Sourdoughs as Natural Enhancers of Bread Quality and Shelf Life: A Review

: Sourdough is a key component in traditional and artisanal bread making. It imparts unique flavors and textures to bread, which are highly sought after by consumers. The use of sourdoughs to prepare bakery products has been researched for more than 30 years, and accumulated research shows the performance of sourdoughs as an alternative to improve the organoleptic characteristics of bread and its shelf life. The purpose of this review is to present an overview of the research carried out on the use of sourdoughs from lactic acid bacteria and their benefits in the quality characteristics of bread, as well as to present relevant and recent information on the use of sourdoughs and their aqueous extracts for the preservation of bakery products. Also, the advances in the identification of antifungal compounds have been revised. In general, it has been shown that incorporating sourdoughs into the bread formulation positively impacts the product’s flavor and helps slow down the bread’s aging process and spoilage. Also, it has been observed that the bioactive compounds formed by lactic acid bacteria (LAB) during sourdough fermentation and their extracts have an antimicrobial, especially antifungal, capacity that significantly helps increase bread’s shelf life. Studying sourdough as part of fermentation processes and product development is essential to improve bread production’s quality, diversity, and sustainability, and to advance our understanding of the science behind this food tradition.


Introduction
Bread is one of the most consumed fermented foods in the world and is an important part of the food base of many countries [1].In general, bread is a product made by baking a dough previously fermented with yeast and, in some types of bread, other micro-organisms that give it the important characteristics and quality attributes of this product.However, fresh bread has a short shelf life, governed mainly by physicochemical alterations known as aging and mold growth on the product's surface.To improve the shelf life of bakery products, the baking industry has used the incorporation of synthetic additives and antimicrobial agents, which have been shown to have the ability to reduce both the aging process of bread and the growth of molds on the surface of baking products.However, consumer demand for natural products or products without synthetic additives has generated the search for new "natural" alternatives that help replace these synthetic additives and maintain the shelf life of bread [2,3].
One of the oldest ways of bread leavening is sourdough fermentation, converting cereal flour into attractive, tastier, and more digestible products [1,4].An alternative approach that has attracted the attention of the scientific community in recent years is the use of sourdough to leaven various bakery products, as well as to incorporate it into the formulation of other products, which creates research opportunities in the use of sourdoughs and their fermentation products with very diverse applications and approaches [1,5].An alternative that has attracted attention in recent years is to use sourdough together with baker's yeast to produce bread at an industrial level and have the benefits of its incorporation in the formulation [6].For many years, sourdoughs have been used in the European and Western regions to make bread [7] since it has been observed that incorporating them helps maintain the product's organoleptic characteristics and delays the growth of molds on the product's surface.This fermented dough has a complex microbiome that mainly includes lactic acid bacteria and yeasts [4].It has been shown that during the formation of sourdoughs, compounds such as organic acids, peptide compounds, and exopolysaccharides are formed, which help maintain the shelf life of the product, preserving the quality characteristics of the bread and reducing the rate in the growth of molds on bread surface [3,[8][9][10][11].Likewise, in recent years, the ability of aqueous extracts from fermented sourdoughs to inhibit the growth of the main molds that spoil bread has been demonstrated, thus generating promising alternatives as natural antimicrobials in bakery products [11][12][13][14][15].
Bread is highly perishable due to its composition, leading to a quality decline from physicochemical and microbiological changes after baking.Post-baking cooling results in moisture loss, affecting crumb and crust texture.Over time, starch retrogradation and moisture loss increase hardness.Mold growth on the bread surface poses a significant economic challenge for the baking industry [16].
Molds proliferate easily in bread because it is rich in carbohydrates (70-80 g carbohydrates/100 g bread), with water activity values between 0.94-0.97and pH values around 6, which are optimal conditions for mold growth.Contamination of bakery products with mold spores, which are naturally found in the bakery environment, has been shown to occur after the baking process, during cooling, slicing, and packaging of the product, these last stages being the determinants to define the microbiological quality of these products [16,17].
Penicillium and Aspergillus are common molds that cause bread spoilage due to their ability to thrive in diverse conditions, produce numerous spores, and exist in the environment.Notable species include Penicillium roqueforti, P. paneum, P. corylophilum, P. chrysogenum, and others, leading to blue-green spots on bread.Aspergillus species such as A. chevalieri and A. niger cause colored spots, ranging from green to black, with some producing yellow pigments.Additional filamentous molds like Rhizopus, Mucor, and Neurospora, while present in various baking products, are less significant than Penicillium and Aspergillus [16,17].
Bacteria such as Serratia marcescens and Endomyces fibulger can cause red or white stains on bread.Ropy bread, characterized by yellow color and sticky viscosity, results from specific strains of Bacillus subtilis.However, bacterial growth on bread is infrequent compared to mold growth, leading the baking industry to prioritize the elimination of mold contamination [17,18].
The baking industry, facing substantial economic losses from mold spoilage of bread, is increasingly developing preservation methods.Propionic, sorbic, and acetic acids, and their salts, are common antifungals for mold prevention [14].In response to consumer demand for "natural" additives, research explores new compounds to enhance the shelf life of baking products.Utilizing sourdoughs or their aqueous extracts in bread formulation is considered an alternative, with numerous studies describing their effectiveness as antifungals.
This review focuses on recent developments in the utilization of sourdoughs, specifically those fermented with lactic acid bacteria and their corresponding aqueous extracts, aimed at augmenting bread quality and prolonging shelf life.The investigation examines the fundamental roles played by lactic acid bacteria and yeasts in generating organic acids, peptide compounds, and exopolysaccharides within sourdoughs.Additionally, it highlights the potential of aqueous extracts derived from sourdoughs to serve as natural antimicrobials, with a targeted focus on molds that compromise bread quality.Also, it attempts to identify the specific compounds inherent in sourdough that are responsible for its antifungal properties.Through this identification, the research aims to elucidate potential paths for future studies and the development of various bakery products where sourdough can be an alternative.As the baking industry contends with substantial eco-nomic losses from mold spoilage, this review contributes to evaluating the possibility of transforming the baking industry towards natural and sustainable conservation methods, emphasizing the multifaceted advantages of sourdough fermentation, thereby contributing to the ongoing scientific search for high-quality bakery products that align with evolving consumer preferences.

Sourdoughs
Sourdough is one of the oldest examples of natural starters, mostly used for making fermented bread as an alternative to baker's yeast and chemical leavening.The term "sourdough" can vary in different countries and languages.However, the different linguistic expressions capture the idea of a natural leaven or sourdough starter, such as in French: levain, Italian: lievito madre, German: Sauerteig, Spanish: masa madre or levadura madre, Portuguese: fermento natural, Dutch: zuurdesem, and many others.Sourdoughs are the result of spontaneous fermentations that are obtained when flours from different grains (mainly wheat or barley) are mixed with an adequate amount of water and maintained under conditions of temperature and time favorable for the growth of microorganisms inherent to the flour or intentionally inoculated [19].In many countries worldwide, sourdoughs continue to be used as part of their formulation due to their unique texture and flavor characteristics in the final product [20].During sourdough fermentation, metabolite production and chemical reactions occur, giving the product distinctive characteristics and a high sensory quality [5].Likewise, in recent years, the use of sourdough in the baking industry has increased significantly due to the discovery of nutritional improvements, such as the enhancement of the bioavailability of minerals, the production of peptides with antioxidant activity, and the preservative effects on bread, which improve the shelf life [3,21].
Microbial analyses and investigations of sourdough microbiota have shown that two types of microorganisms coexist within this ecosystem: lactic acid bacteria (LAB) and yeasts [22].A majority of sourdoughs that are used in bakeries as sole leavening agents include Fructilactobacillus sanfranciscensis (formerly Lactobacillus sanfranciscensis), which is one of the dominant fermentation microbes [23,24].In sourdoughs fermented to achieve high acidity levels, Limosilactobacillus and Lactobacillus species typically dominate [24].In most analyses, the LAB commonly isolated from sourdoughs are of the Lactobacillus genus, whereas Saccharomyces spp.and Candida spp.have mostly been isolated and identified as yeasts.The biodiversity of both LAB and yeasts in sourdoughs mainly depends on their type and production process [25,26].De Vuyst et al. [27] provide the most recent comprehensive compilation of lactobacilli in sourdoughs (more than 700) used for bread making.

Classification of Sourdoughs
According to the preparation process and the metabolic activity of the main LAB and yeasts found in sourdoughs, these doughs can be classified into three or four types [28].Type I sourdough is a mixture of flour and water subjected to incubation at temperatures below 30 • C, inoculated with a previously fermented sourdough, to keep the microorganisms' metabolic activity high.Type I sourdoughs traditionally use the fermented dough as the single leavening agent [28].Examples include the sourdoughs used for sourdough bread, panettone, and San Francisco rye bread.Type I sourdoughs are generally firm doughs with a pH range between 3.8 and 4.5 and are fermented in a temperature range of 20 to 30 In type II sourdough, the mixture of flour and water is incubated at temperatures above 30 • C for extended fermentation times (up to 5 days).In Type II sourdoughs, Saccharomyces cerevisiae is added to leaven the dough; Lactobacillus species are the dominant members of type II sourdoughs.This sourdough can be liquid and is widely used as an acidifying agent and aroma generator in bakery products.This process was adopted by the industry, in part, due to the simplification of the multiple-step build typical of type I sourdoughs [30].
Type III sourdough is a dough that goes through a drying process, generally by spraying, known as "ready-to-use" [28].Microorganisms in an active state are first transformed to a dormant state by drying and then require reactivation and resuspension with water before being used as a bread ingredient.Type III sourdough is used most for large-scale baking products as a premix starter due to its easy handling and long shelf life.However, it is important to mention that the number of viable microorganisms in this dough depends on the drying conditions [25,26,31].Drying-resistant LAB, such as Pediococcus pentosaceus, Lb. plantarum, and Lb.brevis predominate in this sourdough.The drying conditions, time, and temperature depend on the desired characteristics and caramelization of the baked product.Pediococcus pentosaceus SP2 (isolated from kefir grains) was evaluated by Plessas et al. [32] as the starter in sourdough bread making when applied in fresh, freeze-dried, and freeze-dried immobilized (on wheat bran).The freeze-dried immobilized cells led to higher total titratable acidity values, and the bread presented higher resistance to mold and rope spoilage.Finally, type IV ferment (mixed ferment) was more recently characterized and consists of a mixture of types I and II [28].

Lactic Acid Bacteria in Sourdoughs
Sourdough microbiota generally comprise two types of microorganisms: LAB and yeasts.Table 1 shows the common LAB and yeasts isolated from different types of sourdoughs.The populations of the microorganisms can range from 10 6 to 10 9 CFU/g, and the proportion of LAB to yeasts is approximately 100:1; in consequence, the most significant contribution to the production of compounds during the fermentation of sourdoughs is provided by LAB [3,31,32].LAB from the genera Enterococcus, Weissella, Lactococcus, Leuconostoc, Pediococcus, and Lactobacillus have been identified in sourdoughs, Lactobacillus being the most important since more than 60 species of this genus have been isolated.Despite the vast diversity of LAB identified, only some species of lactobacilli survive the maintenance process and feeding cycles to which sourdoughs are subjected [33].In general, obligate heterofermentative LAB survive these processes mainly due to their ability to metabolize carbohydrates, assimilate various amino acids, and resist the acidic environment caused by the production of acids.The species Lb. plantarum and Lb.sanfrancisencis have been identified as the LAB with the highest presence and resistance in sourdoughs.It has been reported that around 50% of fermented sourdoughs present this type of LAB growth during the different feeding cycles [20].
The feeding cycles of sourdoughs, which consist of adding new flour and water to the dough once the fermentation time has been completed, significantly affect the doughs' microbiota diversity.Different investigations have demonstrated that, with feeding cycles increasing, the diversity of sourdough microbiota decreases significantly; Lb. plantarum and Lb.sanfrancisencis predominate from the tenth day of feeding since they present the greatest ability for adaptation and resistance to this process [20,[34][35][36].Meanwhile, species such as Lb.brevis, Lb. paralimentarius, Lb. fermentum, and Lb.casei have a low capacity to adapt to the environment after several cycles of sourdough feeding [20,25,36].Plessas [32] remarked that many factors are important for sourdough preparations; however, the starter culture selection is considered the most critical.Arena et al. [37] investigated the diversity of LAB and yeasts in six spontaneous sourdough fermentations from Italy (Apulian region); the authors reported the identification of Lb. plantarum as the dominant LAB and S. cerevisiae as the main yeast in the analyzed samples.Developments in the last decade have changed the selection criteria used to select sourdough cultures and have expanded the tools used to assess the metabolic potential of specific strains, species, or genera [6].

Metabolic Pathways of LAB during Sourdough Fermentation
The metabolic pathways or processes during lactic acid fermentation depend directly on the type of LAB [6] in the sourdough (Figure 1).Microorganisms' metabolic activity in sourdough affects the dough's technological functionality and the bread's nutritional properties, aroma profile, shelf life, and overall quality [19].From glucose, homofermentative LAB mainly produce lactic acid/lactate (>90%) through glycolysis (homolactic fermentation), while heterofermentative LAB, in addition to lactic acid (around 50%) production, generate CO 2 , acetic acid, or other acids, and ethanol through the 6phosphoglucanate/phosphoketolase (6-PG/PC) pathway.Hexoses other than glucose enter the main glycolytic pathways at glucose-6-phosphate or fructose-6-phosphate, after the isomerization process and/or phosphorylation [3].
In the presence of electron acceptors, obligate heterofermentative LAB reduce fructose to mannitol and produce acetate and ATP, while mannitol can be fermented to lactate.The use of pentoses is not restricted to the LAB type and their type fermentation; facultative heterofermentative LAB, which ferment hexoses through glycolysis, ferment pentoses in the same way as obligate heterofermentative species, with the help of the enzyme fructose-1-6 diphosphate aldose.Pentose fermentation produces equimolar amounts of lactic and acetic acid without the formation of CO 2 [3].
LAB from sourdoughs have proteases, peptidases, and amino acid-converting enzymes, which break down cereal proteins, generating both compounds with bioactive properties and compounds that directly impact the bread's flavor [5].The LAB proteolytic enzymatic system consists of an extracellularly located serine proteinase, specific transport systems for di/tripeptides and oligopeptides (>3 amino acid residues), and a variety of intracellular peptidases [3,38].The transformation of amino acids by LAB into compounds that improve the bread's flavor and aroma is produced by transamination reactions, which require the presence of both an electron acceptor and an amino group, preferably α-ketoglutarate.
reactions, which require the presence of both an electron acceptor and an amino g preferably α-ketoglutarate.
Protein metabolism in lactic acid bacteria can involve the breakdown of variou amino acids, specifically, catabolic reactions involving phenylalanine, where LAB bolic enzymes generate bioactive compounds such as phenylactate and phenylac (Figure 2) [3,38].

Sourdough and Its Effect on Bread Properties
It has been observed that the fermentation processes of sourdoughs play a signifi role in improving the bread's organoleptic attributes.The compounds formed in acid fermentations and their concentration are directly related to improving bread' vors and textures [3,39,40].Protein metabolism in lactic acid bacteria can involve the breakdown of various free amino acids, specifically, catabolic reactions involving phenylalanine, where LAB catabolic enzymes generate bioactive compounds such as phenylactate and phenylacetate (Figure 2) [3,38].
Figure 2 provides an overview of the protein metabolism by LAB during sourdough fermentation.LAB hydrolyze proteins from the dough into amino acids and perform various metabolic reactions, including decarboxylation and deamination, which produce organic acids (such as lactic acid) and biogenic amines.These metabolic processes lead to the acidification of the sourdough and the development of characteristic flavors.The sourdough used in bread fermentation is crucial in leavening and flavor development [39].

Sourdough and Its Effect on Bread Properties
It has been observed that the fermentation processes of sourdoughs play a significant role in improving the bread's organoleptic attributes.The compounds formed in lactic acid fermentations and their concentration are directly related to improving bread's flavors and textures [3,39,40].

Texture
In general, sourdough fermentation affects dough rheology on two levels: in the sourdough itself and in dough that has had sourdough added to it [3].Various studies have assessed the effect of sourdough on bread formulation and the resulting bread texture.In general, the addition of sourdoughs decreases the resistance to extension, reduces the elasticity of the dough, and increases the degree of softening, obtaining a crumb with greater CO2 retention capacity (a greater number of alveoli of a larger size), a firmer crust, and

Texture
In general, sourdough fermentation affects dough rheology on two levels: in the sourdough itself and in dough that has had sourdough added to it [3].Various studies have assessed the effect of sourdough on bread formulation and the resulting bread texture.In general, the addition of sourdoughs decreases the resistance to extension, reduces the elasticity of the dough, and increases the degree of softening, obtaining a crumb with greater CO 2 retention capacity (a greater number of alveoli of a larger size), a firmer crust, and better texture.These changes, both in the structure of the crumb and in the dough, have been attributed by several authors to the proteolysis of gluten proteins caused primarily by the increase in acidity due to the production of organic acids during the sourdough fermentation [8,40,[42][43][44][45][46][47].
Many LAB associated with sourdoughs, especially the Lactobacillus genus, produce a variety of exopolysaccharides (EPS) and oligosaccharides during the lactic acid fermentation of sourdoughs.These compounds have been shown to have the ability to function as hydrocolloids and thus improve the texture properties of bread during storage [3].Torrieri et al. [46] recorded that adding EPS-producing LAB to sourdough significantly improved the texture during bread storage, further to gluten proteolysis.Besides, they observed that bread had a higher moisture content and better mechanical properties during storage, which may be an alternative to reducing the aging process of bread [46].

Flavor
Sourdough bread has a complex flavor profile influenced by the compounds generated during microbial fermentation [48][49][50].Odors and flavors are generated in the sourdough mainly due to enzymatic and microbial processes during the fermentation of these doughs.These compounds belong to different chemical classes: aldehydes, acids, alcohols, ketones, esters, and pyrazines.The flavors and odors from the raw materials are insignificant compared to those generated during fermentation.However, other aromatic and flavor compounds are produced from lipid oxidation processes and Maillard reactions [39].
Two categories of flavor (taste and aroma) compounds are produced during the lactic acid fermentation of sourdoughs.The first of them includes non-volatile compounds, such as organic acids (lactic, acetic, phenyllactic, and phenylacetic acid) that LAB produce and volatile compounds that include alcohols, aldehydes, ketones, esters, and sulfur compounds, which are formed during the fermentation and baking stages of bread [3,39].The generation of aromatic and flavor compounds is closely related to LAB type, temperature, pH, and moisture content since they directly influence the metabolic activity of the microorganisms [3].Mantzourani et al. [50] reported that sourdough (fermented with Lb. paracasei K5) improved bread's sensory properties related to the detected aroma volatiles and consumer preference.Warburton et al. [48] characterized the volatile organic compounds in the crumb of 12 sourdough breads.The authors attributed the observed differences to the culture identified if the fermentation activity was dominated by yeast or the different classes of LAB.The authors associated the profiles in three clusters.In Cluster 1, volatile organic compounds (ethanol, 3-methyl-1-butanol, phenylethanol, 2-methyl-1-propanol, acetaldehyde, and 2,3-butanedione) predominated, along with increased production of lactic acid, indicating the activity of yeast and homofermentative or facultative heterofermentative LAB.Cluster 2 was associated with acetic acid, acetate esters, and acidity, revealing that obligate heterofermentative LAB predominates.Fermentation of lipids (production of aldehydes and lactones) related to yeast fermentation activity was classified in Cluster 3.
Siepmann et al. [40] observed sourdoughs made with Lb brevis and Lb.plantarum and with the fermentation temperature changed from 28 to 35 • C.They reported the formation of furfurals, compounds associated with almond aromas in pieces of bread incorporating doughs fermented at 35 • C. On the other hand, pieces of bread made with sourdough fermented at 28 • C did not show the formation of these compounds, demonstrating that the fermentation temperature in the sourdough is important for forming volatile compounds that affect the bread's flavor.Also, they verified that the type of LAB significantly influences the generation of final flavors in the bread.These researchers observed that sourdoughs fermented with Lb. reuteri, Lb. plantarum, and Lb.amylovorus generated a greater diversity of flavor and aroma compounds in the bread than in loaves of bread to which sourdoughs fermented with Lb. brevis and Lb.plantarum had been added.Xu et al. [51] investigated the influence of different starter cultures (Kazachstania humilis, S. cerevisiae, Wickerhamomyces anomalus, Lb. sanfranciscensis DSM20451T and Lactobacillus sakei LS8, in various combinations) on sourdough wheat bread volatiles.Using headspace solid-phase microextraction and gas chromatography/mass spectrometry analysis (SPME-GC/MS) established that sourdough bread fermented with a combination of lactobacilli and yeast had a more complex profile of volatiles (particularly concerning esters).Finally, Siepmann et al. [40] determined that both the starter culture and the sourdough fermentation temperature significantly affect the final aroma and flavor of bread.
Various investigations have reported different sensory analyses to determine the acceptability of breads formulated with sourdoughs.Sourdough fermentation is known for its ability to produce complex and desirable flavors [32].Understanding sourdough fermentation's microbial dynamics and metabolic pathways [52] can lead to new and unique bread flavors and aromatic profiles.Adding 20 to 30% sourdough to the bread formulation has increased the acceptability of both texture and flavor compared with bread made exclusively with yeast.These results demonstrate that the aromatic compounds formed during the fermentation process of sourdoughs help to improve the sensory acceptability of bakery products [8,13,45,50,53].

Sourdough Effect on the Bread's Shelf Life
It has been shown that LAB produce bioactive compounds with antimicrobial capacity during the lactic acid fermentation process.Substances such as short-chain fatty acids, peptides, diacetyl, hydrogen peroxide, and organic acids have been reported with the potential for inhibiting both pathogenic bacteria and important molds in foods [26,[54][55][56][57][58][59][60].Some of these compounds have been identified and isolated from sourdoughs, which are also related to the antifungal activity in bread when sourdough is used as an ingredient.In addition, an increase in the bread's shelf life was observed.
Various investigations have been carried out on the ability of sourdough, as an additive, to extend the useful life of bakery products [14,15,26,37,50,61,62].Table 2 presents some examples of these investigations carried out in recent years.According to Table 2, adding 20% (w/w) of sourdough to the bread formulation increases the product's shelf life by an average of six days.Garofalo et al. [10] observed that the addition of 30% (w/w) sourdough in the bread formulation considerably increased the shelf life of the product, achieving improvements of 19 to 21 days (Table 2).The type of LAB used for the fermentation of the sourdough affects the effectiveness in increasing the shelf life of the bread.Hernández-Figueroa et al. [15] reported that bread incorporating sourdoughs fermented with Lactobacillus (Lactobacillus acidophilus NRRL B-4495 or L. casei 21/1) had a longer shelf life compared to a traditional one.The addition of a poolish-type sourdough (fermented with Lactiplantibacillus plantarum NRRL B-4496) inhibited fungal growth in bread for ten days [14].As can be seen in Table 2, the sourdoughs fermented with Lb. amylovorus, Lb rossiae, and a mixture of Lb. rossiae and Lb.paralimentarius presented a significant ability to increase the final product's shelf life.
In addition to mold growth, bread aging (retrogradation of starch and loss of moisture) is one of the determining factors for the shelf life of baking products.Torrieri et al. [46] observed that adding 30% sourdough fermented with exopolysaccharide-forming LAB to the bread formulation reduced the moisture loss rate and affected starch retrogradation kinetics.The results were attributed mainly to the production of organic acids, bacterial hydrolysis (by LAB) of starch molecules, and proteolysis of gluten subunits.Finally, they concluded that adding sourdoughs to the bread formulation could be an alternative to produce bakery products free of synthetic additives.* In comparison to control bread that did not have any antimicrobial agent added to it.

Properties of Sourdough Bioactive Compound Extracts
LAB are distinguished by their ability to produce lactic acid, among other interesting metabolites with antimicrobial activity [60,65].Also, cell-free supernatants' antibacterial and antifungal activity in vitro has been widely studied, particularly in members of the Lactobacillus genus.Studies of extracts obtained from sourdoughs focus on analyzing the bioactive capacity presented mainly by peptides and organic acids formed from the fermentation of LAB in the doughs [26].These extracts and their different fractions are obtained using the Osborne method (1907) and a physical separation employing centrifugation.The compounds' separation and identification in these extracts are commonly obtained through different instrumental techniques based on chromatography [11,15,52,57].
For peptide compounds, the peptidases of the lactic acid bacteria in the sourdough participate in secondary proteolysis (Figure 2), releasing free amino acids, which in turn are subjected to various catabolic reactions by the same microorganisms.These peptide compounds, which generally increase during lactic acid fermentation, are native protein fragments with amino acid sequences that can have a positive impact as antimicrobial, antioxidant, and anti-inflammatory agents [21,54,[66][67][68].
Similarly, the organic acids identified and isolated in the sourdough extracts show bioactive properties, mainly functioning as antimicrobial agents against important pathogenic bacteria in food and mold spoilage of bakery products.This property means that the extracts can be used as "natural" preservatives in the development of products free of synthetic additives or clean labels, and in this way, satisfy consumers' demand [14,15,53,55,58,69].

Bioactive Compounds from Sourdoughs with Antimicrobial Properties
As mentioned previously, organic acids and peptide compounds formed during the lactic acid fermentation of sourdoughs have been shown to have antimicrobial activity when used as pure compounds, individually or together, in in vitro studies [53,56,59,63,70].For this reason, various investigations assess the antifungal capacity of the pure compounds and the aqueous extracts obtained from sourdoughs against the bread's spoilage molds [14,15].Table 3 presents the bioactive compounds in the sourdough extracts that have shown antifungal activity in vitro against selected molds.
According to Table 3, organic acids such as lactic, acetic, and phenyllactic acids are identified in sourdoughs fermented with Lb. reuteri, Lb. plantarum, and Lb.brevis presented significant antifungal activity, reducing the radial growth of molds such as Fusarium graminearum and A. niger.When these sourdoughs were incorporated into the bread formulation, they increased the product's shelf life by seven days compared with control bread [64].Similarly, Axel et al. [71] and Luz et al. [11] identified another type of organic acid, which inhibited the growth of molds on the surface of the bread and, in this way, increased the shelf life of the product (Table 3).Fraberger et al. [62] isolated (Lactiplantibacillus plantarum S4.2 and Lentilactobacillus parabuchneri S2.9) from sourdoughs and reported antifungal growth suppression against F. graminearum MUCL43764, Aspergillus fumigatus, A. flavus MUCL11945, A. brasiliensis DSM1988, and P. roqueforti DSM1079.They also found antibacterial activity against Bacillus cereus DSM31, B. licheniformis DSM13, B. subtilis LMG7135, and B. subtilis S15.20.Hernández-Figueroa et al. [14] found that aqueous extracts from poolish-type sourdoughs fermented with Lactiplantibacillus plantarum NRRL B-4496 presented fungistatic capacity primarily attributed to lactic and acetic acids and probably antifungal peptides.Hernández-Figueroa et al. [15] identified lactic acid in concentrations between 1 and 2% in sourdoughs fermented with L. acidophilus, while in those fermented by L. casei, lactic acid (1-2%) and acetic acid (0.1-0.2%) were identified.
Table 3 shows some peptide compounds identified in sourdoughs fermented with LAB.These compounds have shown the ability to inhibit the growth of molds such as P. roqueforti, Aspergillus parasiticus, P. polonicum, and P. paneum in slices of bread.Likewise, the ability of peptide compounds to increase the shelf life of bakery products by up to seven days has been demonstrated [14,54,57].
In addition, bacteriocins produced by LAB have been isolated from the aqueous extracts of sourdoughs.One of the bacteriocins identified in these doughs is reutericycline produced by Lb. reuteri LTH2584.This compound has significant antibacterial capacity against Listeria monocytogenes, B. subtilis, and Staphylococcus aureus [73].Other bacteriocins isolated from aqueous extracts of sourdoughs with antibacterial capacity are bavaricin A produced by Lb. bavaricus MI401, plantaricin ST31 produced by Lb. plantarum, and BLIS C57 produced by Lb. sanfranciscensis C57.Although these bacteriocins have not been approved as antimicrobials or antifungals, studying their antimicrobial capacity and food safety may generate alternatives that can replace synthetic antimicrobials as preservatives [3].

Effects of Bioactive Compounds on the Shelf Life of Bread
Despite the existence of various investigations on the in vitro antifungal capacity of bioactive compounds formed by LAB isolated from sourdoughs, tested in specific culture broths at the laboratory level, studies of this capacity, obtained from sourdough extracts and incorporated into bread for the study of the shelf life of the product, are scarce [14,15,53,55,58,69,74].
Coda et al. [72] demonstrated the addition of aqueous extracts of fermented sourdoughs with Lb. plantarum caused an increase in mold appearance time in bread inoculated with P. roqueforti and without inoculation, of 7 and 21 days, respectively.In another study, Rizello et al. [57] evaluated the addition of 4% (w/w) of a lyophilized extract from sourdough fermented with Lb. rossiaie and Lb.plantarum to the bread formulation, finding that it inhibited mold growth for around 28 days of storage at room temperature.These researchers concluded that the antifungal activity of the extract was due to the synergistic and complex activity between organic acids and peptide compounds formed during lactic acid fermentation.These results open the possibility of obtaining antifungal compounds from fermented sourdough extracts, which can be made with accessible and lower-cost raw materials and be used as natural antimicrobials in a wide range of food products.

Other Possible Beneficial Effects
Functional LAB as starter cultures used in sourdough fermentation have been researched for years [32,75].Acidification caused by LAB in sourdough is a central aspect in activating several cereal enzymes and synthesizing microbial active metabolites, positively influencing the derived products' nutritional/functional and health-promoting benefits [1].Akamine et al. [5] stated that proper management of sourdough fermentation allows gluten levels to be adjusted, delaying starch digestibility and increasing the bioaccessibility of vitamins and minerals.The nutritional quality and the biotransformation observed in the flour during the fermentation process impact the nutrients' bioaccessibility, and the beneficial effects from this process influence the final product and consumers' health [76,77].Perri et al. [78] reported that fermentation of sprouted barley with selected lactic acid bacteria enhanced its nutritional features through increases in free amino acids (35%) and γ-aminobutyric acid concentrations (57%), and a decrease in phytic acid content.The fermentation process strongly mitigated the adverse effects (on dough rheology and baking) related to sprouted barley flour enzymatic activities.
Utilizing food production byproducts back into food production within a circular food economy is one of the driving methods used to improve sustainability within the food industry [7,23,79].Sourdough production can benefit from various byproducts from other industries if they are safe and suitable for consumption.Some examples are fruit and vegetable peelings (apple skins or potato peels) [80], spent grains, yeast slurry byproducts from breweries [5], coffee grounds, and leftovers from grinding nuts and seeds [81].
According to D'Amico et al. [82], certain cereal components, particularly in wheat and rye, which are known to trigger adverse reactions in a small subset of the population, may undergo partial modification or degradation during sourdough fermentation.This process can potentially decrease their harmful effects, depending upon the composition of sourdough microbiota, processing conditions, and the resultant acidification.Nissen et al. [83] investigated the impact of gluten-free bread enriched with spirulina on the colon microbiota of non-celiac volunteers.These authors indicate that sourdough fermentation and algae enrichment can mitigate the negative effect of gluten-free bread on the gut microbiota of non-celiac consumers.To make gluten-free sourdough bread, a gluten-free starter can be produced by fermenting flours, such as brown rice, white rice, sorghum, tapioca, or a blend of gluten-free flours to replace wheat flour [84][85][86], and proper selection of the starter is important [87,88].Gluten-free doughs lack the structure provided by gluten; thus, many formulations incorporate hydrocolloids (such as xanthan gum) to improve the texture and elasticity.These ingredients trap gas produced during fermentation and improve the overall structure of the bread.
Foods with high daily intake, such as cereal-based products, are good vehicles for supplementary probiotics [4,5].The addition of probiotics is challenging for cereal-based baked foods due to the high temperatures applied during baking [5].The use of microencapsulated probiotics, edible films containing probiotics, spore-forming probiotics, and probiotic addition after baking are the main strategies used to formulate probiotic cereal-based baked products [89].Spore-forming probiotics represent the best approach to supplementing foods before baking [89].Da Ros et al. [90] studied industrially made baker's yeast and sourdough bread, which were used to feed for two weeks the gastrointestinal simulator Twin Mucosal-SHIME, and found that only the consumption of sourdough bread has the potential to enhance the synthesis of short-chain fatty acids and free amino acids at the colon level.Akamine et al. [5] identified probiotics (Bacillus licheniformis) isolated from sourdough and analyzed hydrolases that presented the capacity to hydrolyze gluten from Limosilactobacillus fermentum, Pediococcus pentosaceus, and Saccharomyces cerevisiae.
Müller et al. [75] evaluated Leuconostoc citreum DCM65 and Lactiplantibacillus plantarum subsp.plantarum MA418 selected for producing mannitol, exopolysaccharides, and amylolytic and antifungal activity) and their potential as single or co-culture starters in sourdough fermented buns containing different levels of sugar (control 9% and reduced 0, 3, 6%).The authors reported that sourdough buns started with Lactiplantibacillus plantarum had higher volume, larger slice area, and softer crumb; this was in contrast to buns fermented with Leuconostoc citreum.Both starter cultures showed high potential for producing sourdough buns with reduced sugar content.
Sourdough fermentation can partially break down gluten proteins, potentially making bread more tolerable for individuals with mild gluten sensitivities.Also, it can increase the bioavailability of certain nutrients in grains, making them more digestible and nutritious.For example, phytic acid reduction through fermentation can improve mineral absorption.The study of sourdough can lead to healthier and more nutritious bread products.

Final Remarks
Although it remains a traditional process, adding sourdoughs to the formulations of bakery products represents an effective option to enhance the properties of those products.These improvements are primarily attributed to the compounds formed during the sourdough lactic acid fermentation, such as organic acids, peptide compounds, and/or exopolysaccharides.
This review highlights sourdough's significant role in enhancing bread's quality and shelf life.Decades of research has consistently demonstrated that incorporating sourdough into bread formulations positively influences their organoleptic attributes, imparting distinct and desirable flavors and textures.The bioactive compounds generated through sourdough fermentation, especially by lactic acid bacteria, possess antimicrobial properties that play a crucial role in prolonging bread shelf life.These compounds effectively combat spoilage, particularly inhibiting the growth of molds.Selected studies about using extracts from sourdoughs containing these bioactives to extend the shelf life of bread have shown favorable results.This emphasizes the importance of continuing to study and integrate sourdough or its extracts into bread production processes as it can improve the quality and diversity of products and align with broader objectives of improving sustainability.

Figure 1 .
Figure 1.Products generated by homofermentative and heterofermentative lactic acid ba (LAB) metabolic routes during sourdough fermentation.

Figure 2
Figure2provides an overview of the protein metabolism by LAB during sourd fermentation.LAB hydrolyze proteins from the dough into amino acids and perform ious metabolic reactions, including decarboxylation and deamination, which produc ganic acids (such as lactic acid) and biogenic amines.These metabolic processes le the acidification of the sourdough and the development of characteristic flavors.The dough used in bread fermentation is crucial in leavening and flavor development[3

Table 1 .
Main lactic acid bacteria (LAB) and yeasts isolated from different types of sourdoughs.

Table 2 .
Results of improvement in the shelf life of bread with the addition of sourdoughs fermented by different types of lactic acid bacteria (LAB).

Table 3 .
Bioactive compounds identified in sourdoughs fermented with different lactic acid bacteria (LAB) and their effect against bread's spoilage molds.