The Ecological Dose of Nickel in a Semiarid Soil Amended with Sewage Sludge Related to the Unamended Soil

Nickel pollution may affect microbial communitydevelopment and its activity in soil and, therefore, itsfertility, while organic amendment may affect Ni mobilityand bioavailability. The aim of this research was toascertain the extent to which the addition of sewage sludgeto soil may affect Ni toxicity and to establish theecological dose of this heavy metal. Unamended and amendedsoils with sewage sludge were spiked with differentconcentrations of Ni (0–8000 mg kg^-1) and then incubatedfor 3 hours, 12 days and 40 days. Soil dehydrogenaseactivity, ATP content, microbial respiration and microbialbiomass C were measured in the samples containing differentNi concentrations. A mathematical model which describe theinhibition of these soil parameters were used to calculatethe ecological dose of Ni: ED₅₀. Soil microbialbiomass C measurements of all treatments better fitted themodel (r² = 0.95–0.69 with p < 0.05) than the otherparameters studied; but the ED₅₀ values predicted by the modelhad a high coefficient of variation. For both ATP contentand microbial biomass C, the ED₅₀ values calculatedfor the amended soil were higher than for the unamendedsoil and so that the effect of Ni toxicity on theseparameters of microbial activity may be considered lower inthe former. In this study, an increase in ED₅₀ valueswith the time elapsed was also observed.     

Optimization and Validation of a High Throughput UHPLC-MS/MS Method for Determination of the EU Regulated Lipophilic Marine Toxins and Occurrence in Fresh and Processed Shellfish

Under the name of lipophilic marine toxins, there are included more than 1000 toxic secondary metabolites, produced by phytoplankton, with the common chemical property of lipophilicity. Due to toxicological effects and geographical distribution, in European legislation relevant compounds are regulated, and their determination is accomplished with the reference liquid chromatography-tandem mass spectrometry method. In this study a modified ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method has been developed for the identification and quantification of EU-regulated lipophilic toxins. The method optimization included a refinement of SPE-C18 clean-up, in order to reduce matrix interferences. Improved LC conditions and upgraded chromatographic ammonia-based gradient ensured the best separation of all analytes and, in particular, of the two structural isomers (OA and DTX2). Also, different MS parameters were tested, and confirmation criteria finally established. The validation studies confirmed that all parameters were satisfactory. The requirements for precision (RSD% < 11.8% for each compound), trueness (recoveries from 73 to 101%) and sensitivity (limits of quantification in the range 3–8 µg kg⁻¹) were fulfilled. The matrix effect, ranging from −9 to 19%, allowed the use of a calibration curve in solvent (3–320 µg kg⁻¹ in matrix) for quantification of real samples. Method relative uncertainty ranged from 12 to 20.3%. Additionally, a total of 1000 shellfish samples was analysed, providing a first preliminary surveillance study that may contribute to the knowledge of lipophilic marine toxins contamination. Increase in algae proliferation events and intoxication cases, EFSA suggestions for modification of maximum permitted levels and toxicity equivalency factors, and new studies of important toxic effects underline that implementation of reference methods still represents an important task for health and food safety laboratories.     

Effects of filtration of semen doses from subfertile boars through neuter Sephadex columns.

This study was designed to develop a method of improving the quality of sperm obtained from subfertile Piétrain boars. Seminal doses were filtered through neuter Sephadex columns (G-25 Medium, G-50 Fine, G-50 Medium and G-75, length 10 +/- 0.5 cm, flow rate 1 ml/20 s). Doses were prepared by pooling 10 ml semen samples collected from 58 asthenoteratospermic boars and diluted the sperm-cell rich fraction 1 : 6 in Betsville thawing solution extender. Sperm quality was determined before and after the filtering process. Sperm morphology and motility were assessed using the computer program SCA 2002 production, and sperm vitality was evaluated by fluorescence multistaining. ORT and HRT tests were used to determine the osmotic resistance of spermatozoa, and metabolic performance was assessed by measuring l-lactate production. Results indicate that the filtration process rendered increased proportions of mature spermatozoa and of viable spermatozoa with an intact acrosome, nucleus and mitochondrial sheath. Sperm filtration led to decreased percentages of spermatozoa with proximal and distal droplets and of agglutinated spermatozoa, along with slightly diminished ORT values. HRT scores and L-lactate production were unaffected. Our findings indicate that filtering through a Sephadex column improves the sperm morphology and vitality of seminal doses obtained from subfertile boars, but produces no functional changes in the spermatozoa. All four column types yielded similar results.     

Long-term control of the invasive tree Ailanthus altissima: Insights from Mediterranean protected forests

Ailanthus altissima is an invasive tree species which has colonized numerous ecosystems and affected ecosystem processes worldwide. Despite its importance as an invasive species and the high economic costs incurred from its spread, there is a lack of long-term management planning for its control. Although mechanical disturbance is commonly applied, the effect that this treatment may have exhausting its resprouting ability and also its joint effect with phytochemical treatments are poorly understood, especially in Mediterranean environments. We tested three treatments (plus a control) aimed to reduce A. altissima growth in Mediterranean forests throughout 5 years of study. The treatments (one cut stump, double cut stump and cut stump with glyphosate application) were repeated annually. General plant performance (biomass, height and resprout-type density) was measured yearly during the study. Water potential and stomatal conductance were also measured at the end of the study to evaluate particular ecophysiological factors which might affect the response of A. altissima to assayed treatments, together with leaf area index. Our results show that only the cut stump with glyphosate application treatment was able to reduce the long-term growth and spread of A. altissima. The treatments applied favoured collar sprout growth in response to disturbance events (treatments) opposite to the control, where new sprouts grew mainly from the root. Treated resprouts displayed ecophysiological changes depending on the assayed treatment. To our knowledge, this is the only study testing the long-term effect of both physical disturbance and phytochemical application on A. altissima growth. Our study further refines our knowledge on the effects of repeating both commonly and newly used treatments, improving our management techniques to reduce the presence and growth of this invasive tree.     

Comparative topical anti-inflammatory activity of cannabinoids and cannabivarins

A selection of seven phytocannabinoids representative of the major structural types of classic cannabinoids and their corresponding cannabivarins was investigated for in vivo topical anti-inflammatory activity in the Croton oil mouse ear dermatitis assay. Differences in the terpenoid moiety were far more important for anti-inflammatory activity than those at the C-3 alkyl residue, suggesting the involvement not only of cannabinoid receptors, but also of other inflammatory end-points targeted by phytocannabinoids.     

Pomegranate trees performance under sustained and regulated deficit irrigation

The effects of sustained and regulated deficit irrigation (SDI and RDI) on “Mollar de Elche” pomegranate tree performance were investigated in a field trial conducted over three consecutive seasons. In the RDI regimes, severe water restrictions were applied during one of three phases: flowering and fruit set, fruit growth, or the final phase of fruit growth and ripening. In another approach, SDI was applied by watering trees at 50 % of the estimated crop water needs (ET_c) during the entire season. Results showed that even after three consecutive seasons of water restrictions, similar yield levels were obtained in SDI and Control trees watered at 100 % ET_c. This was because a 22 % reduction in average fresh fruit weight recorded in the SDI treatment was compensated by an increase in 28 % in the quantity of fruit collected per tree. This was most likely due to a reduction in the fall of the reproductive organs. However, the SDI strategy led to a reduction in 28 % in the yield value when fruits are sold for fresh fruit markets. Water restrictions applied only during flowering and fruit set also resulted in an increase in the quantity of fruit collected per tree, with only a slight reduction in fruit weight and without affecting the yield value. On the other hand, severe water restrictions applied during the summer (i.e., mid-phase of fruit growth) led to 24 % water savings with only a 7 % reduction in fruit weight. Fruit cracking was very low in all treatments and seasons (2–6 % over the total quantity fruit collected per tree). Only the RDI regime with restrictions during the summer increased cracking in one out of the three seasons. It is concluded that RDI can be used as a measure to cope with water scarcity and high water prices. Among all the RDI explored, the one with restrictions applied early in the season (during flowering and fruit set) was the most convenient strategy.     

Characterization of topical antiinflammatory compounds in Rosmarinus officinalis L

The topical antiinflammatory activity of three extracts at increasing polarity (n-hexane, chloroform, and methanol) from the leaves of Rosmarinus officinalis L. (Labiatae) has been tested using the croton oil ear test in mice. Both the n-hexane and the chloroform (CE-1) extracts from the leaves showed a dose-dependent activity, the last one possessing an antiinflammatory potency similar to that of indomethacin, the nonsteroidal antiinflammatory drug used as a reference drug (ID50 = 83 and 93 microg/cm2, respectively). The bioassay-oriented fractionation of CE-1 led to the identification of tritepenes, ursolic acid, oleanolic acid, and micromeric acid as the main antiinflammatory principles. Furthermore, the CE-1 extract obtained from the residue of the steam distillation of the leaves (extract A) showed the same antiinflammatory potency of CE-1, suggesting this waste product as a source of antiinflammatory products.