Gabriella Pasini

IgE binding to soluble and insoluble wheat flour proteins in atopic and non-atopic patients suffering from gastrointestinal symptoms after wheat ingestion

Background The involvement of IgE‐mediated hypersensitivity reactions in the genesis of gastrointestinal symptoms after ingestion of foods containing wheat has been rarely reported.

Enhanced microbial cell lysis by the use of lysozyme immobilized on different carriers

Abstract Lysozyme was immobilized on chitosan and silica gel by adsorption, on cross-linked polystyrene divinylbenzene matrix (Deacidite KMP) by ionic binding and on non-porous glass beads by covalent attachment to improve the lytic activity towards complex substrates such as Micrococcus luteus . The lysozyme immobilization yields decreased on decreasing the amount of enzyme loaded, whereas the activation yields (defined as percentage ratio of immobilized active enzyme to immobilized enzyme) increased. Chitosan, silica gel, Deacidite KMP and non-porous glass bead preparations showed activation yields of 10%, 13·20%, 26·67% and 12·77%, respectively.

IgE-mediated allergy to corn: a 50 kDa protein, belonging to the Reduced Soluble Proteins, is a major allergen

Background: Although corn is often cited as an allergenic food, very few studies have been devoted to the identification of corn allergens and corn allergy has been rarely confirmed by double‐blind, placebo‐controlled food challenge (DBPCFC). Recently, Pastorello et al. (1) identified some salt‐soluble IgE‐binding proteins of corn flour as potential allergens. One of these, corresponding to corn Lipid Transfer Protein (LTP), appeared to be the major one. The aim of this study was to verify the clinical significance of the skin prick test (SPT) and CAP‐FEIA CAP‐System IgE fluozoenzyme immunosorbent assay (Pharmacia Diagnostic, Uppsala, Sweden) positivities to corn and to identify the presence of IgE‐binding proteins in the corn flour salt‐insoluble protein fractions (comprising up to 96% of the total protein) using sera of patients with DBPCFC‐documented food allergy to corn. In addition the effect of cooking and proteolytic digestion on the corn allergens was investigated.

Immunochemical and mass spectrometry detection of residual proteins in gluten fined red wine.

Recently, wheat gluten has been proposed as technological adjuvant in order to clarify wines. However, the possibility that residual gluten proteins remain in treated wines cannot be excluded, representing a hazard for wheat allergic or celiac disease patients. In this work, commercial wheat glutens, in both partially hydrolyzed (GBS-P51) and nonhydrolyzed (Gluvital 21000) forms, were used as fining agents in red wine at different concentrations. Beside immunoenzymatic analyses using anti-gliadin, anti-prolamin antibodies and pooled sera of wheat allergic patients, a method based on liquid chromatography coupled to mass spectrometry has been proposed to detect residues of gluten proteins. Residual gluten proteins were detected by anti-prolamin antibodies, anti-gliadin antibodies and sera-IgE only in the wine treated with GBS-P51 at concentration 50, 150, and 300 g/hL, respectively, whereas no residual proteins were detected by these systems in the wine treated with Gluvital 21000. In contrast liquid chromatography-mass spectrometry analyses allowed the detection of proteins in red wines fined down to 1 g/hL of Gluvital 21000 and GBS-P51. Our results indicate that MS methods are superior to immunochemical methods in detecting gluten proteins in wines and that adverse reactions against gluten treated wines cannot be excluded.

Production of a bacteriocin active on lactate-fermenting clostridia by Lactococcus lactis subsp. lactis immobilized in coated alginate beads

We report the isolation and immobilization of a nisinogenic strain (NZ1) of Lactococcus lactis subsp. lactis , active on gas-forming lactate-fermenting clostridia responsible for late blowing of Asiago and Montasio cheeses. The bacteriocin (nisin) produced by strain NZ1 is pronase-sensitive and is released in culture media during the growth phase. Using the sensitive indicator strain Lactobacillus delbrueckii subsp. bulgaricus NCDO 1489, a rapid microtitre plate based assay was developed for quantitative determination of the bacteriocin produced by NZ1 cells, either free or immobilized in gel beads. Scanning electron microscopy of cells immobilized in calcium alginate coated beads and viable counts of the surrounding medium showed that no cell leakage occurred during a 24 h assay. The bacteriocin released from immobilized cells reached, after 5 and 24 h, concentrations comparable to that of the free cell system after 3–4 h incubation in culture media.

Immobilized Laccase for Must and Wine Processinga

Whereas the use of enzyme preparations in the food industry is a wellestablished, rapidly expanding technology, their utilization in enology is less frequent. The “classic” wine industry is in fact still largely based on traditional methods. Moreover, musts and wines represent a hostile environment for purified enzymes, i.e., low pH, high ethanol concentration, presence of sulfur dioxide and tannins. White and ‘‘rosk” wines are known to be more sensitive than red wines to browning and to the development of madeirized flavor, which are mainly due to the presence of polyphenols. Aiming at the replacement of traditional stabilization treatments (i-e., sulfur dioxide, fining agents) with mild technologies to prevent madeirization, unconventional systems like the use of oxidative enzymes (chiefly laccase) have been proposed.’,* However, the residual enzyme activity in wine during storage or, in the case of musts, its inactivation by heat treatment could lead to negative effects, i.e., color browning. Therefore, the use of immobilized oxidative enzymes that can be easily recovered from the reaction mixture and reused could be exploited. Studies have been reportedly carried out on fungal laccase immobilized by covalent attachment to CNBr-activated Sepharose 4B and by adsorption to Concanavalin A-Sepharose? covalently bound to a diatomaceous silica product (R-650 Celite): immobilized on porous glass beads5s6 to detoxify the environment polluted with aromatic and phenolic compounds of agricultural or industrial origin. Taking into account our previous trials on the approach to the use of immobilized oxidative enzymes in white wine stabilization,7vu in the present work we report the results on a commercial laccase (p-diphenol: oxygen oxidoreductase; EC 1.10.3.2) immobilized by adsorption on silica gel carrier followed by treatment with glutaraldehyde and by covalent binding on polymer carrier VA-Hydroxy Biosinth, and CNBractivated Sepharose 4B, respectively. In addition, the possibility of an enzymatic removal of phenolics was tested in model solution, in must, and in wine.

IgE binding to almond proteins in two CAP-FEIA-negative patients with allergic symptoms to almond as compared to three CAP-FEIA-false-positive subjects.

Background: Allergy to almonds has been frequently reported, but data on the identification of the almond allergens, as well as on the reliability of the methods for in vitro detection of specific IgE for these allergens, are scant. This study aimed to identify the almond allergens and to evaluate the reliability of the CAP‐FEIA as the standard system for detection of almond‐specific IgE with clinical significance.

The proteins of the grape (Vitis vinifera L.) seed endosperm: Fractionation and identification of the major components

In the present study, grape (Vitis vinifera L.) seed endosperm proteins were characterized after sequential fractionation, according to a modified Osborne procedure. The salt-soluble fraction (albumins and globulins) comprised the majority (58.4%) of the total extracted protein. The protein fractions analysed by SDS-PAGE showed similar bands, indicating different solubility of the same protein components. SDS-PAGE in non-reducing and reducing conditions revealed the polypeptide composition of the protein bands. The main polypeptides, which were similar in all the grape varieties analysed, were identified by LC-MS/MS as homologous to the 11S globulin-like seed storage proteins of other plant species, while a monomeric 43 kDa protein presented high homology with the 7S globulins of legume seeds. The results provide new insights about the identity, structure and polypeptide composition of the grape seed storage proteins.

Contact urticaria and protein contact dermatitis from corn in a patient with serum IgE specific for a salt-soluble corn protein of low molecular weight

Among the cereals, wheat, rye, barley and oats, have been reported to cause protein contact dermatitis. However, in these cases neither the involvement of an immunological mechanism nor the role of specific protein(s) has been demonstrated. We present a case of protein contact dermatitis from corn. The patient presented with a Type I sensitization to corn, as shown by the presence of specific immunoglobulin (Ig)E and positivity to prick tests with both a flour suspension and the salt‐soluble protein fraction of this cereal. The same corn preparations induced a strong urticarial reaction on scratch testing. This reaction was followed several days later by the appearance of erythema and then eczema at the site of application. When boiled, these preparations became inactive on both prick and scratch testing. Patch tests were negative in all cases. Immunoblotting performed with the patients serum showed the presence of a unique IgE‐binding protein band with a molecular weight of around 14 kDa, belonging to the salt‐soluble corn protein fraction. Our results give the first clear evidence that cornflour can induce protein contact dermatitis. The IgE‐binding 14‐kDa protein has characteristics identical to those of the trypsin/α‐amylase inhibitors from cereals.

Allergenic potential of Kamut® wheat

. POLISH WHEAT (Triticum turgidum subspecies (ssp.) polonicum) is an ancient relative of modern durum (pasta) wheat (Triticum turgidum ssp. durum) which originated in the Middle East thousands of years ago. Today, this cereal has been recovered and is traded worldwide under the name Kamut for use in the production of pasta, baked goods and frozen meals. Kamut is frequently recommended as an excellent dietary substitute for wheat for wheathypersensitive (1) people because of its presumed low allergenicity (see for example http://www.kamut.com). However, no data regarding the allergenic properties of this grain are available in the literature. Pooled sera of 10 patients, previously characterized as suffering from IgEmediated food allergy to common and durum wheat products (2), were tested by immunoblotting for IgE-binding to Kamut flour proteins. A sample of T. turgidum ssp. polonicum (kindly provided by Dr E. De Ambrogio, Società Produttori Sementi, Argelato, Italy) and a commercial Kamut sample were compared with Durum wheat.(Fig. 1). Protein extraction, SDS-PAGE and IgEAllergy to fishing bait