A comprehensive investigation of sensitive ecosystems in South Florida with the main goal of determining the identity, spatial distribution, and sources of both organic biocides and trace elements in different environmental compartments is reported. This study presents the development and validation of a fractionation and isolation method of twelve polar acidic herbicides commonly applied in the vicinity of the study areas, including e.g. 2,4-D, MCPA, dichlorprop, mecroprop, picloram in surface water. Solid phase extraction (SPE) was used to isolate the analytes from abiotic matrices containing large amounts of dissolved organic material. Atmospheric-pressure ionization (API) with electrospray ionization in negative mode (ESP-) in a Quadrupole Ion Trap mass spectrometer was used to perform the characterization of the herbicides of interest. The application of Laser Ablation-ICP-MS methodology in the analysis of soils and sediments is reported in this study. The analytical performance of the method was evaluated on certified standards and real soil and sediment samples. Residential soils were analyzed to evaluate feasibility of using the powerful technique as a routine and rapid method to monitor potential contaminated sites. Forty eight sediments were also collected from semi pristine areas in South Florida to conduct screening of baseline levels of bioavailable elements in support of risk evaluation. The LA-ICP-MS data were used to perform a statistical evaluation of the elemental composition as a tool for environmental forensics. A LA-ICP-MS protocol was also developed and optimized for the elemental analysis of a wide range of elements in polymeric filters containing atmospheric dust. A quantitative strategy based on internal and external standards allowed for a rapid determination of airborne trace elements in filters containing both contemporary African dust and local dust emissions. These distributions were used to qualitative and quantitative assess differences of composition and to establish provenance and fluxes to protected regional ecosystems such as coral reefs and national parks.

Lo scopo della tesi è di caratterizzare i rischi per la salute umana, che derivano dall’inquinamento delle acque (acque sotterranee) e dal cibo (riso) tramite l’applicazione e lo sviluppo di nuove tecniche analitiche. La tecnica “diffusive gradients in thin films (DGT)” con resina a base di ferrihydrite, è stata studiata per l'accumulo delle forme inorganiche (As(III) e As(V)) e organiche (DMA(V) e MMA(V)) di Arsenico in matrici acquose. I DGT sono stati applicati su campioni di acque sotterranee, raccolti in sei diverse città del Nord Italia, dove la concentrazione di As è molto alta. Antimonio (Sb) è associato all’As in diversi studi perché le proprietà fisiche e chimiche di questi due elementi sono simili, ed esso è stato recentemente riconosciuto come contaminante dell'acqua. In questa tesi per la prima volta sono state riportate le prestazioni dettagliate dei DGT con ferrihydrite, impiegati in soluzioni acquose di Sb(III) e Sb(V). L’analisi delle forme chimiche di Sb(III) e Sb(V) in campioni acquosi è stata realizzata con l’applicazione della diluizione isotopica. In generale il riso, a differenza di altri prodotti alimentari di origine terrestre, contiene quantità significative di Arsenico inorganico. L’analisi della speciazione di As è stata realizzata in 70 campioni di riso italiano, rappresentativo di diversi tipi di coltivazione. Le forme più abbondanti nel riso erano As(III) e DMA(V). Inoltre è stato interessante studiare la localizzazione dell’As nel chicco di riso, sottoposto a diversi processi di lavorazione (risone, riso integrale e lavorato con o senza parboiling), perché sia la distribuzione che la presenza delle diverse forme chimiche nel chicco sono fattori chiave che controllano la biodisponibilità del contaminante. La distribuzione dell’ As nei chicchi di riso di due varietà (Gladio e Ronaldo) sottoposti a diversi processi, è stato determinato con Laser Ablation ICP-MS. In realtà il contenuto di As era più alto nei chicchi di riso non-parboiled rispetto a quello parboiled. La relazione tra intensità dell’As e le varie parti del chicco di riso ha rivelato che i livelli di As diminuisco dalla parte esterna verso la posizione centrale, mentre i valori di As sembrano essere simili tra la parte media e interna dei non parboiled. / The aim of this thesis was to explore new analytical techniques as well as to carry out further characterisations of human health risks, which derive from water pollution, in particular groundwater, and food, in particular rice. The prevention of water pollution is an environmental aspect, that includes monitoring of both natural enrichment and outside pollution with routine analysis but also with new techniques, e.g. the application of passive sampling techniques and advanced technologies. The diffusive gradients in thin films (DGT) technique with ferrihydrite adsorbent, has been investigated for the accumulation of different species of Arsenic (As), like Inorganic Species (arsenite and arsenate) and Organic Arsenic (dimethylarsinic and monomethylarsenate) in aqueous matrix. To evaluate the performance of DGT method for accumulation of arsenic species, after deployment in synthetic solutions, DGT devices were carried out on groundwaters collected in six different towns in the North of Italy, where the As concentration is very high. Recently, health effects at arsenic exposures have been observed in areas where levels of inorganic As in drinking water are not excessive. Antimony (Sb) is associated to As in several studies because the physical and chemical properties of these two elements are similar, and it has been recently recognized as water contaminant. In this thesis I reported for the first time detailed performance characteristics of the Fe-oxide gel associated to DGT devices deployed in known aqueous solutions of trivalent and pentavalent Sb. Speciation analysis of Sb(III) and Sb(V) in aqueous samples was performed through extraction and on-line determination of isotope dilution concentration after a chromatographic separation. Generally rice, unlike food products of terrestrial origin, contains significant amounts of inorganic arsenic. Recently some Government Organizations (e.g. EFSA) debated the possibility to set an upper limit for total and inorganic arsenic in rice. Arsenic speciation was realized in 70 Italian rice samples from different representative cultivation conditions. The most abundant forms in rice were As(III) and DMA(V). After that, it was fundamental to investigate the localization of As in rice grains in different processes (raw, brown and milled rice with or without parboiling technique), because both speciation and distribution throughout the grain are key factors controlling bioavailability of the contaminant. The As distribution in rice grains of two varieties (Gladio and Ronaldo) from different processes, was determined by LA-ICP-MS. The distribution of As varied between the various parts of the grains (exterior, medium and interior part). During parboiling, the partial boiling of food as the first step in the cooking process, arsenic might have released from the grain to the boiling water. Thus, parboiling of rice grain may reduce the magnitude of arsenic intake in human body. Actually the As content was higher in non-parboiled rice grain than in parboiled rice. The relationship between As intensities and the different parts of rice grain revealed that As levels decreased from the external part towards the middle position, and then the intensity values seem to be similar between medium and internal part in non parboiled products.

Cellular therapy is emerging as a clinically viable strategy in the field of solid organ transplantation, where it is expected to reduce the dependency on conventional immunosuppression. This has produced a demand for highly sensitive methods to monitor the persistence and tissue distribution of administered cells in vivo. However, tracking cells presents significant challenges. In many cases transplanted cells are autologous with the immune system of the transplant recipient, and hence are invisible to typical methods of detection. To enable their differentiation, the cells must be labelled with a suitable, non-toxic and long lifetime label, prior to their administration to patients. In addition, administered cells represent only a small fraction of the recipient's endogenous cells, which necessitates the use of an extremely sensitive detection method. Laser ablation – inductively coupled plasma – mass spectrometry (LA-ICP-MS) is an exquisitely sensitive analytical technique, capable of imaging trace elements in complex samples, at high spatial resolution.

Die Analyse von organischen Polymeren mittels Massenspektrometrie mit induktiv gekoppeltem Plasma nach Laserablation unterliegt starken Matrixeffekten. Mögliche Korrekturmodelle sollten basierend auf der tatsächlich ablatierten Elementmasse sowie der Signalintensität des Kohlenstoffisotops C-13 (13C-Korrektur) entworfen werden. Damit die 13C-Korrektur erfolgreich verlaufen kann, ist ein direkter Zusammenhang zwischen ablatiertem Kohlenstoff und gemessenem 13C-ICP-MS Signal erforderlich. Dies wurde überprüft, indem unter Anwendung der konfokalen Mikroskopie das Ablationskratervolumen bestimmt und der ablatierte Kohlenstoff berechnet wurde. Dieser Zusammenhang konnte sowohl für undotierte, wie auch für elementdotierte Polymere und die Verwendung verschiedener Ablationsgase (Helium, Argon und Sauerstoff) ermittelt werden. Eigens hergestellte polymere elementdotierte Standardmaterialien wurden hinsichtlich der Matrixeffekte charakterisiert. Als größter Einflussfaktor stellte sich die unterschiedliche Ablationsrate bei der Ablation verschiedener Polymere heraus. Unter Anwendung der tatsächlich ablatierten Elementmasse sowie unter Anwendung der 13C-Korrektur konnte die Matrixabhängigkeit deutlich verringert werden.

Le sélénium est un micronutriment essentiel pour des nombreux organismes vivants, y compris l’homme. Son rôle est lié à sa présence dans des sélénoprotéines sous forme d’un acide aminé, génétiquement encodé – la sélénocystéine. Il y a 25 sélénoprotéines encodées dans le génome humain. Leurs fonctions, la cinétique et la hiérarchie d'expression se trouvent au cœur des problématiques de recherche concernant le sélénium et la santé humaine. Il existe également un autre type de protéines où le sélénium est inséré par un remplacement partiel du soufre dans la méthionine mais aussi, potentiellement, dans la cystéine. Ces protéines suscitent l’intérêt dans les sciences de nutrition comme source de sélénium biodisponible dans l’alimentation naturelle et supplémentée. L'objectif de cette thèse a été la mise au point de méthodologies analytiques visant la spéciation du sélénium incorporé dans les protéines à l’échelle du protéome entier. Une procédure inédite a été développée pour la détection globale de protéines séléniées dans des gels d’électrophorèse bidimensionnelle par l’imagerie d’ablation laser ICP MS (spectrométrie de masse plasma à couplage inductif) permettant de s’affranchir de l’utilisation de l’isotope radioactif 75Se. Les autres avancées comprennent la mise en place d’un couplage robuste de HPLC capillaire avec l’ICP MS pour la détection des sélénopeptides dans des microvolumes de digestats trypsiques des protéines extraites du gel ainsi que la mise en place des protocoles d’identification des protéines séléniées par la spectrométrie de masse électrospray en tandem utilisant la trappe orbitale (Orbitrap). Les méthodes développées ont permis (i) la caractérisation de la part du protéome sélénié contenant la sélénocystéine chez la levure séléniée, (ii) l’identification des protéines majeures qui accumulent le sélénium dans le blé, et (iii) le dosage semi quantitatif et la caractérisation globale des sélénoprotéomes (GPx1, GPx4, TRxR1, TRxR2, Sel15kDa) dans les lignées cellulaires. / Selenium is an essential micronutrient for many living organisms including man. Its role is related to selenoproteins which contain genetically encoded selenocysteine. There are 25 selenoproteins encoded in the human genome. Their function, expression kinetics and hierarchy have been a topic of intense research in life sciences. There is another type of proteins which contain selenium inserted non-specifically by partly replacing sulphur in methionine and, potentially, cysteine. They are of interest in nutrition science as source of bio-available selenium in natural and supplemented foods. The goal of this Ph.D. was the development of methodologies for the analysis of selenium-containing proteins on the entire proteome scale. A novel procedure was developed for their global detection in 2D electrophoretic gels par laser ablation inductively coupled plasma mass spectrometry (ICP MS) imaging permitting to avoid the use of the radioactive 75Se. The other developments included (i) a robust capillary HPLC – ICP MS coupling allowing the detection of Se-containing peptides in microliter volumes of the digests of proteins extracted from the gel and (ii) protocols allowing the targeted identification of the Se-containing proteins by a parallel capillary HPLC - electrospray Orbitrap MS/MS. The methods developed allowed (i) the characterisation of the selenocystein-containing part of the selenoproteome of Se-enriched yeast, (ii) identification of the major Se-accumulating proteins in wheat, and (iii) semiquatitive analysis and global identification of the selenoproteomes (GPx1, GPx4, TRxR1, TRxR2, Sel15kDa) expressed in different human cell lines.

No description available.

In dieser Arbeit wurde eine neue Markierungsstrategie von Antikörpern mit dem Markierungsreagenz MeCAT (Metal Coded Tag) unter physiologischen Reaktionsbedingungen, sowie deren Anwendung in einem 8-fach Multiplex-Immunoassay von in Formalin-fixierten und in Paraffin-eingebetteten Gewebeschnitten entwickelt. Für eine aussagekräftige LA-ICP-MS Detektion von MeCAT-modifizierten Antikörpern, wurde eine Standardisierung für biologische Proben auf NC-Membranen, basierend auf einer homogenen Aufbringung eines internen Standards und Kalibrierstandards durch einen kommerziell verfügbaren Tintenstrahldrucker entwickelt und mit der ICP-MS Analyse von Lösungen evaluiert. Die LA-ICP-MS wurde in zwei 8-fach Multiplex-Immunoassays von Tissue Micro Arrays vom Prostatakarzinom und in Maushirngewebeschnitten zur Einschätzung von neurogenerative Erkrankungen erfolgreich eingesetzt werden. Es konnte hierbei gezeigt werden, dass das Nachweisvermögen, der hier entwickelten Methode bereits ausreicht, um die gängigen klinischen Biomarker mit guter Ortsauflösung nachzuweisen. / In this work a new tagging strategy of antibodies with the tagging reagent MeCAT (Metal Coded Tag) was developed under physiological reaction conditions. Their application was proved in an 8-fold multiplex immunoassay of formalin-fixed and paraffin-embedded tissue sections. For a significant LA-ICP-MS detection of MeCAT tagged antibodies standardization for biological samples owere developed. The standardization based on a homogeneous deposition onto the NC membrane via conventional CD-ink-jet printer was validated in addition with the ICP-MS analysis of solutions. The internal standardization of LA-ICP-MS was successfully applied in two 8-fold multiplex immunoassays for Tissue Micro Arrays (TMA) of prostate cancer and for detection of biomarkers for neurodegenerative diseases in mouse brain tissue sections. In both examples it could be shown that the detection capability of the new tagging strategy in combination with the printing standardization allows the detection of the clinical biomarker with good spatial resolutions.

Bei der Schwefelquantifizierung in Kupfer und anderen reinen Metallen zeigte sich in der Vergangenheit eine mangelnde SI-Rückführung und zusätzlich inkonsistente Ergebnisse, wenn verschiedene Methoden verglichen wurden. Um diesen Mangel zu beheben ist ein Referenzverfahren erforderlich, welches SI-rückführbare Werte mit einem zuverlässigen Unsicherheitsbudget ermöglicht. In dieser Studie wurde ein entsprechendes Referenzverfahren zur Quantifizierung von Gesamtschwefel in Kupfer basierend auf der induktiv gekoppelten Plasma-Massenspektrometrie und der Isotopenverdünnungsanalyse (ICP-IDMS) entwickelt. Um diese Probleme zu lösen wurde mit Hilfe der Ionenaustauschchromatographie ein Schwefel-Matrix-Trennverfahren entwickelt. Dieses Trennverfahren wurde mit ICP-IDMS kombiniert, um Schwierigkeiten mit der Kalibrierung zu lösen und fehlende metrologische Konzepte einzuführen. So wurden die in diesem Projekt erzielten IDMS-Messwerte für die Kalibrierung von GDMS und LA-ICP-MS verwendet, beides Verfahren die im industriellen Einsatz üblich sind. Dadurch konnten mit beiden Routineverfahren zuverlässige Ergebnisse erzeilt warden, die zudem auf SI rückführbar sind. Darüber hinaus wurde ein auf der LA-ICP-IDMS basierendes Verfahren entwickelt, um den Probenvorbereitungsschritt von ICP-IDMS mit Schwefel-Matrix-Trennung zu reduzieren. Die Vorteile dieser Methode sind ein geringerer Arbeits- und Zeitaufwand, die SI- Rückführung der Messergebnisse und eine für LA-ICP-MS vergleichsweise hohe Genauigkeit. Die Schlüsselrolle hierbei spielte der innovative Einsatz von Polyethylenfritten als Trägermaterial der aufgelösten Probe. Dadurch war die Quantifizierung von Schwefel in Kupferproben mittels LA-ICP-IDMS möglich. Die wesentlichen Parameter wie Absorptionseffizienz der Fritten und Matrixeffekt wurden untersucht. Das entwickelte Verfahren konnte mit Hilfe der ICP-IDMS vollständig validiert werden. / Sulphur quantification in copper and other pure metals in the past revealed a lack of SI-traceability and also showed inconsistent results, when different methods are compared. Therefore, a reference procedure is required to enable SI-traceable measurement results accompanied by a sound uncertainty budget. In this study, such a procedure was developed for the quantification of total sulphur in copper using inductively coupled plasma-isotope dilution mass spectrometry (ICP-IDMS). For solving these problems ion exchange chromatography was applied, and a sulphur-matrix separation procedure was developed. This procedure was combined with ICP-IDMS to solve difficulties with the calibration and to realize metrological concepts. An application of the IDMS procedure was realized by using the measurement results of specific copper samples values for calibrating glow discharge mass spectrometry (GDMS) and laser ablation ICP-MS (LA-ICP-MS). Both techniques are considered routine techniques. Thus, they could provide reliable results which are traceable to the SI. Additionally, a procedure based on LA-ICP-IDMS was developed to significantly reduce the sample preparation step of ICP-IDMS with sulphur-matrix separation. This procedure is less laborious and the measurement results are still SI traceable and offer a comparatively high accuracy for LA-ICP-MS. Key for this development was the innovative application of polyethylene frits as support material for the dissolved sample. Thus, the quantification of sulphur in copper samples by LA-ICP-IDMS could be realized. The essential parameters are investigated such as the absorption efficiency of the frit and matrix effects. The developed procedure was fully validated by means of the ICP-IDMS results.