Browsing by Author "Tagarelli, Antonio"

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Development and optimization by experimental design of solid phase microextraction gas chromatography triple quadrupole mass spectrometry methods in aqueous matrices
(2012-11-30) Monteleone, Marcello; Tagarelli, Antonio; Gabriele, Bartolo; Bartolino, Roberto
Il presente lavoro di tesi relativo all‘attività di ricerca svolta durante il triennio di dottorato ha riguardato la messa a punto di metodi analitici per la determinazione di analiti in due distinte aree di interesse. La prima di ambito clinico ed ha riguardato la quantificazione in urina di metaboliti riconosciuti come marker in diagnostica clinica. In particolare ci si occupati della sarcosina come biomarker del tumore alla prostata, e di tre acidi: acido omovanillico (HVA), acido vanilmandelico (VMA) ed acido 5-idrossindoloacetico (5-HIAA) come marker urinari del neuroblastoma. Il secondo ambito di lavoro ha riguardato la quantificazione di inquinanti in matrici acquose, vale a dire carbammati ed acidi perfluoroalchilici. In particolare gli analiti, previa derivatizzazione con alchilcloroformiati (eccetto i carbammati), sono stati estratti dalle matrici acquose (acqua e urina) tramite la tecnica della microestrazione in fase solida (SPME) e successivamente analizzati mediante un gascromatografo con analizzatore di massa a triplo quadrupolo (GC-QqQ-MS). Le variabili significative della microestrazione in fase solida in ciascun metodo sono state ottimizzate tramite l‘approccio multivariato dell‘ ― Experimental Design‖. L‘utilizzo della tecnica SPME ha consentito di poter estrarre gli analiti direttamente dal campione da analizzare minimizzando i tempi di preparazione dello stesso e riducendo l‘uso di solventi organici, ottenendo metodi poco costosi e basso impatto ambientale. L‘utilizzo dello spettrometro di massa triplo quadrupolo, ha consentito di raggiungere livelli di sensibilità molto elevati e, nel contempo, di identificare gli analiti con maggiore sicurezza. In tutti i metodi sviluppati, sono stati ottenuti ottimi risultati in termini di linearità accuratezza e precisione. Anche i valori dei limiti di rilevabilità (LOD) e dei limiti di quantificazione (LOQ) ottenuti in ciascun metodo possono essere considerati soddisfacenti.
Development and optimization of analytical protocols based on microextraction techniques for clinical screening and environmental control
(2019-03-21) Elliani, Rosangela; Andò, Sebastiano; Tagarelli, Antonio
The development of analytical protocols for the determination of analytes at trace levels in complex matrices (e.g. biological fluids or contaminated water) is a crucial point for the environmental assessment and monitoring as well as for scientific research in the field of disease biomarkers. An essential part of analytical method development is represented by sample preparation due to its significant impact on most of the subsequent steps and the data quality. In recent years, the application of pro-ecological, automated, solvent-free sample preparation approaches or techniques employing a minimal amount of solvents or safe and non-toxic extractants has become one of the most popular research topics in analytical chemistry. In this context, microextraction techniques represent a suitable choice for the extraction of analytes from complex matrices because these techniques use less organic solvent and allow to perform in a single step extraction and concentration of analytes. Moreover, the use of microextraction techniques for sample preparation reduces the number of errors that commonly result from multi-stage procedures, and limits the negative impact on the environment and the health of analytical chemists performing laboratory work. The goal of this Ph.D project was the development and optimization of analytical methods based on the use of microextraction techniques for the assay of disease biomarkers and environmental contaminants in biological fluids and environmental matrices. The microextraction techniques employed in this thesis were solid phase microextraction (SPME) and microextraction by packed sorbent (MEPS). SPME was used to evaluate the applicability of a new fiber (PDMS/DVB/PDMS) as analytical sampling tool for investigation in raw human urine. The PDMS/DVB/PDMS fiber was exploited to develop a DI-SPME-GC-MS method for the assay of polycyclic aromatic hydrocarbons (PAHs) with 2-6 aromatic rings in untreated human urine samples. Moreover, in the light of the increasing demand of faster and easier protocols allowing the assessment of disease biomarkers, SPME was applied to develop a reliable and rapid GC-MS approach for the determination of polyamines in human urine. Indeed, polyamines are widely recognized as among the most important cancer biomarkers for early diagnosis and treatment. SPME was also applied for the extraction of nine phthalates monoesters in urine samples. These compounds are important metabolites of phthalates and their assay can reliably rank exposures to phthalates over a period. MEPS was used to extract organophosphate ester flame retardant in aqueous matrices and, again, monoesters phthalates in urine. In both methods, in order to improve method sensitivity, programmed temperature vaporization (PTV) was chosen as gas chromatographic injection technique. For polyamines and phthalates monoesters, a prior derivatization step with suitable reagents was carried out before gas chromatographic analysis so as to improve chromatographic elution and resolution by decreasing volatility and polarity of analytes. Derivatization reaction was performed directly in aqueous samples using alkyl chloroformates. The combined use of alkyl chloroformate as derivatizing reagent and SPME for analyte extraction was chosen to develop a simple protocol involving minimal sample handling and no consumption of toxic organic solvents. The variables affecting the different steps of the proposed protocols were optimized by the multivariate approach of experimental design which has allowed for the simultaneous investigation of the different factors in the entire experimental domain and the possible synergic effects between variables. In this thesis, experimental design was used to optimize the parameters influencing SPME extraction, MEPS extraction, PTV process and derivatization reaction. Gas chromatographic analyses were carried out using a GC-QqQ-MS instrument in selected reaction monitoring (SRM) acquisition which has allowed to obtain reconstructed chromatograms with well-defined chromatographic peaks and to achieve high specificity through the selection of appropriate precursor-product ion couples, improving the capability in analyte identification. Finally, during the period as visiting Ph.D student at University Duisburg-Essen, Faculty of Chemistry, Instrumental Analytical Chemistry, the object of research activity, coordinated by Professor Torsten C. Schmidt, concerned the extraction of fatty acid methyl esters (FAMEs) in wastewater by solid phase microextraction arrow (SPME arrow).
Food control quality by isotope dilution liquid chromatography-tandem mass spectrometry
(2014-06-13) Benabdelkamel, Hicham; Tagarelli, Antonio; Gabriele, Bartolo
The late part of the 20th century the advancement of knowledge regarding nutrition and disease prevention provided an opportunity for individuals to affect their own health. This expanding body of information helped people to understand how the environment and their own behaviour affected their body. People now had powerful tools to help maintaining and protecting their health. The understanding of how our diet affects our well being has dramatically changed the lifestyles and attitudes of people, who began to make menu and purchasing decisions based on how foods would affect their heath. A shift toward healthier lifestyles and healthier diets began. Food processors and marketers had to refocus their efforts from promoting foods for pleasure to promoting foods that fit in to a healthy diet. Primarily, the focus was on reducing fat and cholesterol in the diet and supplementing vitamins and minerals. Research began to demonstrate the presence of various phytochemicals in wine and juice make it from fruits and vegetables( such as stilbenes, falvoniods, polyphenols…) and specially in olive oil as ( polyphenol in dialdehyed form: oleochanthal, hydroxyoleocanthal..) These compounds have come to be known as nutraceuticals. The list of nutraceuticals present in wine, fruit drink and olive oil that are believed to have positive biological properties has been expanding. Food processors and developers have become very interested in exploiting these nutraceuticals for the production of foods that are not only part of a healthy diet but also improve the consumer’s health in another specific way. These foods have become known as “functional foods” means quality marker. The aim of this thesis has been to develop a analytical methods to determine the concentration of a group of these nutraceuticals in food , such as, quantitative determination of resveratrol in wine, pterostilbene in blueberry juice and dialdehyde form in olive oil using a sensitive high-performance liquid chromatographic separation method coupling with tandem-mass and isotope dilution to order to optimal the conditions for the analysis method, such as extraction procedure, matrices, column, quality controls, wavelength, mobile phases, run time, optimal separation (gradient, retention times), temperature, capillary voltage, cone voltages, vacuum and labelled internal standards, resulting in the best sensitivity and selectivity,The goodness and satisfactory of the method was performed according to, containing linear measuring range, quantification, lower limit of quantification (LLOQ), lower limit of detection (LLOD), quality controls, precision(RSD %), accuracy, recovery, stability and matrix effects. In conclusion, the described high-performance liquid chromatographic separation method with tandem-mass spectrometry detection and isotope dilution showed a satisfactory overall analytical performance well suited for applications in food quality control.