2). EXPERIMENTAL APPROACH We synthesized KTP-NH 2. This peptide was given systemically to assess its ability to cross the BBB. A wide range of pain models, including acute, sustained and chronic inflammatory and neuropathic pain, were used to characterize analgesic efficacies of KTP-NH 2. Binding to opioid receptors and toxicity were also measured. KEY RESULTS KTP-NH 2, unlike its precursor KTP, was lipophilic and highly analgesic following systemic administration in several acute and chronic pain models, without inducing toxic effects or affecting motor responses and blood pressure. Binding to opioid receptors was minimal. KTP-NH 2 inhibited nociceptive responses of spinal neurons. Its analgesic effects were prevented by intrathecal or i.p. administration of naloxone. CONCLUSIONS AND IMPLICATIONS Amidation allowed KTP to show good analgesic ability after systemic delivery in acute and chronic pain models. The indirect opioid-mediated actions of KTP-NH 2 may explain why this compound retained its analgesic effects although the usual side effects of opioids were absent, which is a desired feature in next-generation pain medications