Project 1: Chemical characterization of breath sample for a non-invasive monitoring of human health
In recent years, exhaled breath analysis has become increasingly attractive for its potential use as an easy, painless and non-invasive tool to monitor physiological and pathological conditions as well as exposure to environmental contaminants. A main advantage of this approach is that it can be used on people of all ages and conditions (e.g. newborns, infants and mechanically ventilated patients) reducing the problems associated with blood sampling (e.g. acceptance from patients, risk of infections, production of potentially infected waste, and need for trained personnel working in dedicated environment). Several studies highlighted the potential of breath analysis as an innovative diagnostic tool since the chemical composition of breath is related to the occurrence of diseases, suggesting the diagnostic utility of breath biomarkers, often complementary or even alternative to those of blood and urine. For example, acetone is potentially useful for monitoring patients suffering from diabetes and heart failure, hydrocarbons and aldehydes for monitoring abnormal lipid peroxidation. Analytical protocols based on thermal desorption coupled to gas-chromatography and mass spectrometry will be normally employed in several pilot studies, conducted in collaboration with medical partners, to monitor the chemical composition of breath sample collected from patients suffering from diabetes, heart failure and asthma.
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Project 2: Development of MS-based analytical approaches to monitor oxidative stress and inflammation in biological fluids
Immune-inflammatory response is regulated among other cells by macrophages through the secretion of various soluble molecules, as cytokines, chemokines and lipid mediators [10]. Lipid mediators, also termed oxylipins, are generated from both n-3 and n-6 polyunsaturated fatty acids (PUFAs) through enzymatic (e.g., prostanoids, epoxy, and hydroxy fatty acids) and non-enzymatic (e.g., isoprostanes) oxidation reactions. PUFAs act as precursors of pro-inflammatory, anti-inflammatory, and specialized pro-resolving lipid mediators (SPMs) through enzymatic oxidation reactions. Pro-inflammatory mediators released at the very beginning of inflammation foster the appearance of its classic signs, whereas the switch to lipoxygenase-derived anti-inflammatory and SPMs leads to a natural resolution of inflammation. PUFAs can also undergo free radical-mediated oxidation, thus generating isoprostanes, currently known as gold standard biomarkers of oxidative stress. The analysis of oxylipins may furnish a clear picture of the oxidant and inflammatory cascade, thus revealing key information about the pathogenesis and pathophysiology of many diseases. In this scenario, we have developed a powerful in-house MS-based platform that couples the new micro-extraction by packed sorbent (MEPS) technique with ultra-high performances liquid chromatography tandem mass spectrometry for the detection of 60 oxylipins in a single run. In the recent years, we have been working on MS-based targeted metabolomics for the determination of oxylipins in different research area, such as cardiovascular, neurological, and neurodegenerative diseases, the evaluation of metabolic adaptation to extreme physical exercise or altitude induced-hypoxia, the study of fetal-to-newborn transition and of host-pathogen response upon viral infection, as in the case of the current COVID-19 pandemic crisis.
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Project 3: Development of analytical approaches to monitor volatile- and semi-volatile organic compounds emitted from plastic debris
Environmental pollution associated to plastic debris is gaining increasing relevance not only as a threat to ecosystems but also for its possible harmful effects on biota and human health. Plastic debris exposed to environmental weathering are prone to degradation mainly due to photo-oxidative, thermal, and hydrolytic processes. Such degradation processes lead to progressive polymer fragmentation into microplastics as well as the release of toxic volatile- and semi-volatile compounds resulting from the chemical degradation induced in particular by heat and UV-light. As example, degradation of polyolefins generates compounds belonging to the families of lactones, esters, ketones, and carboxylic acids. Most of these compounds play a key role in atmospheric chemistry and represent also a source of toxic leachate with negative impact on biota and on human health. Analytical protocols based on thermal desorption coupled to gas-chromatography and mass spectrometry will be normally employed to monitor degradation products. In this scenario, micro-extraction tools (e.g. needle trap micro extraction and stir bar sorptive extraction) will be tested too, with the aim to reduce the amount of plastic debris required for the chemical characterization. Developed methods will be employed to monitor the degradation products emitted from reference materials that will be subjected to artificial aging processes simulating the real environmental conditions.
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Project 4: Mass spectrometry-based assay for the determination of natriuretic peptides in biological fluids
Cardiac natriuretic peptides (NPs), i.e., ANP, BNP, and CNP, are hormones initially synthesized from cardiomyocytes, as prohormones. Once they are released from the heart, prohormones are split into two fragments: a longer N-terminal fragment (NT-proBNP and NT-proANP) and a shorter C-terminal fragment (BNP and ANP). In pathological conditions as heart failure (HF), heart induces the stretching of myocytes due to pressure overload, leading to the release of NPs in blood. Thus, NPs plasma levels increase progressively with the severity of HF and represent the main biomarkers for prognosis and diagnosis of HF. NPs are normally determined in blood using commercial immunoassay kits. This approach is affected by cross-reactions due to the presence of several NP forms showing homologous structures, as those with oxidized methionine. My project aims to the development of a reliable and innovative analytical workflow based on targeted top-down proteomics and UHPLC-MS/MS analysis for the determination of NPs in saliva and plasma samples. The ability to rapidly quantify multiple molecular species with high specificity and accuracy makes mass spectrometry suitable for determining the different NPs forms. Ultrafiltration, fractional protein precipitation, and micro-extraction by packed sorbent (MEPS) will be tested as sample treatment procedures. Finally, the optimized analytical protocol will be employed for the longitudinal monitoring of patients suffering from HF, and for a systematic comparison between the analytical performances of the proposed method and immunoassays used in the clinical routine for the quantification of NPs.
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