1. Introduction
Although the consumption of bread has showed a progressive decrease in recent decades, bread is considered one of the most important staple foods in the world. Its consumption in the last years was reported to be approximately 60 kg per capita per year in the Western countries [
1].
Bread is commonly made using wheat flour, in which gluten is the main protein source. However, a growing part of the population is affected by several diseases related to gluten intake. Indeed, celiac disease (CD) is the most common and increasing food intolerance, affecting approximately 1% of the worldwide population [
2]. In addition to CD, there are several disorders related to gluten intake, called gluten-related disorders, which include non-celiac gluten sensitivity (NCGS), dermatitis herpetiformis, wheat allergy, gluten ataxia, and other chronic inflammatory diseases (e.g., inflammatory bowel diseases, IBD), usually co-occurring with CD [
3,
4].
The primary treatment to avoid complications related to gluten assumption is the strict adherence to a gluten-free diet (GFD) [
5]. Overall, among GF ingredients, corn and rice flours are extensively used to make GF bread [
6]. However, several technological issues lead to difficulties in the production of GF bread. Indeed, gluten is the main structuring agent in wheat dough and confers its baking quality [
7]. Several approaches were investigated to overcome the absence of wheat protein network, including the use of a wide range of ingredients and additives [
8,
9]. Structuring ingredients are usually classified in three different categories, which include (i) water-binding and film-forming ingredients (e.g., hydrocolloids or thickening agents), (ii) structure-forming, volume-filling, and taste-giving ingredients (e.g., proteins, fats, and low molecular weight carbohydrates), and (iii) surface-active substances (e.g., emulsifiers). However, the inclusion of many ingredients and additives give the perception of a high processed food, thus leading to consumers’ dissatisfaction. Indeed, in recent years, the consumer’s choice is continuously evolving toward a niche of products considered safer, which are referred to as “clean-label products” [
10,
11]. “Clean-label” is not a scientific definition, but it is a popular definition broadly accepted by the food industry, consumers, the scientific community, and even regulatory agencies. The definition refers to products made by using as few ingredients as possible, making sure they could be easily identified by the consumers as no-artificial, no-synthetic, wholesome ingredients [
10,
11].
The inclusion of non-wheat plant ingredients and the application of new technologies were recently proposed to overcome this issue. Being water-binding, film-forming, and a source of protein, fiber, and minerals [
12,
13], the use of pseudo-cereals and pulses in breadmaking has largely been encouraged, both in gluten-containing and gluten-free products [
14,
15,
16].
In addition to flours deriving from legumes and pseudo-cereals, whose importance in the GF sector is mainly related to the nutritional and functional properties, other flours gained the attention of the producers as texture improvers. Psyllium flour is reported as one of the most important ingredients already used in commercial GF bread due to its structuring properties [
9,
17]. Other seeds (or derived flours) such as chia and flaxseed have been proposed thanks to their water-binding capacity and production of mucilage [
18], which favor the workability and cohesiveness of the GF doughs, and the hydration and softness of the GF breads.
In addition to the selection of proper ingredients, bioprocessing could moreover affect the nutritional and functional properties of GF bread. In particular, sourdough fermentation has largely been reported as a natural and effective biotechnology to improve the textural, sensorial, and nutritional properties as well as the shelf life of GF baked goods [
19,
20]. A proper selection of lactic acid bacteria (LAB) having specific metabolic traits able to affect the technological and the nutritional features of the product is strongly suggested [
21]. Among all, the synthesis of exopolysaccharides (EPS) has recently gained attention due to the ability to counteract the negative effects associated with high levels of sourdough acidification and enhance the loaf volume [
22]. EPS-producing strains belonging to the
Weissella genus were widely investigated as potential starters to be employed in food fermentations. Indeed, the relevance of these heterofermentative LAB is related to their ability to grow at wide temperature, a
w, and pH ranges [
23] and to produce EPS that may improve the rheological properties of doughs [
24].
This study aimed at optimizing a formulation and breadmaking procedure to obtain a novel “clean-label” GF bread. The formulation of the recipe was based on the selection of different ingredients naturally characterized by (i) high protein content, (ii) structuring properties, and (iii) high sucrose concentration, this latter able to promote the in situ EPS production by the selected Weissella cibaria P9. The bread made with the optimized production protocol was characterized for the main technological, nutritional, and sensory features.
4. Discussion
In addition to celiac patients, the increasing consumer demand for products with “reduced” or “no gluten” content categories to face gluten sensitivity and lactose intolerance disturbances requires the innovation and differentiation of gluten-free products [
47]. Moreover, the amount of attention on the “free-from” foods, mainly because consumers consider them healthier [
48], is also forcing the industries toward the production of “clean-label” products. Although, a unique definition of this term as well as the features that a “clean label” food should highlight are still debated [
10], the preparation of a “clean label” GF bread should avoid the use of chemical additives [
10], which in turn are usually included in such bread formula. Indeed, although the technological process allowed acquiring significant achievements in the overall quality of the gluten-free products, the absence of gluten still represents an industrial challenge to produce baked goods [
49]. Moreover, concerns about the nutritional composition of gluten-free foods are often reported [
50]. To face nutritional deficiencies, the inclusion of a protein source in GF bread is a consolidate strategy; nevertheless, the final protein content tends to be lower than that of wheat-containing counterparts [
50,
51]. Overall, plant-derived proteins are preferred to animal protein sources (e.g., egg and dairy products) because, thanks to their better water-retention capacity, they exert positive effects on the texture of GF bread [
52].
Sourdough technology has widely been used to improve the physicochemical, sensory, and nutritional attributes of GF bread [
53]. Indeed, different GF cereals such as sorghum, millet, rice, corn, and buckwheat have been investigated as ingredients to develop GF sourdough breads [
53,
54,
55,
56].
The positive effects of sourdough are mainly due to the metabolic activities of the lactic acid bacteria performing the fermentation, and recently, the use of
Weissella species as novel starter cultures for sourdough bread production has been reported by several authors [
57,
58,
59,
60].
In this framework, the present study aimed at optimizing a GF bread formulation including not-refined plant-based flours as a source of protein and structuring ingredients. The use of a Type-II sourdough obtained through Weissella cibaria P9 fermentation, an EPS-producing strain, has also been evaluated. The effect of the ingredients and sourdough fermentation on the main structural, sensory, and nutritional properties of the bread has been investigated.
A preliminary evaluation of the effect of protein-rich flours on the dough leavening performance (volume increase) and bread structural properties (specific volume and alveolus percentage) has shown a positive influence of quinoa, teff, and lentil flours. The inclusion of quinoa and teff has already been reported as effective in improving the leavening capability of GF bread [
61,
62], such as the nutritional and sensorial quality of GF bread [
14,
63]. Moreover, Asif and colleagues demonstrated that lentil protein isolates possess good foaming and emulsifying properties [
64]. The effects of teff, quinoa, and lentil flours on the sensory profile of bread were also assessed. The inclusion of quinoa led to the enhancement of crust and crumb color, as previously reported by Turkut and colleagues [
62], meeting the consumer concerns about the common light color of GF bread [
65]. Moreover, the inclusion of 20–25% of quinoa flour was already proposed by several authors as a reliable tool to improve the sensory acceptability of conventional and GF breads [
39,
66].
The natural structuring ingredients chia (raw and hydrated), flaxseed, and psyllium flours were tested singly and as a mixture to evaluate the technological effect in GF bread formulation. Studies on commercial GF breads revealed the use of a combination of several starchy sources or gluten replacers to optimize bread quality [
9]. Despite the increasing inclusion in GF commercial bread, very few research studies incorporate psyllium, and no study evaluated psyllium-based formulas including other natural structuring ingredients [
9,
67]. In this work, the supplementation of all the structuring flours improved the volume increase and the structural properties when singly used, as already reported elsewhere [
26,
67,
68,
69,
70]. However, when psyllium, hydrated chia, and flaxseed were combined, a further enhancement of the textural quality of dough and bread was achieved.
Aiming at setting up a “clean-label” product, EPS were produced in situ by using
W. cibaria P9 on matrices naturally containing sucrose. Although belonging to the natural microbiota of several fermented food products [
71],
Weissella species have not yet been included in the Qualified Presumption of Safety (QPS) list of European Food Safety Authority (EFSA) due to the safety concerns [
71,
72]. However, the rare and opportunistic features of the infection cases related to
Weissella strains have recently been highlighted, and its species been proposed to be granted the QPS status [
72].
EPS display physiochemical properties similar to commercial purified hydrocolloids, allowing the improvement of bread structure and loaf volume, enhancing crumb softness and delaying bread staling [
22]. Moreover, dextrans produced by
W. cibaria were recently associated with a positive modulation of gut microbiota and influence on body weight and metabolism [
73]. The in situ formation of EPS (homo-polysaccharides) in sourdough was reported to be more effective than external addition [
30] and does not require labeling; thus, this strengthens their case for use in “clean-label” products [
74].
As expected, the higher the content of the sucrose in the dough, the higher the EPS content at the end of the fermentation. Although the highest percentage of chestnut flours to avoid sensory or technological adverse effects was used [
47], a low amount of EPS was produced during sourdough fermentation. The relatively low amount of sucrose in the dough and the short fermentation time [
75,
76] were reported as responsible for the poor EPS synthesis during fermentation [
77,
78]. However, the role of texture improvers was, in the proposed formulation, synergistic among different components, including the different structuring flours.
The final formulation of GF bread, including (i) quinoa as a protein source, (ii) the mixture of psyllium, hydrated chia, and flaxseed as a structuring (4:1:1) agent, and (iii) the chestnut sourdough containing EPS as improver, was characterized by a protein content that was more than 50% higher than a GF product usually found in the market [
39,
50], and it also had a high value of in vitro protein digestibility (76.9 ± 2.7) [
39,
51]. Moreover, the optimized bread formula contained a low amount of lipids, which were mainly represented by unsaturated fatty acids, previously characterized by an optimal n-6/n-3 polyunsaturated fatty acids ratio and recognized as carriers of nutraceutical components such as tocopherols and sterols [
18,
79,
80]. Finally, the low sugar content of bread led to a similar in vitro glycemic index as compared to commercial GF bread [
39,
81].
5. Conclusions
Here, the formulation of the novel “clean-label” GF bread has been optimized by using (i) a mixture of corn and rice flour (ratio 1:1); (ii) quinoa flour (10%) as a source of protein, and (iii) psyllium, flaxseed, and hydrated chia (6%) as structuring agents. Moreover, a type-II sourdough, which was obtained by using a selected Weissella cibaria P9 and chestnuts flour as a sucrose source to promote the in situ exo-polysaccharides syntheses, was included (30% of the final dough) in the bread formulation. Overall, the novel GF bread was characterized by good textural properties, high protein content (8.9% of dry matter) and in vitro protein digestibility (76.9%), low sugar (1.0% of dry matter) and fat (3.1% of dry matter) content, and an in vitro predicted glycemic index of 85. Moreover, common sourdough bread features, i.e., acidic taste and darker color of both crust and crumb, were identified in the “clean-label” GF bread. This work demonstrates that a proper selection of raw materials (unprocessed flours), together with the use of traditionally inspired biotechnology, allow the production of high-quality bread, meeting the demand of the modern consumer for novel clean-label GF products.