Toxicologic evaluation of potassium polyaspartate (A-5D K/SD): Genotoxicity and subchronic toxicity.

Potassium polyaspartate (A-5D K/SD) is proposed for use as a stabiliser in wine, with a maximum use level of 300 mg/L and typical levels in the range of 100-200 mg/L. Potassium polyaspartate (A-5D K/SD) tested negative in a bacterial reverse mutation assay performed in accordance with OECD TG 471 and in an in vitro mammalian cell micronucleus test performed in accordance with OECD TG 487. From a 90-day oral toxicity study in male and female Wistar rats performed in accordance with OECD TG 408, a no observed adverse effect level (NOAEL) was set at 1000 mg/kg bw per day, the highest dose tested. In its opinion adopted on 9 March 2016, the EFSA-ANS Panel (European Food Safety Authority - Panel on Food Additives and Nutrient Sources added to Food), considering these data, concluded that "there was no safety concern from the proposed use and use levels of potassium polyaspartate (A-5D K/SD)".

ePlantLIBRA: A composition and biological activity database for bioactive compounds in plant food supplements.

The newly developed ePlantLIBRA database is a comprehensive and searchable database, with up-to-date coherent and validated scientific information on plant food supplement (PFS) bioactive compounds, with putative health benefits as well as adverse effects, and contaminants and residues. It is the only web-based database available compiling peer reviewed publications and case studies on PFS. A user-friendly, efficient and flexible interface has been developed for searching, extracting, and exporting the data, including links to the original references. Data from over 570 publications have been quality evaluated and entered covering 70 PFS or their botanical ingredients.

The effect of in vitro gastrointestinal digestion on the anti-inflammatory activity of Vitis vinifera L. leaves.

Botanicals are widely consumed all over the world for health purposes, with increased usage in the general population, in many different types of products, including foods and plant food supplements. Several reports support for the beneficial effects of botanicals against gastrointestinal inflammation. However, no studies regarding the anti-inflammatory activity in the gastrointestinal tract of red vine leaves have been reported so far. The present work investigates the biological activity of Vitis vinifera L. water extract (VVWE) from dried leaves in two in vitro models of gastric and intestinal inflammation. The extract was characterized by a validated HPLC-DAD method, and tested on human epithelial gastric (AGS) and intestinal (Caco-2) cells with the aim to investigate the inhibitory effect on IL-8 secretion and promoter activity, before and after in vitro gastric or gastrointestinal digestion. Our results show that the water extract from red vine leaves inhibits TNFα-induced IL-8 secretion and expression in human gastric epithelial cells; the effect should be maintained, although to a lesser extent, after gastric digestion. In contrast, the effect after intestinal digestion is dramatically decreased since degradation of the active components in the gut does not allow the extract to efficiently counteract TNFα or IL-1ß induced IL-8 expression and the NF-κB pathway. The main molecular target of VVWE at the gastric level includes TNFα-induced activation of NF-κB and occurs at concentrations easily reachable after PFS consumption based on red vine leaf water extract as the ingredient. Our findings suggest that PFS containing water extracts from Vitis vinifera L. leaves could be useful to inhibit/attenuate gastric inflammation inhibiting IL-8 secretion and expression through impairment of the NF-κB pathway.

Allergy to all mammalian Bovidae proteins but cow's milk in a child

No disponible

Severe anaphylaxis to sheep's milk cheese in a child desensitized to cow's milk through specific oral tolerance induction.

Specific oral tolerance induction to food (SOTI) is a new promising treatmentfor persistent IgE-mediatedfood allergy. Our paper reports a case of a 5-year-old girl with cow's milk allergy, who developed severe anaphylaxis after the ingestion of a croissant containing sheep's milk ricotta cheese, even though she had been previously desensitized to cow's milk through SOTI. The sheep's milk specific allergen causing the severe allergic reaction (a derivative of alpha-casein of 54,1kDa) was identified by SDS-PAGE and immunoblotting. We conclude that SOTI is a species-specific procedure and the induced tolerance to cow's milk doesn't necessarily provide protection against milk of other mammals. Therefore, children desensitized to cow's milk through SOTI should strictly avoid the intake of milk of other mammals, until tolerance to those kinds of milk is documented by an oral food challenge.

Goat milk allergenicity as a function of αs1-casein genetic polymorphism.

Cow milk allergy is the most frequent allergy in the first years of life. Milk from other mammalian species has been suggested as a possible nutritional alternative to cow milk, but in several cases, the clinical studies showed a high risk of cross-reactivity with cow milk. In the goat species, αS1-casein (αS1-CN), coded by the CSN1S1 gene, is characterized by extensive qualitative and quantitative polymorphisms. Some alleles are associated with null (i.e., CSN1S1 0(1)) or reduced (i.e., CSN1S1 F) expression of the specific protein. The aim of this work was to obtain new information on goat milk and to evaluate its suitability for allergic subjects, depending on the genetic variation at αs1-CN. Individual milk samples from 25 goats with different CSN1S1 genotypes were analyzed by sodium dodecyl sulfate PAGE and immunoblotting, using monoclonal antibodies specific for bovine α-CN and sera from children allergic to cow milk. A lower reaction was observed to 2 goat milk samples characterized by the CSN1S1 0(1)0(1) and 0(1)F genotypes. Moreover, a fresh food skin prick test, carried out on 6 allergic children, showed the lack of positive reaction to the 0(1)0(1) milk sample and only one weak reactivity to the 0(1)F sample. The risk of cross-reactivity between cow and goat milk proteins suggests the need for caution before using goat milk for infant formulas. However, we hypothesize that it can be used successfully in the preparation of modified formulas for selected groups of allergic patients. The importance of taking the individual goat CN genetic variation into account in further experimental studies is evident from the results of the present work.

Respiratory allergy induced by exclusive polysensitization to serum albumins of furry animals.

In this report we describe un unusual case of exclusive allergic sensitization to furry animals, as a possible study model to speculate about different modalities ofsensitization to allergens of common and less common mammalian species. A 27-year-old woman referred in our Allergological Centre for the occurrence of conjunctival and severe respiratory symptoms after contact with several animals such as cats, dogs, rabbits, horses, cows etc. Patient underwent clinical and anamnestic evaluation including a detailed information on the modality of exposure to different furry animals. Skin-prick-test (SPT) was performed with our routine panel of commercial standardized extracts (Lofarma Laboratories, Milan, Italy). Some animal allergenic extracts (rabbit, horse, rat, mouse, cavia, cow and hamster) have been tested by SPT one week after the routine SPT A blood sample was taken for measurement of total IgE and specific IgE (CAP System, Phadia, Uppsala, Sweden) as well as Immunoblotting procedures. The results of in vivo and in vitro procedures revealed allergic sensitization only to animal-derived allergens. Total IgE were 59.3 kU/L. Immunoblotting showed a specific IgE-mediated sensitization of the patient to cow's, rabbit's and horse's serum albumins (SA). In conclusion, our case report confirms the role of SA as cross-reacting agent in allergic sensitization to furry animals. This finding suggests to perform SPTs to several furry animal allergens in all individuals with high level of allergic sensitization to common pets (cats and/or dogs) in order to identify allergy to other animals and consequently to avoid future exposures at risk.

Predicting the outcome of oral food challenges with hen's egg through skin test end-point titration.

BACKGROUND: Oral food challenge (OFC) is the diagnostic 'gold standard' of food allergies but it is laborious and time consuming. Attempts to predict a positive OFC through specific IgE assays or conventional skin tests so far gave suboptimal results. OBJECTIVE: To test whether skin test with titration curves predict with enough confidence the outcome of an oral food challenge. METHODS: Children (n=47; mean age 6.2 +/- 4.2 years) with suspected and diagnosed allergic reactions to hen's egg (HE) were examined through clinical history, physical examination, oral food challenge, conventional and end-point titrated skin tests with HE white extract and determination of serum specific IgE against HE white. Predictive decision points for a positive outcome of food challenges were calculated through receiver operating characteristic (ROC) analysis for HE white using IgE concentration, weal size and end-point titration (EPT). RESULTS: OFC was positive (Sampson's score >or=3) in 20/47 children (42.5%). The area under the ROC curve obtained with the EPT method was significantly bigger than the one obtained by measuring IgE-specific antibodies (0.99 vs. 0.83, P<0.05) and weal size (0.99 vs. 0.88, P<0.05). The extract's dilution that successfully discriminated a positive from a negative OFC (sensitivity 95%, specificity 100%) was 1 : 256, corresponding to a concentration of 5.9 microg/mL of ovotransferrin, 22.2 microg/mL of ovalbumin, and 1.4 microg/mL of lysozyme. CONCLUSION: EPT is a promising approach to optimize the use of skin prick tests and to predict the outcome of OFC with HE in children. Further studies are needed to test whether this encouraging finding can be extended to other populations and food allergens.

Assessment of the tolerance to lupine-enriched pasta in peanut-allergic children.

BACKGROUND: Reports of allergy to lupine derivatives (as de novo sensitization or cross-reactivity in subjects allergic to peanut) are increasing as their use in food products increases. OBJECTIVES: The aim of this study was to assess: (1) lupine tolerance in a group of children allergic to peanut, using lupine enriched-pasta instead of raw flour as has been done in previous clinical studies; (2) whether technological treatments of lupine modify its cross-reactivity or co-sensitization with peanut; (3) the role of lupine seed proteins in sensitization, and (4) to identify the eliciting doses (EDs) by using double-blind, placebo-controlled food challenges (DBPCFC). METHODS: Twelve patients with a history of clinical allergic reactions to peanut were evaluated by skin prick tests (SPTs), the ImmunoCAP test, immunoblotting, and DBPCFC. The 12 selected subjects were included in a trial where lupine-enriched pasta and placebo pasta were administered in a DBPCFC protocol. RESULTS: Positive clinical reactions were observed in two children, the EDs being 0.2 and 6.4 g of pasta, corresponding to 50 mg and 1.6 g of lupine proteins, respectively. Beta-conglutin was the protein most involved in SPT positivity. CONCLUSION: Lupine-enriched pasta can be tolerated by most subjects suffering from peanut allergy, but a sizeable minority (2/12 of them in this case) can develop potentially dangerous clinical reactions. Information about possible reactions to lupine derivatives by those allergic to peanuts must be included in the labelling of lupine-enriched products to protect consumers at risk.

Immunoglobulin-A profile in breast milk from mothers delivering full term and preterm infants.

Recent advances in the care of low-birth-weight and preterm neonates have stimulated research into the best dietetic program to improve their survival and short/long term outcome. Some components of human milk that cannot be included in artificial formulas may be critical for survival. Of these, immunoglobulins are important, and in particular secretory immunoglobulins A (sIgA). The concentration of secretory IgA was measured by immunoblotting (an immunoelectrophoretic technique having high specificity and reliability) in milk from mothers delivering at term (TM) or prematurely (PM). In both groups, IgA concentrations were high very early on but quickly decreased during the first week of lactation. The early IgA mean concentration was higher in PM than in TM but, because of high variability in PM milk, the difference rarely reached statistical significance. This variability during lactation reflects the important role of human milk in supplying immunological factors to cope with the gastrointestinal absorption of high molecular weight proteins in the first days of life. Immunological protection is particularly critical for a preterm baby, so it is important to promote feeding with its own mothers milk if possible, paying strict attention to the timing of milk collection.

A hydrolysed rice-based formula is tolerated by children with cow's milk allergy: a multi-centre study.

BACKGROUND: Children allergic to cow's milk are fed a soy- or a hydrolysed cow's milk-based substitute. Neither can rule out a sensitization risk. Previous studies have shown that hydrolysed rice is tolerated by animals and children with multiple food hypersensitivities. OBJECTIVE: A prospective clinical assessment of tolerance to a rice-based hydrolysed formula was carried out in children allergic to cow's milk. Patients and methods One hundred children (42 girls and 58 boys, mean age 3.17+/-2.93 years, median 2.20, range 0.18-14.6 years) with a history of immediate reactions to cow's milk and confirmed at double-blind, placebo-controlled food challenge (DBPCFC) when not contraindicated were assessed for clinical tolerance to cow's milk proteins. Their allergy work-up included skin prick tests with whole milk, alpha-lactalbumin (ALA), beta-lactoglobulin (BLG) and total caseins, and specific IgE determinations using CAP technology were performed against whole milk, ALA, BLG and casein. Sensitization to rice and rice-based hydrolysed formula was similarly investigated. Patients' sera were evaluated at immunoblotting for specific IgE to cow's milk proteins, rice and rice-based hydrolysed formula. DBPCFC was carried out with increasing doses of a rice-based hydrolysed formula. RESULTS: All patients were sensitized to cow's milk and/or at least one cow's milk protein fraction. Eighty-seven out of 99 were positive to cow's milk and/or a cow's milk protein fraction at skin prick test. Positive (>0.35 kUA/L) specific IgE determinations were found for cow's milk and/or milk fractions (92/95), rice (21/91) and hydrolysed rice infant formula (4/91). At immunoblotting, sera from 96 children were positive to alpha-casein (n=54), beta-casein (n=38), ALA (n=57), BLG (n=37) and bovine serum albumin (n=61). Similarly, although patients' sera often contained specific IgE against rice proteins at CAP (21/91) and immunoblotting (70/96), only six very weakly positive responses were observed against rice-based hydrolysed formula. All DBPCFC with rice-based hydrolysed formula were negative. CONCLUSIONS: Rice-based hydrolysed formula is a possible alternative not only for children with multiple allergies, but also for children with cow's milk allergy.

Immunoblotting in the diagnosis of cross-reactivity in children allergic to birch.

PURPOSE: The scientific experiments with new immunological methods (immunoblotting, RAST inhibition) and isolation of recombinant allergens suggest structural similarities in the allergenic components responsible for cross-reactions. Immunochemical and molecular biology studies indicate that epitopes of major allergen (Bet v 1, Mal d 1) contain more IgE binding epitopes than minor allergens (Bet v 2, Mal d 2), what explained clinical importance of major birch and apple allergens, but it is individual. The important role in cross-reactivity play also proteins with low molecular weight; a potentially dangerous allergen is lipid transfer protein (LTP) inducing severe systemic reactions in allergic subjects. The recent studies indicate that the IgE cross-reactivity patterns and the clinical relevance is still not clear and that only some of patients with confirmed IgE cross allergy to Bet v 1 and Mal d 1 demonstrated clinical symptoms after ingesting of apple. The aim of study was to establish the pattern of cross-reactivity between major (Bet v 1) and minor (Bet v 2) birch pollen allergens and apple proteins in children allergic to birch using recombinant allergens and immunoblotting method. MATERIAL AND METHODS: The prospective study were carried out on the group of 13 children aged 4-16 years, referred to the IIIrd Department of Paediatrics in Bialystok and outpatient clinic with clinical symptoms of food and inhalant allergy. Inclusion criteria to the study were: allergy to birch pollen recombinant allergens and apple, confirmed by presence of specific IgE in the sera of patients. The allergens from peel and pulp of apple and birch were separated and loaded onto the polyacrylamide electrophoretic gel and than transferred to membranes by Western blotting. Antigen-IgE complex was detected using goat anti-human IgE antibodies labelled with alkaline phosphatase. RESULTS: Only few sera presented strong reactions in immunoblotting to birch pollen proteins with a molecular weight of 17-18 kDa, corresponding to the main birch allergen Bet v 1. Most of sera having positive reaction vs Bet v 1 showed cross-reactivity with Mal d 1. All sera recognized specifically the main allergen of apple peel Mal d 3 with molecular weight < 10 kDa (Lipid Transfer Protein). CONCLUSIONS: Immunoblotting method allows to verification of cross-reactivity recognized by presence of specific IgE. The nature of proteins responsible for sensitization can influence the spectrum of offending foods and the clinical features of allergic reactions.

Characterization of bovine serum albumin epitopes and their role in allergic reactions.

OBJECTIVE: This review provides updated information on conformational and sequential epitopes identified in bovine serum albumin (BSA) and summarizes available data about the role of structural modifications on BSA antigenicity/allergenicity. DATA SOURCES: Data on beef allergy and BSA antigenicity are reported, with reference both to the basic literature and to clinical results obtained by our group. RESULTS AND DISCUSSION: BSA is an important allergen involved in milk and beef allergy. The presence of conformational epitopes has been suggested by indirect evidence, while at least one sequential epitope has been experimentally identified. The role of structural modifications on BSA antigenicity is discussed as well as the increased tolerance observed in allergic subjects consuming beef as strained (homogenized) and freeze-dried derivatives. CONCLUSION: Study of the molecular characteristics of a known major allergen allows the identification of technological processes that may be capable of improving the tolerance of allergic subjects to a specific food. Even though any hoped for reduced allergenicity must be verified under medical supervision, the use of new products could obviate the need to avoid important foods such as meat in childhood.

Ochratoxin A in conventional and organic cereal derivatives: a survey of the Italian market, 2001-02.

Ochratoxin A is a mycotoxin produced mainly by Penicillium verrucosum and Aspergillus ochraceus. Although typically considered a cereal contaminant, it has also been detected in dried fruit, nuts, meat and derivatives. To estimate the quantity of ochratoxin A that might be ingested by Italian consumers from these foods, 211 cereal derivatives (flours and bakery products) were analysed by high-performance liquid chromatography. Products were from conventional and organic agriculture and from integrated pest management agriculture. All commercial flours and derivatives examined contained ochratoxin A at concentrations very much below the legal limit (3 microg kg(-1)): the highest value, 0.816 microg kg(-1), was detected in a sample of spelt whole flour from organic agriculture. In many samples, the ochratoxin content was below the limit of detection; only rarely did values exceed 0.5 microg kg(-1). In baby foods, four samples were above the particularly restrictive Italian legal limit of 0.5 microg kg(-1). Although some significant differences were found between samples from conventional and organic agriculture when some product categories were examined (namely, baby foods as semolina and rice creams), no important difference was found between the two types of agricultural practice when all types of cereal derivatives were considered together.

Evaluation of residual immunoreactivity in red and white wines clarified with gluten or gluten derivatives.

Gluten or hydrolyzed gluten could be a suitable alternative to animal proteins in the wine clarification process, but their residues could represent a risk for individuals suffering from coeliac disease or allergic to cereal proteins. The aim of this study was to investigate the presence of gluten in wines treated with gluten or its hydrolysate in the clarification process and to assess its antigenicity in commercial products. The presence of residual immunoreactive gluten was evaluated by electrophoresis (SDS-PAGE) and immunoblotting. Data obtained in several red and white wine samples showed that no residue was detectable in any of the red wines. In white wines, gluten reduced the protein content less completely, but most samples showed no immunoreactivity after the wine had been treated with gluten or its derivatives, either alone or combined with bentonite, silica gel or tannins. The use of gluten derivatives coupled with bentonite was the most effective method of removing immunoreactive protein in white wines. In conclusion, the use of gluten derivatives in wine clarification seems to exclude a risk for subjects susceptible to coeliac disease or gluten allergy. However, it is recommended that wine producers continuously monitor the clarification process in order to protect the most sensitive individuals.

Determination of nitrates by a novel ion chromatographic method: occurrence in leafy vegetables (organic and conventional) and exposure assessment for Italian consumers.

A novel ion chromatographic method to detect nitrates in vegetables was developed, and the nitrate contents in green salad (a mixture of endive and prickly lettuce), lettuce, chicory, rocket and spinach were determined from Italian markets in 1996-2002. These leaf vegetables were included because they are currently supposed to provide most of the nitrate intake in the typical Italian diet. The highest content of nitrate was detected in chicory (6250 mg kg(-1)) and rocket (6120 mg kg(-1)), which are consumed in large quantities in some regions of Italy. Green salad and lettuce contained less nitrate (highest values = 4200 and 3300 mg kg(-1), respectively), but because they are consumed more generally, they provided 60% of the total intake of nitrates. Only a few samples were above the legal limits, with seasonal variation. A significantly higher nitrate content was found in organically grown green salad and rocket than in those conventionally produced. These data indicate that the average intake of nitrates from leafy vegetables is below the acceptable daily intake, i.e. 3.7 mg nitrate ion kg(-1) body weight day(-1), but the total intake should be monitored to protect groups at risk, such as children and vegetarians.

Assessment of residual immunoreactivity in red or white wines clarified with pea or lupin extracts.

Vegetable proteins could be a suitable alternative to animal proteins in the clarification of wine, but their residues could represent a risk for subjects with food allergy or intolerance. The aim of this study was to investigate the presence of specific immunoreactivity in red and white wines treated, as must or wine, with vegetable proteins in the clarification process. The proteins considered were prepared from lupins and peas, which are not included among the allergens listed in annex Illbis of Directive 2003/89/EC. The presence of residual immunoreactivity to specific rabbit anti-lupin and anti-pea polyclonal antibodies in treated wines was assessed by electrophoresis (SDS-PAGE) and immunoblotting. Residual protein was not detectable in red wines clarified with lupin, pea or a mixture of pea and lupin proteins or in white wines clarified with pea proteins. A small number of musts treated with lupin or pea proteins and white wines treated with lupin proteins yielded equivocal results, probably because of the presence of interfering material (e.g., sugar-rich proteins from grape and yeast). The use of bentonite as a secondary clarifying agent is therefore recommended since its combination with vegetable proteins is particularly effective in removing overall protein immunoreactivity.