Monica Mattarozzi

Innovative Cavitand-Based Sol-Gel Coatings for the Environmental Monitoring of Benzene and Chlorobenzenes via Solid-Phase Microextraction

An innovative and very selective solid-phase microextraction coating synthesized by sol-gel technology was developed for the determination of environmental pollutants such as aromatic hydrocarbons at trace levels in air, water, and soil samples. The obtained fibers, composed of quinoxaline-bridged cavitand units, were characterized in terms of film thickness, morphology, thermal stability, and pH resistance. Fibers, characterized by an average thickness of 56 +/- 6 mum, exhibited an excellent thermal stability until 400 degrees C and a very good fiber-to-fiber and batch-to-batch repeatability with RSD lower than 6%. Finally, the capabilities of the developed coating for the selective sampling of aromatic hydrocarbons were proved, obtaining LOD values in the subnanogram per liter range. Extraction efficiency at least 2-fold higher than that obtained using commercial devices was proved for chlorobenzenes sampling in river water, obtaining extraction recoveries ranging from 87.4 +/- 2.6% to 94.7 +/- 1.9%. The selective desorption of benzene in the presence of high amounts of other airborne pollutants was also demonstrated.

Culturable endophytic bacteria enhance Ni translocation in the hyperaccumulator Noccaea caerulescens

In this work, both culture-dependent and independent approaches were used to identify and isolate endophytic bacteria from roots of the Ni hyperaccumulator Noccaea caerulescens. A total of 17 isolates were cultured from root samples, selected for tolerance to 6mM Ni and grouped by restriction analysis of 16S rDNA. Bacterial species cultivated from roots belonged to seven genera, Microbacterium, Arthrobacter, Agreia, Bacillus, Sthenotrophomonas, Kocuria and Variovorax. The culture-independent approach confirmed the presence of Microbacterium and Arthrobacter while only other five clones corresponding to different amplified ribosomal DNA restriction patterns were detected. Five selected highly Ni-resistant bacteria showing also plant growth promoting activities, were inoculated into seeds of N. caerulescens, and in vivo microscopic analysis showed rapid root colonisation. Inoculated plants showed increased shoot biomass, root length and root-to-shoot Ni translocation. Root colonisation was also evident, but not effective, in the non-hyperaccumulating Thlaspi perfoliatum. Seed inoculation with selected Ni-resistant endophytic bacteria may represent a powerful tool in phytotechnologies, although transferring it to biomass species still requires further studies and screening.

Experimental design for the optimization of the extraction conditions of polycyclic aromatic hydrocarbons in milk with a novel diethoxydiphenylsilane solid-phase microextraction fiber

An innovative solid-phase microextraction coating based on the use of diethoxydiphenylsilane synthesized by sol-gel technology was used for the determination of polycyclic aromatic hydrocarbons at trace levels in milk. The effects of time and temperature of extraction and acetone addition were investigated by experimental design. Regression models and desirability functions were applied to find the experimental conditions providing the highest global extraction response. The capabilities of the developed fiber were proved obtaining limit of quantitation values in the low microg/l range, enabling the direct analysis of complex matrices like milk and a complete desorption of high-boiling compounds without carryover effects.

Multiplex liquid chromatography-tandem mass spectrometry for the detection of wheat, oat, barley and rye prolamins towards the assessment of gluten-free product safety

Celiac patients should feel confident in the safety of foods labelled or expected to be gluten-free. In this context, a targeted proteomic approach based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) technique was proposed to assess the presence of celiotoxic cereals, namely wheat, oats, barley and rye, in raw and processed food products. To this aim, unique marker peptides were properly selected in order to distinguish between the different cereal types. A revised cocktail solution based on reducing and denaturing agents was exploited for prolamin extraction from raw and processed food; in addition, defatting with hexane was carried out for sample clean-up, allowing to largely reduce problems related to matrix effect. Method validation on fortified rice flour showed good analytical performance in terms of sensitivity (limits of detection in the 2-18 mg kg(-1) range). However, poor trueness was calculated for self-made incurred bread (between 3 and 30% depending on the peptide), probably due to baking processes, which reduce gluten extractability. Thus, it is evident that in the case of processed foods further insights into sample treatment efficiency and reference materials for protein calibration are required to obtain accurate gluten determination. Finally, the developed method was applied for the analysis of market food products, offering the possibility to discriminate among cereals, with good agreement with labelled ingredients for gluten-containing foodstuffs.

Rapid desorption electrospray ionization-high resolution mass spectrometry method for the analysis of melamine migration from melamine tableware

Migration of melamine into foods from melamine tableware has been object of recent Rapid Alert System for Food and Feed (RASFF) notifications. In this context, a rapid and sensitive desorption electrospray ionization-high resolution mass spectrometry (DESI-HRMS) method was developed and validated for the determination of melamine migration from plastic materials. The migration test was performed using acetic acid 3% (w/v) as food simulant. Evaluation of DESI parameters in terms of choice of support, motion profile, geometrical configuration and operating conditions coupled to the use of an orbitrap mass analyzer allowed to achieve significant improvements in terms of selectivity and accuracy obtaining detection and quantitation limits at low microgram per kilogram level. A LC-ESI-MS method was also developed for confirmatory purposes. Both methods were applied to 44 melamine tableware samples available on Italian market in order to assess their compliance with the law. Different concentration levels ranging from 0.00773±0.0006 to 3.0±0.1 mg/kg were found after the third exposure to the simulant in new and used tableware with two samples out of 44 being characterized by a melamine release higher than the legal limit, i.e. 2.5 mg/kg. A two tailed t-test allowed to assess the good agreement between the quantitative results obtained applying the DESI-MS method with those provided by LC-ESI-MS, thus proving reliability of DESI-HRMS as rapid technique for the study of melamine release from plastic materials.

Rapid shotgun proteomic liquid chromatography-electrospray ionization-tandem mass spectrometry-based method for the lupin ( Lupinus albus L.) multi-allergen determination in foods.

Allergy to lupin is a growing food safety problem because this legume, increasingly exploited in the food industry, is one of the allergens that, according to law, must be declared on the labels of food products in the European Union. In this context, a rapid targeted proteomic approach based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis was proposed and aimed to unequivocal confirmation and reliable determination of the major lupin allergens, i.e., conglutins, in pasta and biscuits. Detected concentrations were around 1 mg of lupin/kg of pasta and biscuits, proving the capabilities of the MS-based method in terms of the sensitive allergen screening method. Good precision was observed in terms of both intra- and interday repeatability, with relative standard deviation (RSD) lower than 23%. Recoveries from 95 ± 10 to 118 ± 12% and from 103 ± 1 to 110 ± 12% ranges were calculated for biscuits and pasta, respectively. Finally, the applicability of the devised method was investigated by analyzing market samples containing lupin and samples that may possibly contain traces of lupin deriving from cross-contamination between products and production lines.

An amperometric immunosensor for diagnosis of celiac disease based on covalent immobilization of open conformation tissue transglutaminase for determination of anti-tTG antibodies in human serum.

A new amperometric immunosensor based on the covalent immobilization of tissue transglutaminase enzyme in its open conformation (open-tTG) was developed and optimized for determination of anti-tissue transglutaminase antibodies (anti-tTG) in human serum. Experimental design allowed us to find the optimal conditions for quantification of both IgA and IgG isotypes of anti-tTG in order to assess suitability of the device for diagnostic purposes. The glassy carbon electrodic substrate was electrochemically functionalized with gold nanoparticles and subsequently derivatized with a self-assembled monolayer of 11-mercaptoundecanoic acid for the covalent anchoring of the enzyme. This step was performed under carefully controlled conditions in order to keep the open conformation of the tTG. The immunosensor showed good analytical performance with limit of detection levels (1.7 AU mL(-1) for IgA and 2.7 AU mL(-1) for IgG) below the diagnostic threshold value (3.0 AU mL(-1)) and inter-sensor reproducibility giving RSD lower than 10%. The developed sensor was validated in serum samples from pediatric patients for clinical applications, using two ELISA kits specific for the determination of anti-tTG IgA and IgG antibodies as reference methods; good recovery rates ranging from 74% to 117% were calculated.

Combined endophytic inoculants enhance nickel phytoextraction from serpentine soil in the hyperaccumulator Noccaea caerulescens.

This study assesses the effects of specific bacterial endophytes on the phytoextraction capacity of the Ni-hyperaccumulator Noccaea caerulescens, spontaneously growing in a serpentine soil environment. Five metal-tolerant endophytes had already been selected for their high Ni tolerance (6 mM) and plant growth promoting ability. Here we demonstrate that individual bacterial inoculation is ineffective in enhancing Ni translocation and growth of N. caerulescens in serpentine soil, except for specific strains Ncr-1 and Ncr-8, belonging to the Arthrobacter and Microbacterium genera, which showed the highest indole acetic acid production and 1-aminocyclopropane-1-carboxylic acid-deaminase activity. Ncr-1 and Ncr-8 co-inoculation was even more efficient in promoting plant growth, soil Ni removal, and translocation of Ni, together with that of Fe, Co, and Cu. Bacteria of both strains densely colonized the root surfaces and intercellular spaces of leaf epidermal tissue. These two bacterial strains also turned out to stimulate root length, shoot biomass, and Ni uptake in Arabidopsis thaliana grown in MS agar medium supplemented with Ni. It is concluded that adaptation of N. caerulescens in highly Ni-contaminated serpentine soil can be enhanced by an integrated community of bacterial endophytes rather than by single strains; of the former, Arthrobacter and Microbacterium may be useful candidates for future phytoremediation trials in multiple metal-contaminated sites, with possible extension to non-hyperaccumulator plants.

Liquid chromatography-full scan-high resolution mass spectrometry-based method towards the comprehensive analysis of migration of primary aromatic amines from food packaging.

European Union legislation has established that plastic food contact materials shall not release primary aromatic amines (PAAs), which are toxic compounds and suspected human carcinogens. As valid alternative to existing methods for PAA determination, which are based on spectrophotometric test or targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) approaches, in this study a LC-Orbitrap-full scan-high resolution mass spectrometry (HRMS) method was devised and validated for the determination of migration levels of 22 PAAs from food contact materials, thus exploiting the specificity of accurate mass measurement. Direct injection of the simulant (acetic acid 3%, w/v) into the LC-MS system after migration, without any pre-treatment step, makes the developed method of great value for rapid screening analysis of a large number of amines. A very fast and efficient separation (<11min) of PAAs was achieved. Detection limits in the 0.06-0.7μgkg(-1) range were calculated for 17 out of 22 of the investigated PAAs, however obtaining values within 5.3μgkg(-1) for the other 5 amines. Good dynamic linear ranges from two to four orders of magnitude (r(2)≥0.990) were obtained and satisfying results were achieved in terms of intra-day (RSDs<10%) and inter-day repeatability (RSDs<17%). Trueness values in the 70±1-131±5% range proved reliability of the developed method for PAAs quantification also at very low concentration levels. Finally, the method was successfully applied to a range of different real plastic multilayer food packaging materials, noticing in all cases levels below the established limits of detection.

An innovative method based on quick, easy, cheap, effective, rugged, and safe extraction coupled to desorption electrospray ionization-high resolution mass spectrometry for screening the presence of paralytic shellfish toxins in clams

The capabilities of desorption electrospray ionization-high resolution mass spectrometry (DESI-HRMS) were tested for screening the presence of some paralytic shellfish toxins in clams. A quick, easy, cheap, effective, rugged, and safe (QuEChERS) approach is proposed for sample clean-up. QuEChERS extraction was optimized by using a full factorial design followed by the multicriteria method of the desirability functions. Quantitation limits in the microgram per kilogram range proved reliability of the method for the detection of the investigated toxins in accordance to the rules laid down by European legislation. The optimized QuEChERS-DESI-HRMS based-method allowed for a rapid reliable screening of the investigated compounds at levels of interest, thus being useful for high-throughput analyses.