Anti-listerial activity of a polymeric film coated with hybrid coatings doped with Enterocin 416K1 for use as bioactive food packaging

In this study, Enterocin 416K1, a bacteriocin produced by Enterococcus casseliflavus IM 416K1, was entrapped in an organic-inorganic hybrid coating applied to a LDPE (low-density polyethylene) film for its potential use in the active food packaging field. The antibacterial activity of the coated film was evaluated against Listeria monocytogenes NCTC 10888 by qualitative modified agar diffusion assay, quantitative determination in listeria saline solution suspension and direct contact with artificially contaminated food samples (frankfurters and fresh cheeses) stored at room and refrigeration temperatures. All investigations demonstrated that enterocin-activated coatings have a good anti-listeria activity. Qualitative tests showed a clear zone of inhibition in the indicator lawn in contact with and around the coated film. During the quantitative antibacterial evaluation the L. monocytogenes viable counts decreased to 1.5 log units compared to the control. The inhibitory capability was confirmed also in food-contact assays. In all food samples packed with coated films we observed a significant decrease in L. monocytogenes viable counts in the first 24 h compared to the control. This difference was generally maintained up to the seventh day and then decreased, with the exception of the cheese samples stored at refrigeration temperature.     

A new competitive fluorescence assay for the detection of patulin toxin

Patulin is a toxic secondary metabolite of a number of fungal species belonging to the genera Penicillum and Aspergillus. It has been mainly isolated from apples and apple products contaminated with the common storage-rot fungus of apples, Penicillum expansum, but it has also been extracted from rotten fruits, moldy feeds, and stored cheese. Human exposure to patulin can lead to serious health problems, and according to a long-term investigation in rats, the World Health Organization has set a tolerable weekly intake of 7 ppb body weight. The content of patulin in foods has been restricted to 50 ppb in many countries. Conventional analytical detection methods involve chromatographic analyses, such as HPLC, GC, and, more recently, techniques such as LC/MS and GC/MS. However, extensive protocols of sample cleanup are required prior to the analysis, and to accomplish it, expensive analytical instrumentation is necessary. An immunochemical analytical method, based on highly specific antigen-antibody interactions, would be desirable, offering several advantages compared to conventional techniques, i.e., low cost per sample, high selectivity, high sensitivity, and high throughput. In this paper, the synthesis of two new derivatives of patulin is described, along with their conjugation to the bovine serum albumin for the production of polyclonal antibodies. Finally, a fluorescence competitive immunoassay was developed for the on-line detection of patulin.     

The reaction of hydrogen atoms with methionine residues: A model of reductive radical stress causing tandem protein-lipid damage

The occurrence of tandem damage, due to reductive radical stress involving proteins and lipids, is shown by using a biomimetic model. It is made of unsaturated lipid vesicle suspensions in phosphate buffer in the presence of methionine, either as a single amino acid or as part of a protein such as RNase A, which contains four methionine residues. The radical process starts with the formation of H(.) atoms by reaction of solvated electrons with dihydrogen phosphate anions, which selectively attack the thioether function of methionine. The modification of methionine to alpha-aminobutyric acid is accompanied by the formation of thiyl radicals, which in turn cause the isomerization of the cis fatty acid residues to the trans isomers. The relationship between methionine modification and lipid damage and some details of the reductive radical stress obtained by proteomic analysis of irradiated RNase A are presented.     

Metal content of southern Italy honey of different botanical origins and its correlation with polyphenol content and antioxidant activity

Seventy‐eight samples of southern Italy honey from five different floral origins (chestnut, eucalyptus, citrus, multifloral and sulla) were screened to quantify the polyphenol and metal contents, evaluate the antioxidant activity and determine the correlations between the parameters analysed. The average polyphenol content was 12.06 mg gallic acid equivalent per 100 g honey and 7.92 mg quercetin equivalent per 100 g honey, for total phenolic and flavonoid contents, respectively. The antioxidant activity ranged from 58.40% (eucalyptus honey) to 60.42% (chestnut honey) in the ABTS assay, from 152.65 μm Fe (II) (citrus honey) to 881.34 μm Fe (II) (chestnut honey) in the FRAP assay, and from 54.29% (citrus honey) to 78.73% (chestnut honey) in the DPPH assay. Fe and Zn were the most abundant among the tested metals, while Cd, Co and Mo were those less present. Chestnut honey presented the highest polyphenol content, antioxidant activity and metal content. The correlations between the analysed parameters were statistically significant (P < 0.05). The correlations between metal content and both total phenolic and antioxidant activities were particularly interesting, suggesting a relationship between metal and polyphenol contents in honey.     

Seismic reliability assessment of a two-story steel-concrete composite frame designed according to Eurocode 8

Recent earthquakes have shown that some buildings designed primarily for life safety according to the seismic provisions of current building codes could be subjected to severe damage to structural and non-structural elements and building contents. In order to overcome this drawback, the scientific community is developing more accurate methodologies to assess seismic performances of structures. In this framework, the present paper aims at the evaluation of mean annual frequency (MAF) of exceeding a limit state (limit value of interstory drift ranging from 0.002 to 0.05) following the probabilistic approach introduced by Pacific Earthquake Engineering Research Center adopting (PEER) the application procedure suggested by Jalayer and Cornell. Reference is made to a single two-story steel-concrete composite frame, designed according to Eurocode 8 and tested in real scale at the ELSA Laboratory of Ispra. The reason for investigating this frame is related to the availability of results both of pseudo-dynamic tests performed on 3D steel-concrete composite frame and of monotonic and cyclic tests performed on internal and external beam-to-column joint sub-assemblages belonging to the building. As a consequence, it has been possible to assess the seismic reliability of a 3D steel-concrete composite building, designed according to Eurocode 8, taking into account both randomness and uncertainty.     

Ionic Liquids, 3. Synthesis and Utilisation of Protic Imidazolium Salts in Homogeneous Catalysis

Protonation of 1‐alkylimidazoles provides halogen‐free salts which act as ionic liquids and proton reservoir in proton‐ and metal‐assisted catalytic processes like dimerisation of methyl acrylate and ring closing metathesis, and lead to significant improvements both in activity and selectivity.     

Investigating the Short Peptidome Profile of Italian Dry-Cured Ham at Different Processing Times by High-Resolution Mass Spectrometry and Chemometrics

Short peptides have been spiking interest owing to their significant contribution to the taste and functional properties of dry-cured ham. In this study, a suspect screening approach based on high-resolution mass spectrometry was employed for the comprehensive characterization of the short endogenous peptidome in dry-cured ham samples at different processing stages (14, 22, and 34 months). After careful manual spectra interpretation, a chemometric approach based on principal component analysis was employed for highlighting the differences between the three sets of samples. A total of 236 short peptide sequences was tentatively identified, including 173 natural short peptides and 63 sequences containing non-proteinogenic amino acids, the highest number ever reported for endogenous sequences in dry-cured ham. Samples in the latest processing stages presented a generally higher abundance of dipeptides, indicating residual proteolytic activity. Moreover, the several annotated modified short peptides, mainly pyroglutamination and lactoyl conjugation, allowed hypothesizing several reactions occurring over time. For the first time, several lactoyl-dipeptides were tentatively identified in dry-cured ham samples with maximum concentration in the late processing stage samples. The presented results significantly contribute to the understanding of the reaction involving short peptides that affect the sensory and functional properties of dry-cured ham.     

Bioactivated collagen-based scaffolds embedding protein-releasing biodegradable microspheres: tuning of protein release kinetics

In tissue engineering, the recapitulation of natural sequences of signaling molecules, such as growth factors, as occurring in the native extracellular matrix (ECM), is fundamental to support the stepwise process of tissue regeneration. Among the manifold of tissue engineering strategies, a promising one is based on the creation of the chrono-programmed presentation of different signaling proteins. This approach is based upon the integration of biodegradable microspheres, loaded with suitable protein molecules, within scaffolds made of collagen and, in case, hyaluronic acid, which are two of the fundamental ECM constituents. However, for the design of bioactivated gel-like scaffolds the determination of release kinetics must be performed directly within the tissue engineering template. In this work, biodegradable poly(lactic-co-glycolic)acid (PLGA) microspheres were produced by the multiple emulsion-solvent evaporation technique and loaded with rhodamine-labelled bovine serum albumin (BSA-Rhod), a fluorescent model protein. The microdevices were dispersed in collagen gels and collagen-hyaluronic acid (HA) semi-interpenetrating networks (semi-IPNs). BSA-Rhod release kinetics were studied directly on single microspheres through confocal laser scanning microscopy (CLSM). To thoroughly investigate the mechanisms governing protein release from PLGA microspheres in gels, BSA-Rhod diffusion in gels was determined by fluorescence correlation spectroscopy (FCS), and water transport through the microsphere bulk was determined by dynamic vapor sorption (DVS). Moreover, the decrease of PLGA molecular weight and glass transition temperature (T_g) were determined by gel permeation chromatography (GPC) and differential scanning calorimetry (DSC), respectively. Results indicate that protein release kinetics and delivery onset strongly depend on the complex interplay between protein transport through the PLGA matrix and in the collagen-based release media, and water sequestration within the scaffolds, related to the scaffold hydrophilicity, which is dictated by HA content. The proper manipulation of all these features may thus allow the obtainment of a fine control over protein sequential delivery and release kinetics within tissue-engineering scaffolds.