Angelo Faberi

Peptidomic approach, based on liquid chromatography/electrospray ionization tandem mass spectrometry, for detecting sheep's milk in goat's and cow's cheeses

A common fraud in the dairy field is the addition of sheeps milk to goats cheeses, because it has a very similar taste to goats milk, but is more available, and is commonly considered to have a better capacity to curdle. For similar reasons, and due to economic convenience, sheeps cheeses may also contain fraudulent cows milk. In order to detect this fraud, an EU official method may be used, but it is only a qualitative method (presence/absence of cows milk). A method able to quantify the presence of sheeps milk during cheese production in goats and cows cheeses was developed. The method is based on liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) analysis of peptides of a casein extract from the cheese. By a simple procedure, caseins are extracted from cheeses, solubilized, digested with plasmin, and subsequently analyzed by LC/ESI-MS/MS. A typical sheeps peptide produced by plasmin hydrolysis (m/z = 860) was accurately selected and analyzed to understand if, and by how much, a declared pure goats cheese contains sheeps milk. By analyzing the same peptide it is also possible to detect if, and by how much, a declared pure sheeps milk contains, or not, cows milk. The method was applied to several goats and cows cheese samples. Quantitation was performed with a calibration curve obtained by analyzing curd cheeses containing different percentages of sheeps milk. The method detection limit and method quantitation limit were evaluated. This method appears accurate and suitable for detecting up to 2% of sheeps milk in cheeses.

Multi-detection of preservatives in cheeses by liquid chromatography–tandem mass spectrometry

The incorrect use of preservatives in cheeses may compromise food safety and damage consumers. According to the law, more than one preservative may be contemporarily used in cheeses. So a method for their contemporary detection may be useful for both manufacturers and control agencies quality control. In this research a liquid chromatography-tandem mass spectrometric with electrospray ionization method for the multi-determination of seven preservatives (benzoic acid, citric acid, hexamethylenetetramine, lysozyme, natamycin, nisin and sorbic acid) in cheese was developed. The preservatives were contemporarily extracted from cheese by a single procedure, and analyzed by RP-LC/ESI-MS/MS (Ion Trap) in positive ionization mode, with single reaction monitoring (SRM) acquisition. Three sample types (hard, pasta filata and fresh cheese) were used for method evaluation. Recoveries were mostly higher than 90%; MDLs ranged from 0.02 to 0.26 mgkg(-1), and MQLs were included between 0.07 and 0.88 mgkg(-1). Due to matrix effect, quantitation was performed by referring to a matrix matched calibration curve, for each cheese typology. This method was also applied to commercial cheese samples, with good results. It appears fast, reliable and suitable for both screening and confirmation of the presence and quantitation of the preservatives in a single, multi-detection analysis.

Simultaneous quantitation of free and conjugated phytoestrogens in leguminosae by liquid chromatography-tandem mass spectrometry

Phytoestrogens are diphenolic compounds that are present in several edible plants and are particularly abundant in soybeans. Because of their estrogenical, antriestrogenical, anticarcinogenic and antioxidant activities in animal and humans, they became of great interest. Dietary factors are considered important in determination of risks, in fact, studies have revealed beneficial or protective effects of the consumption of vegetables, in particular soy and soybean products. So that in the present paper the simultaneous determination of eight isoflavones and coumestrol in vegetables is reported. The quantitative analysis has been made by means of LC separation combined with tandem mass spectrometry. In particular, a new simple and fast extraction methodology and a clean-up, based on cold aided de-fatting, is proposed. Method performance was evaluated by comparison with a reference procedure. The developed procedure was then used for a survey of phytoestrogens concentration in some selected vegetables.

Application of an innovative matrix solid-phase dispersion–solid-phase extraction–liquid chromatography–tandem mass spectrometry analytical methodology to the study of the metabolism of the estrogenic mycotoxin zearalenone in rainbow trout liver and muscular tissue

The metabolic fate of the estrogenic mycotoxin zearalenone in rainbow trout is presently unknown. In this study, the tissue concentration of zearalenone and its principal metabolites (α-zearalenol and β-zearalenol) was determined. A known amount of zearalenone was administered as a single bolus to ten fish, and the biological tissue concentration was determined at various times following administration. The analytes were extracted from liver and muscular tissue using an on-line matrix solid-phase dispersion–solid-phase extraction sample preparation protocol, and their concentration determined by HPLC–Turboionspray–tandem mass spectrometry. The results showed that zearalenone is mainly metabolized into α-zearalenol in both liver and muscular tissues. The maximum concentrations of each analyte found in liver were 76.1, 211.2 and 63.7 ng/g respectively for zearalenone, α-zearalenol and β-zearalenol, while in muscular tissue they were 10.7, 8.2 and 6.5 ng/g. These values were reached after 2 h in liver tissue and 12 h in muscular tissue. Moreover the data obtained showed that the elimination rate in liver is quite fast since 48 h after the exposure less than 7% of the maximum concentration found is still present. In muscular tissue, however, about one-third of the maximum concentration found is still present after 48 h.

The use of IRMS, (1)H NMR and chemical analysis to characterise Italian and imported Tunisian olive oils.

Isotope Ratio Mass Spectrometry (IRMS), (1)H Nuclear Magnetic Resonance ((1)H NMR), conventional chemical analysis and chemometric elaboration were used to assess quality and to define and confirm the geographical origin of 177 Italian PDO (Protected Denomination of Origin) olive oils and 86 samples imported from Tunisia. Italian olive oils were richer in squalene and unsaturated fatty acids, whereas Tunisian olive oils showed higher δ(18)O, δ(2)H, linoleic acid, saturated fatty acids β-sitosterol, sn-1 and 3 diglyceride values. Furthermore, all the Tunisian samples imported were of poor quality, with a K232 and/or acidity values above the limits established for extra virgin olive oils. By combining isotopic composition with (1)H NMR data using a multivariate statistical approach, a statistical model able to discriminate olive oil from Italy and those imported from Tunisia was obtained, with an optimal differentiation ability arriving at around 98%.

Fatty acid composition and δ13C of bulk and individual fatty acids as marker for authenticating Italian PDO/PGI extra virgin olive oils by means of isotopic ratio mass spectrometry†

European Regulation (EEC) 2568/91 has been setting the minimum requirements in order to allow labeling of oil as extra virgin. These general requirements, are based on physical-chemical and organoleptic parameters directly linked to the freshness and quality of the product. Isotope ratio mass spectrometry (IRMS) was demonstrated to be a useful tool for the discrimination of the origin of unknown samples, because the obtained data are practically independent of the cultivar employed and the production technique. In this work, the evaluation of the composition of fatty acid methyl esters (FAME) alongside with the determination of stable isotope ratio of C in bulk oils and in main FAME constituents have been investigated as a tool to improve geographical discrimination of Italian Protected Designation of Origin/Protected Geographical Indication (PDO/PGI) samples. For this purpose, authentic PDO/PGI extra virgin olive oils were sampled at oil mills and grouped into different sets according to their areas of provenience. The use of principal component analysis and partial least squares discriminant analysis multivariate analysis techniques demonstrated that discrimination of olive oil samples can be done using geographical and pedoclimatic parameters predominantly by using δ(13) C results of bulk and individual fatty acids. Results showed that δ(13) C values are a more reliable marker of origin with respect to fatty acid composition.

Determination of Maize and Grain Herbicides and their Transformation Products in Soil by Use of Soil Column Extraction then Liquid Chromatography with Tandem Mass Spectrometry

SummaryA multiresidue method has been developed for identification and quantitation of the herbicides most commonly used in cultivation of maize and grain, and of their transformation products, in soil samples. The analytes were isolated by soil column extraction (SCE) and the extracts were purified by use of a Carbograph-1 cartridge. Analysis was performed by liquid chromatography coupled with tandem mass spectrometric detection (LC-TISP-MS-MS) in negative-ion mode. To optimize the extraction conditions affecting the performance of SCE, e.g. the extracting solvent used, temperature, extracting volume, and solvent flow rate, were studied. To evaluate the matrix effect in SCE, recovery experiments were performed on soil samples, with different physical and chemical characteristics, fortified with the 100 ng g−1 of the target compounds. Recovery data were satisfactory and the method detection limits were between 3 and 100 ng g−1, depending on the compound.

Ethyl esters versus fermentative organoleptic defects in virgin olive oil.

The quality and genuineness of extra-virgin olive oils (EVOOs) were assessed following the methods and parameters of EU Commission Regulation N° 2568/91/EEC and subsequent modifications, which also set specific limits for fatty-acid ethyl esters (FAEEs). This study included a subset of EVOOs from among 399 samples analysed as part of a monitoring study for FAEEs in EVOOs. The subset was subjected to statistical evaluation to quantify the relationships between FAEE content and sensory defects associated with fermentation: fusty/muddy sediment, musty/humid/earthy, and winey/vinegary. The use of multiple regression analysis demonstrates that FAEE content can be inferred as a function of the intensity of organoleptic defects for samples with high alkyl esters content. The intensity of the rancid defect negatively influences the accuracy of this model, because of underestimation of the fermentation defects that are also present.

Pressurized-liquid extraction for determination of imidazolinone herbicides in soil

SummaryA rapid and simple method has been developed for determination of imidazolinone (IMI) residues in soil.Extraction of the analytes from the soil matrix was performed with a pressurized-liquid-extraction apparatus built in this laboratory. Four different types of soil sample (clay, clay loam, sandy clay loam, and silty loam) were fortified with target compounds at levels of 10 and 50 ng g−1 by a procedure which can mimic weathered soils. The samples were then dried and packed in a 25 cm×4.6 mm i. d. stainless steel column; this was placed inside a GC oven and extracted by passing an aqueous solution of KCl (0.1m, 20 mL) through the column at 90°C.Quantification of the analytes in the final extract (50-μL injection) was performed by reversed-phase liquid chromatography-mass spectrometry with a TurbolonSpray interface. Recoveries of the analytes were greater than 83% andRSD less than 7%. The method detection limit was in the 1–2.5ng g−1 range in analysis by time-scheduled selected-ion monitoring (SIM).

Italian Cheeses Discrimination by Means of δ 13 C and δ 15 N Isotopic Ratio Mass Spectrometry

Protection of the quality products and particularly Protected Designation of Origin (PDO)/Protected Geographical Indication (PGI) foods is a strategic issue in the EU economy, in terms of protection of market competition and safety. Having reliable tools for the assessment of key parameters useful for the identification of authenticity and/or frauds is therefore of main interest. In this work, the isotope ratios of stable elements variability of four PDO cheeses (Taleggio PDO, Asiago PDO, Pecorino Toscano PDO, and Provolone Valpadana PDO) were investigated with the aim to find discrimination among different kinds of cheeses. The specificity of isotope ratios of stable elements can be profitably used when sample characteristics, conditions, or degradation strongly suggest looking directly at the atoms rather than to the molecules. We analyzed five isotopic parameters: δ13C and δ15N on casein fraction and on cheese as a whole; δ13C on the fat fraction of the cheese. The dataset was composed by 118 cheese samples coming from five different Italian regions and collected over a 3-year period. Data elaboration showed that beyond interesting differences already observed on each individual cheese on the basis of some parameters taken into account (year, season, province, altitude), the characteristic isotopic ratios of each cheese are stable within a narrow range of δ‰. Univariate analysis showed that single parameters were not enough to provide a clear discrimination between each cheese, while principal component analysis (PCA) and partial least squares regression-discriminant analysis (PLS-DA) showed a good separation between cheese classes, particularly for the Pecorino Toscano cheese type. This data suggested a positive indication to the possibility of introducing in the production disciplinary of the concerned cheeses also a range of isotopic ratios of C and N as a further tool for the protection of this four types of PDO cheeses.