Vita Di Stefano

Applications of liquid chromatography–mass spectrometry for food analysis

HPLC-MS applications in the agrifood sector are among the fastest developing fields in science and industry. The present tutorial mini-review briefly describes this analytical methodology: HPLC, UHPLC, nano-HPLC on one hand, mass spectrometry (MS) and tandem mass spectrometry (MS/MS) on the other hand. Analytical results are grouped together based on the type of chemicals analyzed (lipids, carbohydrates, glycoproteins, vitamins, flavonoids, mycotoxins, pesticides, allergens and food additives). Results are also shown for various types of food (ham, cheese, milk, cereals, olive oil and wines). Although it is not an exhaustive list, it illustrates the main current directions of applications. Finally, one of the most important features, the characterization of food quality (including problems of authentication and adulteration) is discussed, together with a future outlook on future directions.

Mycotoxin contamination of animal feedingstuff: detoxification by gamma-irradiation and reduction of aflatoxins and ochratoxin A concentrations

Mycotoxins are fungal secondary metabolites identified in many agricultural products screened for toxigenic moulds. They have been reported to be carcinogenic, teratogenic, tremorogenic, haemorrhagic and dermatitic to a wide range of organisms. With the increasing stringent regulations for mycotoxins imposed by importing countries such as those of the European Union, many cereals that are not safe for human consumption are used in formulations intended for animal feed. Gamma-rays are reported in the scientific literature to destroy ochratoxin A and aflatoxin in food crops and feed. The present study provides preliminary data for establishing the effect of dose of gamma-irradiation, ranging from 0 to 15 kGy, on aflatoxins and ochratoxin A reduction in commercial animal feed. The mycotoxin levels were determined by means of immunoaffinity clean-up (IAC) and HPLC with fluorescence detection (HPLC-FLD). The maximum reductions found at 15 kGy were 23.9%, 18.2%, 11.0%, 21.1% and 13.6% for ochratoxin A, aflatoxin B1, aflatoxin B2, aflatoxin G1 and aflatoxin G2, respectively. Results showed that the gamma-rays even at 15 kGy were not effective in the complete destruction of ochratoxin A and aflatoxins in the tested feed.

Natural co-occurrence of ochratoxin A, ochratoxin B and aflatoxins in Sicilian red wines

The natural occurrence of ochratoxin A, ochratoxin B, aflatoxin B1, aflatoxin B2, aflatoxin G1 and aflatoxin G2 (OTA, OTB, AFB1, AFB2, AFG1, AFG2) in red wines was investigated by HPLC/FLD after immunoaffinity column clean-up in 57 market samples produced in Sicily (Italy). The results showed a very low incidence of these mycotoxins in analysed samples, confirming the high degree of quality and safety of Sicilian red wines. The results indicated 71.9% and 64.9% positive samples for OTA and OTB respectively, with an average level of 0.13 μg l–1, well below the European maximum permitted levels (MLs). The aflatoxin most frequently detected in the samples was AFG1, present in 57.9% of samples, while the other aflatoxins were rarely present. Recovery experiments were carried out on eight mycotoxin-free red wines spiked with OTA, OTB, AFB1, AFB2, AFG1 and AFG2 at two different levels. The limits of detection (LODs) in wines were 0.02 µg l–1 for OTA, 0.04 µg l–1 for OTB, 0.03 µg l–1 for AFG1, AFG2 and AFB2, and 0.05 µg l–1 for AFB1. A good correlation was found, with good performances in term of precision for the method.

Essential oil components of orange peels and antimicrobial activity

Abstract In this study, the orange peel of 12 cultivars of Citrus sinensis from central-eastern Sicily was employed to obtain essential oils and extracts. The ones were extracted through steam distillation, the others through extraction in hexane. Chemical constituents were evaluated in terms of qualitative and quantitative analyses by gas chromatography/mass spectrometry. Fifty-four components were identified in the steam essential oils and 44 in the extracts. In all the cultivars, the main component is d-limonene (73.9–97%); discrete percentages of linalool, geraniol and nerol were also found. Cluster analysis based on essential oils composition showed a certain degree of affinity between cultivars of the same type. The antimicrobial activity was investigated against three micro-organisms (Staphylococcus aureus, Listeria monocytogens and Pseudomonas aeruginosa). ‘Sanguinello’ and ‘Solarino Moro’ essential oils are significantly active against L. monocytogenes, while ‘Valencia’ hexanic extract against all the tested micro-organisms.

Triacylglycerols in edible oils: Determination, characterization, quantitation, chemometric approach and evaluation of adulterations

Vegetable oils are a dietary source of lipids that constitute an essential component of a healthy diet. The commonly used vegetable oils differ significantly for their triacylglycerol (TAG) profile. TAGs represent the principal components of oils and may contain different fatty acids (FA) esterified with glycerol leading to several positional isomers. To differentiate individual TAGs species in edible oils, advanced analysis systems and innovative methods are therefore required. TAGs can be considered as good fingerprints for quality control and many studies have been performed to develop rapid and low cost analytical methods to determinate the authenticity, origin and eventually evidence frauds or adulterations. The present manuscript provides a general overview on the most common vegetable oils TAGs compositions and on the related analytical methodologies recently used. Finally, the chemometric applications developed to assess the authenticity, quality and botanical origin of various edible oils are discussed.

Antimicrobial and antiproliferative activity of Athamanta sicula L. (Apiaceae)

Background: Athamanta sicula L., a member of Apiaceae, is an annual perennial herb and it is known in Sicilian popular medicine with the name of “spaccapietre” (rock splitters), because fresh roots infusions are indicated as diuretic and used in the treatment of diseases of the urinary tract, and to dissolve kidney stones. Materials and Methods: Acetone extracts of leaves, flowers, and stems of A. sicula L. were investigated in vitro for antibacterial and cytotoxic activities. Antimicrobial activity was carried out against bacterial and fungal strains and antiproliferative activity against a group of human cancer cell lines (K-562, NCI-H460, and MCF-7). Results: All acetone extracts, apiol and myristicin, resulted inactive as antimicrobial agents at the maximum tested concentration of 200 μg/mL, but they induced significant antiproliferative activity on the tested cancer cell lines. Conclusions: Our study show that both apiol and myristicin could be tested as novel treatment in cancer chemotherapy.

Quantitative evaluation of the phenolic profile in fruits of six avocado (Persea americana) cultivars by ultra-high-performance liquid chromatography-heated electrospray-mass spectrometry

ABSTRACT The phenolic profiles of six varieties of avocado (Persea americana) grown in Sicily were investigated. The ultra-high-performance liquid chromatography-heated electrospray-mass spectrometry method was developed to determine qualitative and quantitative changes in fruits at two different ripening stage. Nineteen individual phenolic compounds were detected in avocado pulp extracts. Gallic acid, sinapinic acid, vanillin, p-coumaric acid, and gentisic acid were present only in ripe fruits. On the contrary, epicatechin decreased with fruit ripening, whereas protocatechuic acid, 4-hydroxybenzoic acid, chlorogenic acid, and benzoic acid were relatively stable or exhibited erratic changes with fruit ripening. The different avocado cultivars exhibited different phenolic profiles and total quantities and it was found that, among the tested cultivars, “Orotawa” avocados may provide the highest nutritional and health contribution to human diet. The qualitative and quantitative differences among cultivars and maturation stages by multivariate analysis allowed for the individuation of a set of phenolic compounds that have a great potential in the discrimination and identification of different genetic groups.

Fast UPLC/PDA determination of squalene in Sicilian P.D.O. pistachio from Bronte: optimization of oil extraction method and analytical characterization

A fast reversed-phase UPLC method was developed for squalene determination in Sicilian pistachio samples that entry in the European register of the products with P.D.O. In the present study the SPE procedure was optimized for the squalene extraction prior to the UPLC/PDA analysis. The precision of the full analytical procedure was satisfactory and the mean recoveries were 92.8±0.3% and 96.6±0.1% for 25 and 50mgL-1 level of addition, respectively. Selected chromatographic conditions allowed a very fast squalene determination; in fact it was well separated in ∼0.54min with good resolution. Squalene was detected in all the pistachio samples analyzed and the levels ranged from 55.45-226.34mgkg-1. Comparing our results with those of other studies it emerges that squalene contents in P.D.O. Sicilian pistachio samples, generally, were higher than those measured for other samples of different geographic origins.

Electrospray ion mobility mass spectrometry of positively and negatively charged (1R,2S)-dodecyl(2-hydroxy-1-methyl-2-phenylethyl)dimethylammonium bromide aggregates.

RATIONALE Self-assembling processes of surfactants in the gas phase constitute a developing research field of interest since they allow information to be gained on the peculiar structural organization of these aggregates, on their ability to incorporate from small molecules up to proteins and on their possible use as carriers of drugs in the gas phase or as cleaning agents and exotic reaction media. METHODS The mass spectra of charged aggregates of the chiral surfactant (1R,2S)-dodecyl(2-hydroxy-1-methyl-2-phenylethyl)dimethylammonium bromide (DMEB) in the gas phase have been recorded using a Synapt G2-Si mass spectrometer in the positive and negative ion mode. For comparison purposes, the mass spectra of sodium bis(2-ethylhexyl)sulfosuccinate and sodium octane sulfonate aggregates have also been recorded under the same experimental conditions. The collisional cross sections of positively and negatively charged DMEB aggregates were obtained through an appropriate calibration of the measured drift times. RESULTS For all the surfactants investigated, it has been found that there is a lowest and a highest limit of the aggregation number at each charge state: no aggregates are found outside this range. Moreover, the occurrence at each aggregation number and extra charge of a unique value of drift time points toward aggregates whose conformations do not show discernible shape change in the experiment time scale. The analysis of the collisional cross sections emphasizes that the DMEB aggregates are nearly spherical clusters somewhat affected by the charge state and constituted by interlaced polar and apolar domains. CONCLUSIONS The analysis of all the experimental findings indicates that in the gas phase DMEB forms supramolecular aggregates characterized by an internal organization whose stability is triggered by the charge state. The comparison of the behavior of DMEB aggregates with that of sodium bis(2-ethylhexyl)sulfosuccinate and sodium octane sulfonate aggregates allows us to highlight the effects on the aggregate organization in gas phase due to nature of the head group and alkyl chain steric hindrance.

Essential Oil of Leaves and Fruits of Athamanta sicula L. (Apiaceae)

Abstract The essential oils obtained separately from the leaves and the fruits of Athamanta sicula L. collected in Sicily were studied by capillary GC/MS. The main component in the leaf oil was myristicin (97.6%), while β-pinene (2.8%), β-phellandrene (5.8%), δ-3-carene (22.6%), terpinolene (20.5%) and myristicin (41.3%) were the main components of the fruit oil.