EU water framework directive: will the nitrate load reduction from diffuse sources produce the same results in all estuaries?

The impact of urban waste-water and non-point nitrate discharges in estuarine and near-shore coastal waters are analyzed. The study is focused on the effects of applying the European directives 91/271/EEC and 91/676/EEC to these systems. 4 Portuguese estuaries and two coastal lagoons with different characteristics are studied. A modelling system is applied and calibrated in each system. Three nitrate load scenarios are examined. It is shown that the morphologic and hydrodynamic characteristics of the domain largely control the ecological processes in these systems. The primary production limitation factors are split into “biologic” and “hydrodynamic” components. The physical limitation due to hydrodynamic and residence time is the most important factor. The combined limitation of “biologic” factors (temperature, light and nutrients availability) control productivity only in the systems where physical limitation is not important.

Effects of active pharmaceutical ingredients in mussel mytilus galloprovincialis

More than 3000 types of active pharmaceutical ingredients (APIs) are applied in Human and veterinary medicine practice. These compounds are considered an emergent class of environmental contaminants with the ability to cause damage and unexpected effects to aquatic organisms, namely in species of high commercial value. APIs are ubiquitous in the environment being frequently detected in influents and effluents of waste water treatment plants (WWTPs), surface waters and more distressingly in the public tap water in concentrations ranging from ng to μg.L-1. Considering these premises, the present thesis focused on APIs detection in the Arade river water, the impact of summer period in APIs’ concentration alterations applying the passive sampler device, POCIS (polar organic compound integrative sampler), as well as, the assessment of the effects caused by non-steroidal anti-inflammatory drugs (NSAID) ibuprofen (IBU) and diclofenac (DCF) and antidepressant selective serotonin reuptake inhibitor (SSRI) fluoxetine as single and mixture exposures along with a classical contaminant copper (Cu) on a non-target species, mussel Mytilus galloprovincialis. For this purpose, a multibiomarker approach was applied namely including biomarkers of oxidative stress (antioxidant enzymes activities of superoxide dismutase – SOD, catalase – CAT, glutathione reductase – GR and Phase II glutathione-S-transferase), damage - lipid peroxidation (LPO), neurotoxic effects (through the activity of acetylcholinesterase enzyme - AChE) and endocrine disruption (through vitellogenin-like proteins measurement applying the indirect method of alkali-labile phosphate - ALP) after exposure of mussel species’ to selected APIs at environmental relevant concentrations. The main results highlighted the occurrence of 19 APIs in the river Arade from several distinct therapeutic classes. Stimulant caffeine, antiasthmatic theophylline, NSAID ibuprofen and analgesic paracetamol presented the highest concentrations. Summer impact was inconclusive due to each API transient concentration in each month. The multibiomarker results revealed distinct responses towards each selected API (as single exposure or as mixtures) that were tissue and time dependent. Several multistressor interactions were proposed for each biomarker. The results also revealed APIs potential to induce oxidative stress, LPO, neurotoxicity and endocrine disruption even at extremely low concentrations on a species extremely vulnerable to APIs presence highlighting the urgency on the development of methodologies able to prevent its entrance in the aquatic environment.

Liver-like biosensor: immobilized CYP3A4 sensors as tools for ligand binding and enzyme activity measurements

Cytochromes P450 constitute a super-family of enzymes involved in the metabolism of Xenobiotics, where human cytochrome P450 3A4 is the most abundant of all P450s, accounting for about 50% of all human liver cytochromes. This membrane anchored protein is responsible for the metabolization of a wide array of environmental drugs and intoxicants, mainly due to its haem domain properties, and active site cavity volume. These properties make this protein an excellent subject for biosensor application, although CYO3A4 enzyme is also famous for its instability. Enzyme inactivation at room temperature is a normal conversion process that this enzyme undergoes, that may hamper any biosensing approach.

Root ferric chelate reductase is regulated by iron and copper in straberry plants

In the present experiment, we studied the interaction between copper (Cu) and iron (Fe) in strawberry plants grown in nutrient solutions containing different concentrations of Fe. Plants grown in the absence of iron (Fe0) had the characteristic symptoms of Fe deficiency, with smaller chlorotic leaves, less biomass, acidification of the nutrient solution, and roots that were smaller and less ramified, while no symptoms of Fe deficiency were observed in plants grown with Fe. A greater amount of Cu was found in roots of chlorotic plants than in those grown with Fe, while plants grown with 20M of Fe (Fe20) in the nutrient solution had a greater amount of Fe compared with plants from the other treatments. Chlorotic plants (Fe0) and plants grown with the greatest level of Fe (Fe20) had a greater root ferric chelate reductase (FC-R; EC 1.16.1.17) activity compared with the other treatments with 5 or 10M Fe in the nutrient solution. The same pattern was obtained for relative FC-R mRNA concentration and for the sum of Fe and Cu contents in shoots (leaves plus crowns). The DNA obtained from amplification of the FC-R mRNA was cloned and several of the inserts analysed by single strand confirmation polymorphism (SSCP). Although there were different SSCP patterns in the Fe20 treatment, all the inserts that were sequenced were very similar, excluding the hypothesis of more than one FC-R mRNA species being present. The results suggest that Cu as well as Fe is involved in FC-R expression and activity, although the mechanism involved in this regulation is unknown so far. Both small contents of Fe and Cu in plants led to an over-expression of the FC-R gene and enhanced FC-R activity in strawberry roots.

Effects of vitamin K deficiency and its mechanisms in vertebrate's early development: zebrafish as a model

Disertação de mestrado, Ciências Biomédicas, Departamento de Ciências Biomédicas e Medicina, Universidade do Algarve, 2015