Federico Rubino | Università degli Studi di Milano - State University of Milan (Italy) (original) (raw)

Papers by Federico Rubino

RAZIONALE. La melatonina (MLT) \ue8 un potente neurormone prodotto in tracce dal triptofano nella... more RAZIONALE. La melatonina (MLT) \ue8 un potente neurormone prodotto in tracce dal triptofano nella ghiandola pineale dei mammiferi e coinvolto in molte funzioni fisiologiche: la regolazione dei ritmi circadiani di sonno-veglia, l\u2019immunostimolazione, la difesa antiossidante e dai tumori (1). Le piante producono MLT con funzioni simili: aumentare la resistenza allo stress, regolare i ritmi circadiani, promuovere la germinazione e la maturazione. Anche lieviti come Saccaromyces cerevisiae e altri ceppi producono MLT durante la vinificazione. Lo scopo della ricerca \ue8 ottimizzare una procedura analitica per identificare e quantificare l\u2019insieme dei metaboliti indolici del triptofano (indoloma) in matrici complesse vegetali e animali, per selezionare microrganismi alto-produttori di MLT per l\u2019alimentazione, misurare la farmacocinetica di MLT somministrata secondo diverse vie e formulazioni farmaceutiche a confronto con un kit ELISA e studiare l\u2019indoloma plasmatico di soggetti sottoposti a deregolazione dei cicli di buio-luce. MATERIALI E METODI. L\u2019analisi impiega un sistema di cromatografia liquida \u2013 spettrometria di massa composto da un cromatografo Dionex UltiMate\uae 3000 LC Systems (con autocampionatore, pompa binaria e termostato) e da uno spettrometro di massa a triplo quadrupolo AB Sciex 3200 QTRAP\uae. La colonna UPLC \ue8 una Agilent Zorbax\uae Bonus-RP 80A Amide (3x100 mm; 3,5 \u3bcm). Come standard interni sono utilizzati la 5-F-Triptamina per tutti gli indoli, il D5-Ar-Triptofano e la OCD3-MLT per i rispettivi analiti. La separazione cromatografica \ue8 ottenuta con un gradiente di acido formico allo 0,1% e formiato di ammonio 5 mM in acqua (A) e acido formico 0,1% in metanolo (B), flusso 0,4 ml / min, volume di iniezione 10 \u3bcl, tempo totale di analisi 10 min. La preparazione dei campioni impiega la precipitazione delle proteine con acetonitrile - 0,1% acido formico e la SPE con cartucce Supelco HybridSPE-Phospholipid (30mg, 1 ml). RISULTATI. La scelta delle transizioni analitiche e delle condizioni strumentali ottimali \ue8 stata ottenuta con lo studio preliminare mediante infusione in modalit\ue0 di ionizzazione positiva di 11 composti (soluzioni singole, ~0,8 nM). Sono state misurate, con la funzione Ramp del software Analyst, le curve del Declustering Potential (DP) dello strumento (intervallo di \uf044V tra 0 e 70-90 V) e le curve di abbondanza dei frammenti in funzione dell\u2019energia di collisione (curve di breakdown nell\u2019intervallo di \uf044V tra 0 e 50-70 V). Tutte le elaborazioni sono state eseguite, a partire dai dati del software Analyst esportati come file .txt, in fogli di calcolo personalizzati (2), che calcolano i valori dei potenziali strumentali corrispondenti alla resa massima dei frammenti ionici identificati come reporter analitici, che sono stati poi utilizzati nel metodo finale. Il metodo cromatografico sviluppato separa e analizza 11 composti indolici in 10 min, con curve di calibrazione lineari (R2>0,998) nei due intervalli utili di concentrazione 0,012-1,25 ng (curva bassa) e 1,25-10 ng (curva alta). La presenza di triptofano ad elevate concentrazioni (69,7 e 18,4 \ub5g/ml, rispettivamente) in terreni YPD e YNB utilizzati per la fermentazione di lieviti \ue8 stata confermata, mentre tutti gli altri indoli ricercati sono risultati assenti. In estratti metanolici di farina di semi di soia \ue8 stato misurato triptofano libero (2,72 e 3,12 mg/g) e sono state identificate tracce di N-acetil-5OH-Triptamina (0,16 e 0,16 \ub5g/g) e di MLT (5,65 e 0,95 \ub5g/g). Questo metodo ha inoltre consentito di quantificare MLT nel plasma per lo studio di diverse formulazioni farmaceutiche e per la valutazione dei suoi ritmi circadiani nell\u2019uomo. 1. Lionella Palego, Laura Betti, Alessandra Rossi, Gino Giannaccini. Journal of Amino Acids Volume 2016, Article ID 8952520, 13 pageshttp://dx.doi.org/10.1155/2016/8952520 2. Rubino et al., Rapid Communications in Mass Spectrometry, 22(23), pp. 3935\u20133948. doi: 10.1002/rcm.3810

Regulatory Toxicology and Pharmacology, Aug 1, 2020

Pesticides represent an economical, labor-saving, and efficient tool for pest management, but the... more Pesticides represent an economical, labor-saving, and efficient tool for pest management, but their intrinsic toxic properties may endanger workers and the general population. Risk assessment is necessary, and biological monitoring represents a potentially valuable tool. Several international agencies propose biological exposure indices (BEI), especially for substances which are commonly absorbed through the skin. Biological monitoring for pesticide exposure and risk assessment seems a natural choice, but biological exposure limits (BEL) for pesticides are lacking. This study aims at establishing equivalent biological exposure limits (EBEL) for pesticides using real-life field data and the Acceptable Operator Exposure Level (AOEL) of mancozeb as the reference. This study included a group of 16 vineyard pesticide applicators from Northern Italy, a subgroup of a more extensive study of 28 applicators. Their exposure was estimated using "patch" and "hand-wash" methodologies, together with biological monitoring of free ethylene-bis-thiourea (ETU) excretion in 24-hour pre-and post-exposure urine samples. Modeling was done using univariate linear regression with ETU excretion as the dependent variable and the estimated absorbed dose as the independent variable. The median skin deposition of mancozeb in our study population was 125 µg, leading to a median absorbed dose of 0.9 µg/kg. The median post-exposure ETU excretion was 3.7 µg. The modeled EBEL for mancozeb was 148 µg of free ETU or 697 µg of total ETU, accounting for around 75% of the maximum theoretical excretion based on a mass balance model. Although preliminary and based on a small population of low-exposed workers, our results demonstrate a procedure to develop strongly needed biological exposure limits for pesticides. .

Abstracts, Mar 1, 2023

Occupational epidemiology has traditionally relied on large records-based studies of workers to e... more Occupational epidemiology has traditionally relied on large records-based studies of workers to examine the association between an industrial exposure and a chronic disease outcome. These studies are highly informative for worker protection and environmental standards. However, establishing and maintaining large occupational cohorts has always been time and resource intensive. These studies are becoming increasingly challenging to initiate. Furthermore, the nature of work is changing; industrial cohort data cannot always address complex exposures and working conditions or the social and organizational factors that affect contemporary workers. In this symposium, I will present current efforts to develop a cohort of workers exposed to carbon nanomaterials, and discuss the challenges and opportunities associated with this ongoing effort. I will also discuss emerging themes in occupational epidemiology which can be used to address contemporary challenges in research development, including methodological developments, legal epidemiology, open-source software, occupational health equity, and social and structural determinants of occupational health.

... Abstract. Indicatori di esposizione a idrocarburi aromatici policiclici : il paradosso pirene... more ... Abstract. Indicatori di esposizione a idrocarburi aromatici policiclici : il paradosso pirene-idrossipirene / FM Rubino, C. Verduci, M. Buratti, S. Fustinoni, L. Neri, G. Brambilla, A. Colombi. - In: Giornale Italiano di Medicina del Lavoro ed Ergonomia. - ISSN 1592-7830. ...

Elsevier eBooks, 2021

Pesticides are among the most employed synthetic chemicals, and for which there, is the largest d... more Pesticides are among the most employed synthetic chemicals, and for which there, is the largest database of chemical and toxicological information that allows establishing health-based exposure limits. Protection of pesticide applicators and of the general population from over-exposure is both an ethical necessity and a contribution to planetary health. So far the exposure occupational and population exposure limits could not be efficiently used for health protection, due to the limitations of current techniques to be employed on a wide scale. Biological monitoring of exposure is a viable technique for occupational protection and for the assessment of chemical risk of the population Pesticides are intrinsically toxic, and their manipulated quantity and wide use requires adequate protection to reduce the potential impact on applicators\u2019 health. Before marketing the exposure-related health risk of each active substance is assessed through models, to demonstrate that at least in one application scenario its use does not cause an unacceptable health risk to the exposed workers. This reasonably safe scenario identifies the modalities for an effective and safe use that are coded as \u201cGood Agricultural Practices\u201d (GAP) and summarized in the label's instructions. However, real-life work may be conducted out of the frame of GAP, with a consequent health risk for the exposed workers, which must be assessed and managed. In real-life working conditions, risk assessment is seldom, if at all, performed since the task is linked to economic cost, the limited availability of trained personnel and logistics necessary to reach small, family-based enterprises, which are poorly covered by occupational health services. Additional difficulties are represented by the variability of working patterns, climatic conditions, and the frequent use of mixtures of pesticides. The main tools currently available for the exposure and risk assessment \u201cin the field,\u201d namely, biological and environmental monitoring, show important limits for their use in agriculture. In particular, assessment of dermal contamination involves very complicated and expensive procedures that cannot be carried out on a routine basis. Biological monitoring faces strong limitations, including lack of fully validated biomarkers and biological exposure limits. The exposure estimate and risk estimate done for a specific worker at a specific time during one of his workdays using any of the aforementioned methods represent only one point on a map of exposures and risk for workers applying pesticides. Instead of a point, we are interested in the whole map of exposure and risk. To make risk assessment available to all workers in various working conditions, there is a need of simple, user-friendly, and reliable approaches to estimate the levels of exposure (and of related occupational risk) experienced by the workers during typical, rather than actual, activities. We refer to these typical conditions as exposure and risk profiles. This chapter underlines the variables influencing exposure and exposure assessment in real-life work scenarios, reviews the drawback and difficulties of performing exposure assessment using environmental and/or biological monitoring, introduces the concept of exposure scenarios and risk profiles, reviews the current state of risk profile development and their main characteristics, and presents a way forward to integrate the exposure assessment through the exposure score (or index) and the toxicity score to create a risk assessment scheme or a map of exposure and risk in typical exposure scenarios. when measurements can be compared with suitable limit values. This possibility is currently employed only for few metals and organics of industrial interest, but only insufficiently for pesticides. We propose a paradigm to assess exposure and exposure-related risk of pesticides based on the measurement of their metabolites excreted in urine and on the elaboration of results from field exposure studies. Regression models allow forecasting the pesticide excretion associated to a systemic dose of pesticide corresponding to its acceptable occupational exposure limit (AOEL) for agricultural pesticide applicators. For this indicator and its corresponding limit value, we propose the name of equivalent biological exposure limit (EBEL). To demonstrate the proposed procedure and to highlight its utility and current limitations, we report data and elaborations from a previously published study that allows establishing a provisional value for an herbicide. This procedure can be adapted to fill the identified sources of uncertainty and to derive a limit value tha

... industriale (2004). NERI, LUCA,; RUBINO, FEDERICO MARIA,; BRAMBILLA, GABRI,; COLOMBI, ANTONIO... more ... industriale (2004). NERI, LUCA,; RUBINO, FEDERICO MARIA,; BRAMBILLA, GABRI,; COLOMBI, ANTONIO LUCIANO PAOLO,; A. Colombi, FM Rubino, M. Buratti, S. Fustinoni, C. Verduci, L. Neri, G. Brambilla. Abstract. Il metabonoma ...

Journal of Chromatography B: Biomedical Sciences and Applications, Nov 1, 2001

Methotrexate (MTX) is the prototype folate antagonist cytotoxic drug, employed in the therapy of ... more Methotrexate (MTX) is the prototype folate antagonist cytotoxic drug, employed in the therapy of solid tumors and leukaemias, and recently also as an immunosuppressive agent in organ transplantation, in the treatment of some autoimmune diseases and in the therapy of severe asthma. MTX is one of the very few antineoplastic drugs the therapeutic concentration monitoring of which is currently employed in clinical practice and can be routinely measured in biological samples by a number of different analytical techniques, among which are immunoenzymatic and chromatographic methods. Each technique has of course its own advantages in terms of sensitivity, specificity, speed, cost and level of expertise required. Along with therapeutic drug concentration monitoring and clinical pharmacology, fundamental research into the mechanism of action of antifolate drugs is still a field which requires the measurement of MTX, of its new analogues and of their metabolites in biological samples. This review summarizes the instrumental conditions and the performance of several published chromatographic methods employed to measure MTX, its metabolites and some analogues in clinical and biological research. More than 70 papers describing chromatographic assays for MTX and its metabolites have been published in the literature between 1975 and 2000. A wide array of experimental conditions for sample preparation, analyte separation and detection have been employed. According to their chemical properties, MTX, its metabolites and analogue drugs present in several biological samples (plasma, serum, saliva, urine, cerebrospinal fluid, tissue specimens) can be extracted, separated and detected under a variety of chromatographic conditions, i.e. on different stationary phases, under a wide choice of mobile phase conditions (acidic or neutral, employing ion-pair or micellar chromatography), followed by several detection techniques (UV-Vis spectrophotometry, pre-or post-column oxidation and fluorimetry, electrochemistry, mass spectrometry). Optimized methods allow simultaneous measurement within a few minutes of the plasma levels of MTX and its main metabolites at concentrations in the low-nM range. One special field which needs sensitive, fast and inexpensive methods for the detection and measurement of MTX is the monitoring of contamination in workplace environments, such as pharmaceutical industries and oncological hospital pharmacies, and in sewage waters. The measurement of the intracellular g-oligo-glutamate metabolites of biological folates, of MTX and of some analogue drugs is of great importance in basic pharmacological research. The existence of empirical quantitative relationships between the retention of individual oligomers under different chromatographic conditions and the number of added glutamic acid units allows identification of the metabolites even when authentic standards are not available.

Journal of Food Composition and Analysis, Sep 1, 2019

Tree nuts are healthy foods rich in bioactive phytochemicals. Their regular, moderate consumption... more Tree nuts are healthy foods rich in bioactive phytochemicals. Their regular, moderate consumption has been associated with a reduced risk of chronic-degenerative diseases, in the context of a healthy diet and lifestyle. This study aimed to investigate the phytochemical profile of almonds and pistachios, in order to add new elements in the complex scenario of nut chemistry. A LC-MS/MS method was developed to quantify melatonin and ceramides in almonds and pistachios. In general, pistachios were richer in melatonin (2609±3096 vs 1222±500 pg/g) and total ceramides (302±77 vs 165±21 pmol/g) than almonds. Among total ceramides fatty acyl homologs, the most represented was the C16:0 species, both for ceramides and dihydroceramides, and both in almonds (37-40%) and pistachios (51-74%). Overall, these results add a piece of information to elucidate the chemical composition of almonds and pistachios and provide a rationale for the nutraceutical potential of nuts in the Mediterranean diet.

The triple quadrupole mass spectrometer (TSQ) has boosted discovery and quantification of trace c... more The triple quadrupole mass spectrometer (TSQ) has boosted discovery and quantification of trace components in biological samples using class-selective scan modes. The use of suitable molecular precursors and of characteristic fragment ions allows identifying and measuring ceramides with micro-heterogeneous fatty acid or sphingosine. Collision energy in the TSQ is a key parameter to extract information from MS-MS experiments for compounds of which authentic standards are not available. We describe the theoretical basis and proof-of-principle application of a novel scan mode of a commercial TSQ by which the molecular precursor ions of all ceramides in a sample experiment the same Center-of-Mass Collision Energy (CoM-CE), irrespective of their different m/z values over a range of homologous or chemical analogs. At this value of CoM-CE, the different precursor ions have, for the same fragmentation channel, essentially the same fragment ion cross section, thus the same response factor for quantification. This scan mode can be applied to Precursor and Neutral Loss scans, and is performed by modulating the voltage drop of a scanning mass filter according to a specific linear function of precursor ion mass. Under this instrumental condition, all precursor ions collide under the same conditions and fragment to a common charged (in Precursor Ion scan) or neutral (in Neutral Loss scan) sub-structure, and response factors are thus intrinsically homogeneous, especially for first-generation fragments. As a proof-of-principle application in the sphingolipid field, isoenergetic fast scanning during LC separation of lipid extracts highlights the presence of unanticipated ceramides that can be quantified as equivalents of known analogs without resorting to the construction of individual calibration curves. The use of a scan mode coupled to the LC separation allows improving the amount of information on the sphingolipidome that can be extracted from complex samples, and discovering components with unusual or modified fatty acids. Among displayed proof-of principle examples are the sphingolipid fractions of organs and tissues of experimental animals, and of edible seeds and nuts

Giornale italiano di medicina del lavoro ed ergonomia, 2004

Il metil tert-butil etere (MTBE) \ue8 un additivo ossigenato delle benzine. Per valutare l\u2019e... more Il metil tert-butil etere (MTBE) \ue8 un additivo ossigenato delle benzine. Per valutare l\u2019esposizione a vapori di benzina e a traffico autoveicolare \ue8 stato condotto uno studio su 10 benzinai, 8 vigili urbani e 10 impiegati attraverso la determinazione di MTBE urinario. Le concentrazioni di MTBE urinario sono risultate simili in vigili e controlli e maggiori nei benzinai. Questo indicatore biologico sembra adatto a valutare l\u2019esposizione a vapori di benzina ma non a traffico autoveicolare

Introduction Ceramide (Cer) is one of the most important sphingolipids subclasses actively involv... more Introduction Ceramide (Cer) is one of the most important sphingolipids subclasses actively involved in apoptosis and cell death. Recently, it has been proposed as pharmaceutical target in pathologies related to its over productions and, in this optic, its specific involvement within the more complex sphingolipid pathways is needed. Liquid Chromatography coupled to mass spectrometry (LC-MS/MS) is the most important technique to study the Cer metabolism, and the unique able to provide an in-depth profile of all sphingolipid species. Here we present the optimized LC-MS/MS procedure to quantify Cer species in different biological matrices and in different biological models related to deregulation of Cer metabolism. Materials and methods Sphingolipids from different tissues and cells were fortified with internal standard N-lauroyl-D-erythro-sphingosine (0.2 nmol), and extracted with a customized procedure modified from the one described by Merrill et al. (1). The LC-MS/MS (Dionex Ultimate 3000 tandem AB Sciex 3200 Q Trap), operated in the positive electrospray ionization (ESI) mode. Identification and quantification of all targeted Cer was accomplished by multiple reactions monitoring (MRM) by following the transition from the [MH+] species to the common ion fragment 264.4 m/z. The Cer content was expressed as pmol normalized to total protein content (mg). The analysis run to run lasted 22 min at flow rate 0.3 mL/min. Results and Conclusions As low as 50 fmoles can be detected on-column and quantified with \ub115% precision and within \ub120% of target level; the method was linear up to 400 pmoles in a typical 150 microL extract. Examined biological samples (\u2c350), include mouse retinas and hearts, human Chordoma tumor biopsies, C38 and meningioma cell cultures. The method here presented resulted suitable for all the tested samples provided by in vivo and in vitro studies. In particular, the method was useful as a direct read-out for pharmacological treatments aimed to lower the Cer content in different pathological models related to the Cer overproduction