Stability of food allergens to digestion in vitro

Abstract

An integral part of the safety assessment of genetically modified plants is consideration of possible human health effects, especially food allergy. Prospective testing for allergenicity of proteins obtained from sources with no prior history of causing allergy has been difficult because of the absence of valid methods and models. Food allergens may share physicochemical properties that distinguish them from nonallergens, properties that may be used as a tool to predict the inherent allergenicity of proteins newly introduced into the food supply by genetic engineering. One candidate property is stability to digestion. We have systematically evaluated the stability of food allergens that are active via the gastrointestinal tract in a simple model of gastric digestion, emphasizing the major allergens of plant-derived foods such as legumes (peanuts and soybean). Important food allergens were stable to digestion in the gastric model (simulated gastric fluid). For example, soybean beta-conglycinin was stable for 60 min. In contrast, nonallergenic food proteins, such as spinach ribulose bis-phosphate carboxylase/oxygenase, were digested in simulated gastric fluid within 15 sec. The data are consistent with the hypothesis that food allergens must exhibit sufficient gastric stability to reach the intestinal mucosa where absorption and sensitization (development of atopy) can occur. Thus, the stability to digestion is a significant and valid parameter that distinguishes food allergens from nonallergens.

Mass spectrometric characterization of glycated β-lactoglobulin peptides derived from galacto-oligosaccharides surviving the in vitro gastrointestinal digestion

“…As expected, both proteins were very resistant to pepsinolysis (data not shown) because ␤-LG is a poor substrate for pepsin [27][28][29][30]. Nevertheless, the duodenal digestion (trypsin and chymotrypsin) of the native and glycated ␤-LG gave rise to a very complex mixture of peptides after 1 h of incubation as was previously observed by reversed-phase LC-UV [13].…”

Structure of the Major Peanut Allergen Ara h 1 May Protect IgE-Binding Epitopes from Degradation

“…It is a 65-kDa glycoprotein with an acidic isoelectric point. It is an abundant protein in the peanut (11) that survives food-processing methods intact (6) and is stable up to 1 h within the in vitro digestion systems designed to mimic the gastrointestinal tract (6). Resistance of allergens to digestive enzymes have been attributed to various factors including protease inhibitors or nonprotein components present in the extracts analyzed (7,24), direct effects on the secretion of endogenous proteins and/or the structure of the allergen itself (25).…”

“…glycoproteins (Ͻ70 kDa) with acidic isoelectric points that are highly abundant in food. These proteins are usually resistant to proteases, heat, and denaturants, allowing them to resist degradation during food preparation and digestion (6,7). Several studies have shown that the most allergenic portion of the peanut is the protein fraction of the cotyledon (8)(9)(10).…”

A Pepsin-Resistant 20 kDa Protein Found in Red Kidney Bean (Phaseolus vulgarisL.) Identified as Basic Subunit of Legumin

“…The digestibility of protein causes another concern in respect of risk of allergenicity. Astwood et al 2) suggested that stability to pepsin digestion is a significant and valid parameter that distinguishes food allergens from non-allergens. In this study, the protein quality of red kidney bean, one of the representative varieties of common beans in Japan, was assessed by in vitro pepsin digestion.…”

“…The protein concentration of the extracts was estimated by the micro assay procedure for a protein assay reagent (BioRad, Tokyo). The in vitro pepsin digestibility of intact flour and the heat-processed paste was examined by the method of Astwood et al 2) Intermolecular and intramolecular disulfide bonds were analyzed by nonreducing/reducing two-dimensional electrophoresis. Bean flour (200 mg) was mixed with 2 ml of extraction buffer (30 mM Tris-HCl, pH 7.9) containing 1 mM phenylmethanesulphonyl fluoride (PMSF) and 0.02% NaN 3 , then centrifuged at 10;000 Â g for 15 min.…”