Nuevas estrategias analíticas para la determinación de residuos de insecticidas y alcaloides ergóticos en muestras ambientales y alimentarias

Résumé

In this Thesis, different analytical strategies have been proposed as efficient alternatives for the monitoring of several insecticides and ergot alkaloids, considered as relevant chemical hazards in food and environmental samples. Neonicotinoids (NNIs) are a class of insecticides widely used in the last decades to protect mainly plants (i.e., crops, fruits, and vegetables) from pest insect attack acting selectively on the nicotinic acetylcholine receptors (nAChRs) in the central nervous system (CNS) of insects. Due to their high solubility and relatively small molecular weight, they can be easily incorporated into the plant tissues via xylem staying for a long time after application. Thus, NNIs can be found in nectar and pollen, being easily available for non-target and beneficial insects such as pollinators. For these reasons, they have been related with the rapid loss of entire beehives phenomenon known as Colony Collapse Disorder (CCD). Although the action mode of fipronil (FPN) and its metabolites (FPN-sulfide, FPN-sulfone) is different, they have been also related to the death of honeybees. They act as potent disruptors of the insect CNS interfering with the passage of chloride ions through the γ-aminobutyric acid- (GABA-) regulated chloride ion channel, which triggers a severe paralysis and death of the insect. In addition, recent studies have revealed that the fungicide boscalid (BCL) can interact with other pesticides such as NNIs reducing the lethal time and lethal doses (LD50) for bees, as well as decreasing the ATP concentration, pollen consumption and protein digestion in bees. Due to these effects, the use of these pesticides has been banned or restricted in several European regulations. In addition, due to their systemic nature, these compounds can be found in food samples and pose a health risk in humans. For this reason, the EU has established limitations of their use and maximum residue levels (MRLs) have been set in a great number of food commodities to guarantee food safety. In light of these concerns, analytical methods for the determination of these insecticides are required in environmental and food safety fields. They must present high efficiency, selectivity and sensitivity in order to fulfill the European legislation. In this sense, novel analytical methods accomplishing these conditions to determine the above-mentioned insecticides in environmental and food samples have been developed in this Thesis. On the other hand, ergot alkaloids (EAs) are mycotoxins produced mainly by fungi of the Claviceps genus, as Claviceps purpurea, which parasitize the seeds of plants, especially in cereals (i.e. rye, triticale, wheat, oat, and barley) replacing the developing grain with fungal structures known as sclerotia that contain toxic alkaloid substances. The sclerotia are harvested together with the cereals leading to their contamination with EAs which can cause intoxications and ergotism in humans and animals. This illness is characterized by symptoms such as vomiting, burning sensation of the skin, insomnia, and hallucinations, and in some cases gangrenous limbs or even death. Ergotism, whose effects have been known since the fifth century, was historically known as St. Anthony’s Fire or holy fire. Cleaning improvements in grain processing have reduced the presence of sclerotia in cereals; however, EAs are still found in food and feed samples. Consequently, the European Commission has established a maximum level for ergot sclerotia and a tolerable daily intake for total EAs. Multiple factors affect EA production; the EA pattern and contents in sclerotia vary with the fungal strain, geographical region, host plant, and weather conditions. For that reason, the European Food and Safety Authority (EFSA) has asked for more studies including the analysis of EAs in cereal samples especially in processed foods destined for human consumption. In this regard, the determination of the more predominant EAs together with their epimers, in such samples using different analytical methods has been carried out in this Thesis. In general, to face with the analytical challenges above mentioned, the last technical advances in terms of miniaturization, increased efficiency, high sensitivity, high resolution, high capacity of identification and low consumption of solvents and sample have been taken into consideration. Different miniaturized separation techniques such as capillary electrophoresis (CE) and capillary liquid chromatography (CLC) have been selected because they involve low solvent consumption as it is recommended by the trends in green analytical chemistry. Moreover, the use of high efficient techniques such ultra-HPLC (UHPLC) provides lower solvent consumption than traditional LC methods, and also shorter analysis time. In addition to UV-Vis detection, mass spectrometry (MS) has been considered because of its high sensitivity and the capability for the unequivocal identification of compounds. Finally, the coupling of ion mobility spectrometry (IMS) to a LC-MS workflow has been evaluated to provide complementary information to mass spectra and retention time by means of the so-called collision cross section (CCS). It is important to emphasize that this Thesis presents for the first time the application of CLC as well as the use of MEKC-MS/MS technique for the determination of NNIs. Furthermore, it is the first time that the main EAs have been characterized in terms of CCS using travelling wave ion mobility (TWIM)-time of flight high resolution mass spectrometry (TOF-HRMS). In addition, different sample treatments involving on-line and off-line preconcentration methodologies and miniaturized extraction techniques have been proposed. These strategies have allowed a sensitivity and efficiency enhancement in the extraction and preconcentration of the target compounds in environmental and food samples. Moreover, these sample treatments are environmental-friendly being in accordance with current trends in Green Chemistry.