Use of fungal proteases and selected sourdough lactic acid bacteria for making wheat bread with an intermediate content of gluten Carlo Giuseppe Rizzello a , José Antonio Curiel a , Luana Nionelli a , Olimpia Vincentini b , Raffaella Di Cagno a , Marco Silano b , Marco Gobbetti a , Rossana Coda a, * a Department of Soil, Plant and Food Sciences, University of Bari, 70126 Bari, Italy b Unit of Human Nutrition and Health, Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, 00161 Roma, Italy article info Article history: Available online 5 July 2013 Keywords: Lactic acid bacteria Sourdough Gluten Proteolysis Bread abstract This study was aimed at combining the highest degradation of gluten during wheat flour fermentation with good structural and sensory features of the related bread. As estimated by R5-ELISA, the degree of degradation of immune reactive gluten was ca. 28%. Two-dimensional electrophoresis and RP-FPLC analyses showed marked variations of the protein fractions compared to the untreated flour. The com- parison was also extended to in vitro effect of the peptic/tryptic-digests towards K562 and T84 cells. The flour with the intermediate content of gluten (ICG) was used for bread making, and compared to whole gluten (WG) bread. The chemical, structural and sensory features of the ICG bread approached those of the bread made with WG flour. The protein digestibility of the ICG bread was higher than that from WG flour. Also the nutritional quality, as estimated by different indexes, was the highest for ICG bread. Ó 2013 Elsevier Ltd. All rights reserved. 1. Introduction Wheat is the most widely grown crop with more than 25,000 different cultivars (Sapone et al., 2012). The large part of this global production is consumed after processing into bread, other baked goods, pasta and noodles, or, as in the case of Middle East and North Africa, into bulgur and couscous. The functional properties of the gluten proteins make also the wheat flour as an irreplaceable ingredient for other foods (Sapone et al., 2012). Gluten is the abundant structural protein complex of wheat, with equivalent proteins found in other cereals (e.g., rye and barley). Although the wide spread of gluten containing grains initiated ca. 10,000 years ago, more recently wheat breeding was addressed to massive selection of cultivars with an unusual and elevated content of gluten. The daily human exposure to such elevated levels of gluten suggested the possibility that this evolu- tionary challenge also created conditions for related human dis- eases (Sapone et al., 2012). Wheat allergy (WA) and celiac disease (CD), which are mediated by adaptive immune systems, are the most known diseases related to gluten (Sapone et al., 2012). Under both these conditions, gluten reaction occurs via T-cell activation at the gastrointestinal mucosa level. Cross-linking between immu- noglobulin (Ig)E and gluten epitopes is responsible for WA, and it triggers the release of chemical mediators (e.g., histamine) from basophils and mast cells. CD is an autoimmune disorder, which mainly involves the response of serum anti-tissue trans- glutaminase (tTG) and anti-endomysial antibodies (EMA). Other cases of reaction to gluten are commonly described as gluten sensitivity (GS), and they do not involve allergic or autoimmune mechanisms. Intestinal (e.g., diarrhea, abdominal discomfort or pain, bloating) or extra-intestinal (headache, lethargy, attention- deficit/hyperactivity disorder, ataxia or recurrent oral ulceration) symptoms are often manifested during GS (Di Sabatino and Corazza, 2012). Since the clinical symptoms are somewhat over- lapping, the correlation between irritable bowel syndrome (IBS), CD and GS recently received a marked interest. It is well established that GS symptoms decrease or disappear after gluten is withdrawn from the diet (Di Sabatino and Corazza, 2012). The information about the level of gluten that is respon- sible for the disease and about the mechanisms that cause Abbreviations: BV, biological value; CD, celiac disease; CS, chemical score; DY, dough yield; EAA, essential amino acid; EAAI, essential amino acid index; EMA, anti-endomysial antibodies; FAA, free amino acids; FHWF, fully hydrolyzed wheat flour; GS, gluten sensitive; IBS, irritable bowel syndrome; ICG, intermediate content of gluten; IEF, isoelectric focusing; IPG, immobilized pH gradient; MAC, minimal agglutinating capacity; NI, nutritional index; OPA, o-phtaldialdehyde; PER, protein efficiency ratio; PT-, peptic/tryptic; TFA, trifluoroacetic acid; TPA, texture profile analysis; TTA, titratable acidity; tTG, anti-tissue transglutaminase; WA, wheat al- lergy; WG, whole gluten. * Corresponding author. Tel.: þ39 080 5442948. E-mail address: rossana.coda@gmail.com (R. Coda). Contents lists available at SciVerse ScienceDirect Food Microbiology journal homepage: www.elsevier.com/locate/fm 0740-0020/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.fm.2013.06.017 Food Microbiology 37 (2014) 59e68