Heat-Induced Cross-Linking and Degradation of Wheat Gluten, Serum Albumin, and Mixtures Thereof Ine Rombouts,* Bert Lagrain, and Jan A. Delcour Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium ABSTRACT: Some wheat-based food systems, such as cakes, cookies, and egg noodles, contain mixtures of animal and plant (gluten) proteins and are processed under (mildly) alkaline conditions. Although changes in these proteins during processing can affect end product quality, they have seldom been studied. This study investigated protein cross-linking and degradation during heating (0-120 min, pH 8.0, 50-130 °C) of (mixtures of) wheat gluten and bovine serum albumin (BSA). The decrease in protein extractabilities in sodium dodecyl sulfate containing buffer under (non)reducing conditions and the levels of (cross- linked) amino acids were measured. No indications for polymerization at 50 °C were found. Below 100 °C, BSA polymerized more readily than wheat gluten. Above 100 °C, the opposite was observed. The kinetics of heat-induced polymerization of a 1:1 gluten-BSA mixture were similar to that of isolated gluten, implying that gluten decelerated BSA denaturation. Severe heating (130 °C, >15 min) induced degradation reactions in gluten but not in BSA. At all conditions used in this study, disulfide (SS) bonds contributed to the extractability loss. In addition, above 110 °C, β-elimination of cystine led to non-SS cross-links. Intramolecular SS bonds more often transformed in intermolecular non-SS bonds in BSA than in gluten. KEYWORDS: bovine serum albumin, wheat gluten, aggregation, polymerization, protein network, degradation, disulfide, β-elimination, lanthionine, lysinoalanine, dehydroalanine ■ INTRODUCTION The most important food protein source worldwide is wheat, with an average consumption of 16 g of protein/capita/day. 1,2 Wheat flour proteins are classified into albumin, globulin, gliadin and glutenin fractions by sequential extraction in water, dilute salt solutions, aqueous ethanol, and dilute acid. 3 Albumins and globulins represent about 20% of all wheat flour proteins. Gliadins and glutenins are quantitatively more important and develop into gluten upon mixing flour with water. Gluten proteins are of great technological importance in many food systems. 4 Baking, boiling, or extrusion steps during cereal-based food production induce reshuffling of existing and formation of additional intermolecular disulfide (SS) cross- links, thereby forming a strong gluten network, which greatly determines the structure of, for example, bread 5 and pasta. 6 SS cross-links, which can be cleaved by reduction, are the most studied cross-links with regard to gluten network formation. In addition, at high temperatures and/or alkaline conditions, β- elimination of cystine releases a free sulfhydryl group and the reactive intermediate dehydroalanine (DHA). DHA can then react with cysteine or lysine to form the nonreducible cross-link lanthionine (LAN) or lysinoalanine (LAL), respectively. 7 Such cross-links are usually less abundant, but are important because of their more permanent character. In contrast with most proteins, gluten proteins fail to show a definite denaturation peak in differential scanning calorimetry (DSC) thermograms. 8 Therefore, heat-induced gluten changes are typically evaluated on the basis of gluten extractability loss in sodium dodecyl sulfate (SDS)-containing media, 9-13 which generally follows first-order kinetics. 5,14,15 The impact of temperature on the loss of gluten extractability during heating at mildly alkaline pH has, to the best of our knowledge, not yet been reported. The formation of nonreducible cross-links is accompanied by the loss of extractability in SDS-containing media even if they contain a reducing agent. 7 β-Elimination of cystine during hydrothermal treatment of isolated gliadin also follows first- order kinetics. 16 For the entire gluten protein mixture, the kinetics of β-elimination of cystine during heat treatments at alkaline pH remain to be studied. In most bread and pasta products, gliadins and glutenins are by far the most abundant proteins. However, some cereal-based foods also contain milk, soy, or egg proteins for structural, nutritional, or taste-related reasons. In, for example, cakes, cookies, pancakes, milk bread, and egg noodles, which are sometimes processed at (mildly) alkaline pH due to the use of stored eggs and/or baking powder, wheat gluten proteins co- occur with albumins from eggs and/or milk. Here, changes in both wheat gluten proteins and albumins during processing can affect end product quality. One of the most important functional properties of albumins is their ability to denature. Denaturation depends on temperature, pH, and protein concentration. It starts with the exposure of hydrophobic groups above the protein unfolding or denaturation temper- ature and can result in protein aggregation due to interprotein noncovalent bonding (hydrogen and hydrophobic) as well as covalent cross-linking (often through SS bonds). 17 When intermolecular interactions are depressed, the denaturation is reversible. 18 However, more typical is the case of irreversible denaturation, 19 in which large clusters are formed and gel Received: June 6, 2012 Revised: August 21, 2012 Accepted: September 5, 2012 Article pubs.acs.org/JAFC © XXXX American Chemical Society A dx.doi.org/10.1021/jf3024672 | J. Agric. Food Chem. XXXX, XXX, XXX-XXX