RESUMO
Metal sub-microparticles (SMPs) and nanoparticles (NPs) presence in food is attributable to increasing pollution from the environment in raw materials and finished products. In the present study, a multifaceted analytical strategy based on Environmental Scanning Electron Microscopy and High-Angle Annular Dark-Field-Scanning Transmission Electron Microscopy coupled with Energy-Dispersive X-ray Spectroscopy (ESEM-EDX, HAADF-STEM-EDX) and Inductively Coupled Plasma Mass Spectrometry (ICP-MS) was proposed for the detection and characterization of metal and metal-containing SMPs and NPs in durum wheat samples, covering a size measurement range from 1 nm to multiple µm. ESEM-EDX and ICP-MS techniques were applied for the assessment of SMP and NP contamination on the surface of wheat grains collected from seven geographical areas characterized by different natural and anthropic conditions, namely Italy, the USA, Australia, Slovakia, Mexico, Austria, and Russia. ICP-MS showed significant differences among the mean concentration levels of metals, with the USA and Italy having the highest level. ESEM-EDX analysis confirmed ICP-MS concentration measurements and measured the highest presence of particles < 0.8 µm in size in samples from Italy, followed by the USA. Less marked differences were observed when particles < 0.15 µm were considered. HAADF-STEM-EDX was applied to a selected number of samples for a preliminary assessment of internal contamination by metal SMPs and NPs, and to expand the measurable particle size range. The multifaceted approach provided similar results for Fe-containing SMPs and NPs. ICP-MS and ESEM-EDX also highlighted the presence of a significant abundance of Ti- and Al-containing particles, while for STEM-EDX, sample preparation artifacts complicated the interpretation. Finally, HAADF-STEM-EDX results provided relevant information about particles in the low nm range, since, by applying this technique, no particles smaller than 50 nm were observed in accordance with ESEM-EDX.
Assuntos
Espectrometria de Massas , Nanopartículas Metálicas , Triticum , Triticum/química , Nanopartículas Metálicas/química , Espectrometria de Massas/métodos , Espectrometria por Raios X/métodos , Tamanho da Partícula , Metais/análise , Metais/química , Grão Comestível/química , Microscopia Eletrônica de VarreduraRESUMO
The potential of a voltametric E-tongue coupled with a custom data pre-processing stage to improve the performance of machine learning techniques for rapid discrimination of tomato purées between cultivars of different economic value has been investigated. To this aim, a sensor array with screen-printed carbon electrodes modified with gold nanoparticles (GNP), copper nanoparticles (CNP) and bulk gold subsequently modified with poly(3,4-ethylenedioxythiophene) (PEDOT), was developed to acquire data to be transformed by a custom pre-processing pipeline and then processed by a set of commonly used classifiers. The GNP and CNP-modified electrodes, selected based on their sensitivity to soluble monosaccharides, demonstrated good ability in discriminating samples of different cultivars. Among the different data analysis methods tested, Linear Discriminant Analysis (LDA) proved to be particularly suitable, obtaining an average F1 score of 99.26%. The pre-processing stage was beneficial in reducing the number of input features, decreasing the computational cost, i.e., the number of computing operations to be performed, of the entire method and aiding future cost-efficient hardware implementation. These findings proved that coupling the multi-sensing platform featuring properly modified sensors with the custom pre-processing method developed and LDA provided an optimal tradeoff between analytical problem solving and reliable chemical information, as well as accuracy and computational complexity. These results can be preliminary to the design of hardware solutions that could be embedded into low-cost portable devices.
Assuntos
Ouro , Aprendizado de Máquina , Solanum lycopersicum , Solanum lycopersicum/classificação , Solanum lycopersicum/química , Ouro/química , Análise Discriminante , Nariz Eletrônico , Nanopartículas Metálicas/química , Eletrodos , Polímeros/química , Cobre/química , Compostos Bicíclicos Heterocíclicos com Pontes/químicaRESUMO
Aptamers are recognition elements increasingly used for the development of biosensing strategies, especially in the detection of proteins or small molecule targets. Lysozyme, which is recognized as an important biomarker for various diseases and a major allergenic protein found in egg whites, is one of the main analytical targets of aptamer-based biosensors. However, since aptamer-based strategies can be prone to artifacts and data misinterpretation, rigorous strategies for multifaceted characterization of the aptamer-target interaction are needed. In this work, a multitechnique approach has been devised to get further insights into the binding performance of the anti-lysozyme DNA aptamers commonly used in the literature. To study molecular interactions between lysozyme and different anti-lysozyme DNA aptamers, measurements based on a magneto-electrochemical apta-assay, circular dichroism spectroscopy, fluorescence spectroscopy, and asymmetrical flow field-flow fractionation were performed. The reliability and versatility of the approach were proved by investigating a SELEX-selected RNA aptamer reported in the literature, that acts as a positive control. The results confirmed that an interaction in the low micromolar range is present in the investigated binding buffers, and the binding is not associated with a conformational change of either the protein or the DNA aptamer. The similar behavior of the anti-lysozyme DNA aptamers compared to that of randomized sequences and polythymine, used as negative controls, showed nonsequence-specific interactions. This study demonstrates that severe testing of aptamers resulting from SELEX selection is the unique way to push these biorecognition elements toward reliable and reproducible results in the analytical field.
Assuntos
Aptâmeros de Nucleotídeos , Aptâmeros de Nucleotídeos/química , Muramidase , Reprodutibilidade dos Testes , Técnica de Seleção de Aptâmeros/métodos , Anticorpos AntinuclearesRESUMO
The European Union is among the top wheat producers in the world, but its productivity relies on adequate soil fertilisation. Biofertilisers, either alone or in combination with biochar, can be a preferable alternative to chemical fertilisers. However, the addition of biofertilisers, specifically plant growth promoting microbes (PGPM), could modify grain composition, and/or deteriorate the soil composition. In this study, the two wheat cultivars Triticum aestivum (Bramante) and T. durum (Svevo) were cultivated in open fields for two consecutive years in the presence of a commercial PGPM mix supplied alone or in combination with biochar. An in-depth analysis was conducted by collecting physiological and agronomic data throughout the growth period. The effects of PGPM and biochar were investigated in detail; specifically, soil chemistry and rhizosphere microbial composition were characterized, along with the treatment effects on seed storage proteins. The results demonstrated that the addition of commercial microbial consortia and biochar, alone or in combination, did not modify the rhizospheric microbial community; however, it increased grain yield, especially in the cultivar Svevo (increase of 6.8 %-13.6 %), even though the factors driving the most variations were associated with both climate and cultivar. The total gluten content of the flours was not affected, whereas the main effect of the treatments was a variation in gliadins and low-molecular-weight-glutenin subunits in both cultivars when treated with PGPM and biochar. This suggested improved grain quality, especially regarding the viscoelastic properties of the dough, when the filling period occurred in a dry climate. The results indicate that the application of biofertilisers and biochar may aid the effective management of sustainable wheat cultivation, to support environmental health without altering the biodiversity of the resident microbiome.
Assuntos
Consórcios Microbianos , Triticum , Grão Comestível , Carvão Vegetal/farmacologia , Solo/químicaRESUMO
The use of PGPR is widely accepted as a promising tool for a more sustainable agricultural production and improved plant abiotic stress resistance. This study tested the ability of PVr_9, a novel bacterial strain, homologous to Beijerinckia fluminensis, to increase salt stress tolerance in A. thaliana. In vitro plantlets inoculated with PVr_9 and treated with 150 mM NaCl showed a reduction in primary root growth inhibition compared to uninoculated ones, and a leaf area significantly less affected by salt. Furthermore, salt-stressed PVr_9-inoculated plants had low ROS and 8-oxo-dG, osmolytes, and ABA content along with a modulation in antioxidant enzymatic activities. A significant decrease in Na+ in the leaves and a corresponding increase in the roots were also observed in salt-stressed inoculated plants. SOS1, NHX1 genes involved in plant salt tolerance, were up-regulated in PVr_9-inoculated plants, while different MYB genes involved in salt stress signal response were down-regulated in both roots and shoots. Thus, PVr_9 was able to increase salt tolerance in A. thaliana, thereby suggesting a role in ion homeostasis by reducing salt stress rather than inhibiting total Na+ uptake. These results showed a possible molecular mechanism of crosstalk between PVr_9 and plant roots to enhance salt tolerance, and highlighted this bacterium as a promising PGPR for field applications on agronomical crops.
Assuntos
Arabidopsis , Beijerinckiaceae , Arabidopsis/metabolismo , Tolerância ao Sal/genética , Proteínas de Plantas/genética , Beijerinckiaceae/metabolismo , Agrobacterium tumefaciens , Regulação da Expressão Gênica de Plantas , Raízes de Plantas/genéticaRESUMO
Sustainable agriculture aims at achieving a healthy food production while reducing the use of fertilizers and greenhouse gas emissions using biostimulants and soil amendments. Untargeted metabolomics by ultra-high performance liquid chromatography-ion mobility-high-resolution mass spectrometry, operating in a high-definition MSE mode, was applied to investigate the metabolome of durum wheat in response to sustainable treatments, i.e., the addition of biochar, commercial plant growth promoting microbes, and their combination. Partial least squares-discriminant analysis provided a good discrimination among treatments with sensitivity, specificity, and a non-error rate close to 1. A total of 88 and 45 discriminant compounds having biological, nutritional, and technological implications were tentatively identified in samples grown in 2020 and 2021. The addition of biochar-biostimulants produced the highest up-regulation of lipids and flavonoids, with the glycolipid desaturation being the most impacted pathway, whereas carbohydrates were mostly down-regulated. The findings achieved suggest the safe use of the combined biochar-biostimulant treatment for sustainable wheat cultivation.
Assuntos
Metabolômica , Triticum , Cromatografia Líquida de Alta Pressão/métodos , Metabolômica/métodos , Espectrometria de Massas/métodosRESUMO
Dental calculus is a valuable resource for the reconstruction of dietary habits and oral microbiome of past populations. In 2020 the remains of Duke Alessandro Farnese and his wife Maria D'Aviz were exhumed to get novel insights into the causes of death. This study aimed to investigate the dental calculus metabolome of the noble couple by untargeted metabolomics. The pulverized samples were decalcified in a water-formic acid mixture, extracted using methanol/acetonitrile and analyzed by ultra-high performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS) using a reversed-phase separation followed by electrospray ionization and full scan in positive and negative ion mode. Waters Synapt-G2-Si High-Definition hybrid quadrupole time-of-flight mass spectrometer was used. Significant features were then identified using MSE acquisition mode, recording information on exact mass precursor and fragment ions within the same run. This approach, together with data pre-treatment and multivariate statistical analysis allowed for the identification of compounds able to differentiate between the investigated samples. More than 200 metabolites were identified, being fatty acids, alcohols, aldehydes, phosphatidylcholines, phosphatidylglycerols, ceramides and phosphatidylserines the most abundant classes. Metabolites deriving from food, bacteria and fungi were also determined, providing information on the habits and oral health status of the couple.
Assuntos
Cálculos Dentários , Metabolômica , Humanos , Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas , Metabolômica/métodos , MetabolomaRESUMO
Mass spectrometry (MS)-based proteomics has recently attracted the attention from forensic pathologists. This work is the first report of the development of a shotgun bottom-up proteomic approach based on rapid protein extraction and nano-liquid chromatography/high-resolution mass spectrometry applied to full-thickness human skin for the differential analysis of normal and ecchymotic tissues to identify new biomarkers for bruise characterization and dating. We identified around 2000 proteins from each pooled extract. The method showed excellent precision on independent replicates, with Pearson correlation coefficients always higher than 95%. Glycophorin A, a known biomarker of vital wounds from immunochemical studies, was identified only in ecchymotic tissues, as confirmed by Western blotting analysis. This finding suggests that this protein can be used as a MS-detectable biomarker of wound vitality. By focusing on skin samples from individuals with known wound dating, besides Glycophorin A, other proteins differentially expressed in ecchymotic samples and dependant on wound age were identified, although further analysis on larger datasets are needed to validate these findings. This study paves the way for an in-depth investigation of the potential of MS-based techniques for wound examination in forensic pathology, overcoming the limitations of immunochemical assays.
Assuntos
Glicoforinas , Espectrometria de Massas em Tandem , Humanos , Espectrometria de Massas em Tandem/métodos , Proteômica/métodos , Patologia Legal , Proteínas/metabolismo , BiomarcadoresRESUMO
Traditional techniques for food analysis are based on off-line laboratory methods that are expensive and time-consuming and often require qualified personnel. Despite the high standards of accuracy and metrological traceability, these well-established methods do not facilitate real-time process monitoring and timely on-site decision-making as required for food safety and quality control. The future of food testing includes rapid, cost-effective, portable, and simple methods for both qualitative screening and quantification of food contaminants, as well as continuous, real-time measurement in production lines. Process automatization through process analytical technologies (PAT) is an increasing trend in the food industry as a way to achieve improved product quality, safety, and consistency, reduced production cycle times, minimal product waste or reworks, and the possibility for real-time product release. Novel methods of analysis for point-of-need (PON) screening could greatly improve food testing by allowing non-experts, such as consumers, to test in situ food products using portable instruments, smartphones, or even visual naked-eye inspections, or farmers and small producers to monitor products in the field. This requires the attention of the research community and devices manufacturers to ensure reliability of measurement results from PAT strategy and PON tests through the demonstration and critical evaluation of performance characteristics. The fitness for purpose of methods in real-life conditions is a priority that should not be overlooked in order to maintain an effective and harmonized food safety policy.
Assuntos
Inocuidade dos Alimentos , Reprodutibilidade dos Testes , Inocuidade dos Alimentos/métodos , Controle de Qualidade , Padrões de ReferênciaRESUMO
The present review throws a spotlight on new and emerging food safety concerns in view of a well-established food allergen risk arising from global socio-economic changes, international trade, circular economy, environmental sustainability, and upcycling. Food culture globalization needs harmonization of regulations, technical specifications, and reference materials towards mutually recognised results. In parallel, routine laboratories require high-throughput reliable analytical strategies, even in-situ testing devices, to test both food products and food contact surfaces for residual allergens. Finally, the currently neglected safety issues associated to possible allergen exposure due to the newly proposed bio- and plant-based sustainable food contact materials require an in-depth investigation.
Assuntos
Alérgenos , Hipersensibilidade Alimentar , Comércio , Alimentos , Humanos , InternacionalidadeRESUMO
Carbon nanotube (CNT)-based electrodes are cheap, highly performing, and robust platforms for the fabrication of electrochemical sensors. Engineering programmable DNA nanotechnologies on the CNT surface can support the construction of new electrochemical DNA sensors providing an amperometric output in response to biomolecular recognition. This is a significant challenge, since it requires gaining control of specific hybridization processes and functional DNA systems at the interface, while limiting DNA physisorption on the electrode surface, which contributes to nonspecific signal. In this study, we provide design rules to program dynamic DNA structures at the surface of single-walled carbon nanotubes electrodes, showing that specific DNA interactions can be monitored through measurement of the current signal provided by redox-tagged DNA strands. We propose the use of pyrene as a backfilling agent to reduce nonspecific adsorption of reporter DNA strands and demonstrate the controlled formation of DNA duplexes on the electrode surface, which we then apply in the design and conduction of programmable DNA strand displacement reactions. Expanding on this aspect, we report the development of novel amperometric hybridization platforms based on artificial DNA structures templated by the small molecule melamine. These platforms enable dynamic strand exchange reactions orthogonal to conventional toehold-mediated strand displacement and may support new strategies in electrochemical sensing of biomolecular targets, combining the physicochemical properties of nanostructured carbon-based materials with programmable nucleic acid hybridization.
Assuntos
Nanotubos de Carbono , DNA/química , Eletrodos , Nanotecnologia , Nanotubos de Carbono/química , Hibridização de Ácido NucleicoRESUMO
Aptamers are biomimetic receptors that are increasingly exploited for the development of optical and electrochemical aptasensors. They are selected in vitro by the SELEX (Systematic Evolution of Ligands by Exponential Enrichment) procedure, but although they are promising recognition elements, for their reliable applicability for analytical purposes, one cannot ignore sample components that cause matrix effects. This particularly applies when different SELEX-selected aptamers and related truncated sequences are available for a certain target, and the choice of the aptamer should be driven by the specific downstream application. In this context, the present work aimed at investigating the potentialities of asymmetrical flow field-flow fractionation (AF4) with UV detection for the development of a screening method of a large number of anti-lysozyme aptamers towards lysozyme, including randomized sequences and an interfering agent (serum albumin). The possibility to work in native conditions and selectively monitor the evolution of untagged aptamer signal as a result of aptamer-protein binding makes the devised method effective as a strategy for shortlisting the most promising aptamers both in terms of affinity and in terms of selectivity, to support subsequent development of aptamer-based analytical devices.
Assuntos
Aptâmeros de Nucleotídeos , Técnica de Seleção de Aptâmeros , Aptâmeros de Nucleotídeos/metabolismo , Ligantes , Ligação Proteica , Técnica de Seleção de Aptâmeros/métodosRESUMO
An outlook on the current status of different strategies for magnetic micro- and nanosized bead functionalization with aptamers as prominent bioreceptors is given with a focus on electrochemical and optical apta-assays, as well as on aptamer-modified magnetic bead-based miniaturized extraction techniques in food control. Critical aspects that affect interaction of aptamers with target molecules, as well as the possible side effects caused by aptamer interaction with other molecules due to non-specific binding, are discussed. Challenges concerning the real potential and limitations of aptamers as bioreceptors when facing analytical problems in food control are addressed.
Assuntos
Aptâmeros de Nucleotídeos/química , Análise de Alimentos/métodos , Fenômenos Magnéticos , Microesferas , Técnicas Biossensoriais , Microbiologia de Alimentos , Ouro , NanocompostosRESUMO
Two commercial stationary phases allowing both reversed phase mechanism and anion-exchange with different selectivity, i.e. CSH C18 and Atlantis PREMIER BEH C18 AX, were tested for the separation of a complex mixture of 21 fatty acids (FAs) encompassing saturated medium-, long- and very long chain FAs, unsaturated long and very long chain FAs, cis/trans isomers, and isomers of odd- and branched-chain FAs. For this purpose, the role of surface area of stationary phase and the effect of pH of the mobile phase on the retention of the analytes were investigated. Separation was performed by ultra-high-performance liquid chromatography coupled with high resolution mass spectrometry (UHPLC-HRMS). BEH C18 AX was shown to be more versatile and to offer superior retention of these analytes to CSH C18 owing to a higher surface area and anion-exchange capacity up to pH 8.5. The UHPLC system allows shortening analysis time, the chromatographic analysis being accomplished in about 5 min, affording a high throughput of samples without the need for derivatization or ion-pairing reagents compared to techniques based upon gas chromatography approaches or LC. Finally, the application of the BEH C18 AX column using UHPLC-HRMS was demonstrated for the separation and unambiguous identification of FAs of nutritional interest in a dietary supplement sample.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Cromatografia por Troca Iônica/métodos , Ácidos Graxos não Esterificados/análise , Espectrometria de Massas/métodos , Ânions , IsomerismoRESUMO
The nutritional and physicochemical properties of sorghum proteins and starch make the use of this cereal for food production challenging. Sprouting is a cost-effective technology to improve the nutritional and functional profile of grains. Two drying treatments were used after sorghum sprouting to investigate whether the drying phase could improve the protein and starch functionalities. Results showed that the drying treatment at lower temperature/longer time (40 °C for 12 h) extended the enzymatic activity that started during sprouting compared to the one performed at higher temperature/shorter time (50 °C for 6 h). An increased protein hydrolysis and water- and oil-holding capacity were found in the flour obtained by the former treatment. Higher protein matrix hydrolysis caused high exposure of starch to enzymes, thus increasing its digestibility, while worsening the technological functionality. Overall, modulating drying conditions could represent a further way, in addition to sprouting, to improve sorghum flour's nutritional profile.
RESUMO
Prompt cancer diagnosis and treatment represent fundamental aspects to significantly improve patient survival rate. Human epididymis protein 4 (HE4) has recently been identified as promising single serum biomarker of epithelial ovarian cancer with improved diagnostic performances respect to current reference biomarkers. In this study we present the first competitive immunosensing strategy for HE4 determination implemented on magnetic microbeads functionalized with HE4 antigen. A full factorial design and multiple linear regression allowed to find the optimal experimental conditions providing the maximum inhibition rate within the explored domain. Method validation was performed in serum to ensure reliable data to support decision in clinical practice. This method allowed matching the clinically relevant concentration values for the serum biomarker, limits of detection and quantification being 2.8 and 23.0 pM, respectively. Also recovery rate in the 89 ± 7-103 ± 5% range resulted suitable for method applicability for diagnostic purposes.
Assuntos
Biomarcadores Tumorais/sangue , Técnicas Biossensoriais , Carcinoma Epitelial do Ovário/sangue , Técnicas Eletroquímicas , Neoplasias Ovarianas/sangue , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos/análise , Feminino , HumanosRESUMO
Tissue regeneration often requires the use of biocompatible resorbable scaffolds to support the ingrowth of cells from neighboring tissues into a localized tissue defect. Such scaffolds must possess surface molecular cues that stimulate cells to populate the device, the first necessary condition for the formation of a healthy tissue. Chitosan is a natural polymer that has long been tested in biomedical applications because of its high biocompatibility, which can be further increased by modifying its formulation, e.g. adding D-(+) raffinose. We used this formulation in an ad hoc designed 3D printer to create regularly ordered scaffolds, which we then enriched with type IV collagen, an isoform of collagen that is exclusively found in basement membranes. Human epithelial A549 cells were then seeded on control scaffolds or on scaffolds coated with collagen, which was precipitated, or on scaffolds first collagenized and then exposed to either UVB or UVC radiation. Observations by the transmission light microscope, confocal microscope after staining with calcein-AM/propidium iodide, and by environmental scanning electron microscope revealed that collagen-enriched UV-treated scaffolds promoted the attachment of a higher number of cells, which covered a more extensive area of the scaffold, as also confirmed by alamar blue viability assay. Together these data confirm that coating 3D-printed scaffolds made of D-(+) raffinose-modified chitosan with type IV collagen and exposing them to UV light sensibly increases the cell compatibility of scaffolds, making them a better candidate to serve as a tool for the regeneration of epithelia.
Assuntos
Materiais Biocompatíveis/química , Quitosana/química , Colágeno Tipo IV/química , Células Epiteliais/metabolismo , Impressão Tridimensional , Rafinose/química , Alicerces Teciduais/química , Células A549 , Adesão Celular , Materiais Revestidos Biocompatíveis/química , Colágeno/química , Fluoresceínas/química , Humanos , Teste de Materiais , Microscopia Confocal , Polímeros/química , Propídio/química , Regeneração , Temperatura , Engenharia TecidualRESUMO
An outlook on the current status and trends in desorption electrospray ionization-mass spectrometry (DESI-MS), one of the most common spray-based techniques for ambient ionization, is given with a focus on the main advances recently achieved or still in progress regarding studies of surface properties affecting the signal stability and efficiency of the DESI process. Future directions that the field may take in the years to come are discussed, with particular focus on bioanalytical research.
RESUMO
Nowadays, analytical techniques are moving towards the development of smart biosensing strategies for the point-of-care accurate screening of disease biomarkers, such as human epididymis protein 4 (HE4), a recently discovered serum marker for early ovarian cancer diagnosis. In this context, the present work represents the first implementation of a competitive enzyme-labelled magneto-immunoassay exploiting a homemade IoT Wi-Fi cloud-based portable potentiostat for differential pulse voltammetry readout. The electrochemical device was specifically designed to be capable of autonomous calibration and data processing, switching between calibration, and measurement modes: in particular, firstly, a baseline estimation algorithm is applied for correct peak computation, then calibration function is built by interpolating data with a four-parameter logistic function. The calibration function parameters are stored on the cloud for inverse prediction to determine the concentration of unknown samples. Interpolation function calibration and concentration evaluation are performed directly on-board, thus reducing the power consumption. The analytical device was validated in human serum, demonstrating good sensing performance for analysis of HE4 with detection and quantitation limits in human serum of 3.5 and 29.2 pM, respectively, reaching the sensitivity that is required for diagnostic purposes, with high potential for applications as portable and smart diagnostic tool for point-of-care testing.
Assuntos
Biomarcadores Tumorais/sangue , Técnicas Eletroquímicas , Imunoensaio/métodos , Neoplasias Ovarianas/diagnóstico , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos/análise , Algoritmos , Biomarcadores Tumorais/normas , Calibragem , Feminino , Humanos , Imunoensaio/normas , Internet das Coisas , Limite de Detecção , Magnetismo , Sistemas Automatizados de Assistência Junto ao Leito , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos/normasRESUMO
Microalgal species growing in marine and aquaculture environments can be responsible for harmful events because of their ability to produce potent natural toxins that can accumulate in edible mollusc species. Their consumption can cause severe illness and even be lethal. The European Union provides comprehensive regulations covering various general food safety aspects to manage the risk of contamination in shellfish farms. Many analytical methods have been proposed to evaluate algal toxins presence in the environment and in food products, for conducting surveillance studies of the main molluscs production sites and, where necessary, immediate monitoring of possible contamination of shellfish. In this work, a one-year analytical surveillance study was carried out to verify the possible presence of algal biotoxins in molluscs from a Mediterranean breeding area. Water and molluscs were sampled from a district of the North-East coast of Sicily, consisting of a unique brackish ecosystem of two lakes connected to each other and to the sea by narrow canals. Water samples were collected to investigate phytoplankton i by microscope analysis to assess the presence of potentially toxin-producing species, such as Pseudo-nitzschia spp, Alexandrium spp and Gonyaulax spinifera, although the presence of toxic phytoplankton has never reached alert levels. Mussels and clams samples were submitted to analysis of paralytic shellfish poisoning toxins, amnesic shellfish poisoning toxins and lipophilic toxins by liquid chromatography-based methods Only a few yessotoxins were detected, having concentrations always below the regulation limits. An existing liquid chromatography-tandem mass spectrometry-based multiresidue method for lipophilic biotoxins was adopted and extended to cover emerging biotoxins such as cyclic imines. The performance of the analytical method for Gymnodimine A and Spirolide 13-desMeC was assessed, obtaining respective quantitation limits of 20 and 10 µg kg-1, a precision always lower than 13% and trueness in the 81-120% range. Method applicability was confirmed using certified materials and a naturally contaminated sample.