Lactic Acid Bacteria-Derived Exopolysaccharides: Dual Roles as Functional Ingredients and Fermentation Agents in Food Applications

Abstract

Exopolysaccharides (EPSs) produced by lactic acid bacteria (LAB) have received special attention as valuable products due to their potential applications as techno-functional and bioactive ingredients in foods. EPS production and consumption are an age-old practice in humans, as evidenced by fermented foods. Over the last two decades, extensive research has examined, analyzed, and reported a wide variety of EPSs from several LAB strains, as well as their techno-functional properties in foods. Also, research efforts focused on EPS characterization and yield production have been carried out. In food applications, EPS quantification and characterization in situ (direct fermentation) took place in various matrices (dairy, bread, plant-based fermented, and meat products). EPS direct application (ex situ) has been less investigated despite its better structural–functional control and use in non-fermented foods. Fewer EPS investigations have been conducted related to health benefits in humans and their mechanisms of action. The composition and functionality of EPSs vary depending on the LAB strain and food matrix used to produce them; thus, various challenges should be addressed before industrial applications are performed. This review aims to compile and summarize the recent findings on EPSs produced by LAB, highlighting their yield, culture production, techno-functional role in foods, food applications, and health benefits in clinical trials. It examines their dual applications, whether as purified functional ingredients (ex situ) or as fermentation products (in situ), and critically assesses both technological and bioactive implications. Also, it explores production challenges, regulatory considerations, and future perspectives for sustainable and tailored applications of EPSs in food innovation.

1. Introduction

The demand for clean-label, functional, and health-promoting foods has driven a growing interest in naturally derived biopolymers, particularly exopolysaccharides (EPSs) synthesized by lactic acid bacteria (LAB). EPSs are high-molecular-weight polysaccharides secreted extracellularly by many LAB strains during fermentation. These microbial polysaccharides have received increasing attention due to their dual role in food systems: they enhance sensory and physicochemical properties while also exhibiting bioactive potential that may contribute to consumer health [1,2,3]. LAB-derived EPSs can be categorized into homopolysaccharides (homo-EPSs), composed of a single type of monosaccharide (dextran, levan), and heteropolysaccharides (hetero-EPSs), which consist of various sugar monomers arranged in complex branched structures [4,5]. The latter are most commonly produced by LAB strains used in fermented food systems, where they contribute to improved viscosity, texture, water-holding capacity, and overall mouthfeel, especially in low-fat and plant-based formulations [6,7]. These techno-functional properties have made EPSs valuable in the development of dairy products, bakery products, gluten-free baked goods, and novel emulsified or fermented plant-based foods as safe-food additives and natural functional food ingredients (soluble fiber and prebiotics) with economic importance because of the possibility of replacing or reducing hydrocolloids [2,8,9].

Beyond their technological utility, EPSs from LAB also display a range of biofunctional activities. Several studies in in vitro and animal models have demonstrated their immunomodulatory, antioxidant, cholesterol-lowering, and prebiotic effects, linking their use to potential benefits in gut health and metabolic wellness [10,11,12,13]. Given that many EPS-producing LAB strains are also considered probiotics or are used in starter cultures for food fermentation, their integration into foods can simultaneously deliver both structural improvements and microbiota-modulating effects.

Two primary routes are currently explored for integrating EPSs into food systems: (1) the direct addition (ex situ) of purified EPS as food additives or natural hydrocolloids (less studied); and (2) in situ production through fermentation using EPS-producing LAB strains. Purified EPSs could be commercialized as a current hydrocolloid (e.g., xanthan gum), as their easy handling and commercial distribution allow for more precise dosing and standardization. The latter approach aligns particularly well with clean-label and consumer-preferred fermentation-based innovations [11]. Also, in situ fermentation leverages the natural biosynthesis process, minimizing purification costs and often enhancing both texture and microbial stability in the final product [3,14], as occurs with EPS-producing starter cultures for yogurt, fermented milk, fermented cream, and fermented cereal beverages. Despite their promise, challenges remain in the broader application of LAB-derived EPSs, including variability in EPS yield, structural heterogeneity, scale-up limitations, and regulatory constraints. However, advancements in microbial biotechnology, fermentation optimization, and omics-driven strain selection are highlighting the way for expanded use of EPSs in both conventional and functional food systems.

This review aims to provide an overview of EPSs produced by LAB, highlighting their yield, culture production, technological and functional roles in foods, food applications, and health benefits in clinical trials. It examines their dual applications, whether as purified functional ingredients (ex situ) or as fermentation products (in situ), and critically assesses both technological and bioactive implications. The article also explores production challenges, regulatory considerations, and future perspectives for sustainable and tailored applications of EPSs in food innovation.

2. Exopolysaccharide-Producing Lactic Acid Bacteria

LAB are a phylogenetically diverse group of Gram-positive, acid-tolerant, and generally non-sporulating microorganisms that play a crucial role in various fermented food systems. LAB produce lactic acid from the central carbon metabolism and accumulate it in their surroundings, at concentrations of up to around 1% during sugar fermentation [15,16]. Among their many beneficial traits, one of the most functionally significant is the ability of certain strains to synthesize EPS-extracellular carbohydrate polymers of biological origin that participate in the formation of microbiological aggregates or slimy coating around the cells [3,17].

EPSs are key components of the cell wall of bacteria that protect against harsh and harmful environmental factors (dehydration, extreme temperatures, acidity, osmotic stress, antimicrobials) and aid bacteria in biofilm formation and attachment, as well as their strain–host interaction capability [3,4]. For instance, deletion of three eps gene clusters (responsible for capsular polysaccharide synthesis, mediated polysaccharide chain elongation, and EPS biosynthesis) in Lactiplantibacillus plantarum CSK thinned the cell wall, lowered the biofilm formation capacity, diminished the auto-aggregation capability at early growth stages (<9 h), reduced cell surface hydrophobicity, lessened the electron acceptor and donor capacities, and decreased bacteria stability in solution (changes in zeta potential value which assess the surface electrical charge characteristics associated with bacteria stability in suspension) in mutants, reducing strain stability under adverse conditions such as gastric and intestinal environments [18].

2.1. Genera and Species Involved in EPS Production

EPSs produced by LAB are structurally diverse carbohydrate polymers secreted into the extracellular environment or associated with the bacterial cell surface. These molecules can be loosely bound (slime EPS) or tightly attached as capsular EPSs, and their biosynthesis is a metabolically regulated process that reflects the species, strain, and environmental conditions [19]. EPS production is not uniformly distributed across all LAB; rather, it is a strain-specific trait found across multiple genera. Notably, EPS-producing strains have been identified in the genera Lactobacillus (including various reclassified genera), Streptococcus, Leuconostoc, Weissella, Pediococcus, and Enterococcus [4,5,20]. Common species widely distributed and involved in fermented foods are EPS producers such as Limosilactobacillus, Enterococcus, Lactococcus, and Streptococcus in yogurt, where the EPS production levels ranged from 28 to 62 μg/mL [21]; Leuconostoc mesenteroides and Leuconostoc kimchii from pulque [22]; and Lactobacillus brevis and Pediococcus damnosus from oat beverages [23]. Some of the most extensively studied EPS-producing strains include Streptococcus thermophilus, which is widely used in yogurt and cheese production and is known for producing capsular EPSs that contribute to viscosity and reduce syneresis. Lactobacillus delbrueckii subsp. bulgaricus is often co-cultured with S. thermophilus in yogurt fermentation, enhancing texture through synergistic EPS synthesis. Lacticaseibacillus rhamnosus and Lacticaseibacillus casei produce EPSs with reported immunomodulatory and antioxidant properties. Ln. mesenteroides is well-known for dextran production in traditional sourdough and fermented vegetables. Weissella cibaria and Weissella confusa are of emerging interest due to their EPSs with unique branching patterns and promising prebiotic activities [2,8,9]. Several LAB strains have been reported as EPS producers; probably, each fermented food or microbial ecosystem (plants, animals, gastrointestinal tract, etc.) is a potential source. Therefore, LAB strains from all over the world have demonstrated EPS production and are expected to be identified in the future. The identification and knowledge of EPS and their LAB strains could provide inexhaustible sources of natural food additives.

2.2. EPS Types and Biosynthesis

The EPS produced by LAB can be divided into two groups: homo-EPS and hetero-EPS. The primary difference between homo-EPS and hetero-EPS is the enzymes involved in their synthesis. Homo-EPS synthesizes outside of the cell and requires specific substrates (sucrose, glucose, etc.) that form the polymer by enzymatic action. By contrast, the hetero-EPS forms repeating precursors intracellularly and then are transported across the membrane for their extracellular polymerization [3]. Homo-EPSs are composed of a single type of monosaccharide (glucose or fructose), usually produced extracellularly from sucrose by specific enzymes such as β-glucansucrase or β-fructansucrase (these enzymes cleave sucrose and polymerize the glucose or fructose moieties). Hetero-EPSs are composed of repeating units of two or more different monosaccharides (glucose, galactose, rhamnose, mannose), often more complex in structure and biosynthesis [4,5]. These are typically synthesized intracellularly via membrane-associated pathways involving glycosyltransferases and exported across the cell wall [4,5,24]. Homo-EPS can be grouped into four types based on linkage bonds and nature of monomeric units, such as α-D-glucans (dextran, alternan, mutan, and reuteran); β-D-glucans; fructans (levans, inulin-type); and others (polygalactan) [24]. Table 1 shows the main glycosidic linkages in homo-EPSs and selected examples of hetero-EPSs, the monomeric units, and their branching. Weissella, Leuconostoc, and some Lactobacillus strains commonly produce these EPS types.

Table 1. Glycosidic linkages, monomeric units, and branching of selected exopolysaccharides (EPSs).

EPS	Linkage	Monomer Units	Branching	Charge	Reference
Dextran	α(1→6); α(1→3)	Glucose	Branched	Neutral	[13]
Reuteran	α(1→6); α(1→4)
Mutan	α(1→3)		Linear
Alternan	α(1→6); α(1→3)
β-D-glucan	α(1→3)
Levan	β(2→6); β(2→1)	Fructose	Branched	Neutral
Kefiran		Glucose, galactose	Branched	Neutral
BC-25 EPS from Lactobacillus plantarum BC-25	(1→2)-linked Man, (1→2,6)-linked Glc, (2→6)-linked Man, and (2→6)-linked Gal	Mannose, galactose, glucose	Branched	ND	[25]
LgEPS from Lactobacillus gasseri FR4	1,6 linked-α-D-Glcp; 1,4 linked-α-D-Galp, 1,3,4 linked-α-D-Manp, 1,3 linked-α-L-Rhap, 1,4 linked-α-L-Fucp, 1,4 linked-β-D-Glcp, and β-D-Galp-1	Glucose, galactose, mannose, rhamnose, fucose	Branched	ND	[26]
R-17-EPS from Lactobacillus pentosus LZ-R-17	→2)-α-D-Galp-(1→4)-β-D-Glcp-(1→4)-β-D-Glcp-(1→4)-β-D-Glcp-(1→	Galactose, glucose	Linear	ND	[27]
LPE-1 EPS from L. plantarum AR307	Backbone: 1,4-β-D-Glcp, 1,4-α-D-Glcp, and 1,4,6-β-D-Galp; branched 1,6-β-D-Galp	Glucose, galactose	Branched	ND	[28]
EPS-T1 from L. plantarum T1	1,4-linked Glcp and 1,6-linked Galp	Glucose, galactose	Branched	ND	[29]

ND: Not determined.

Table 1. Glycosidic linkages, monomeric units, and branching of selected exopolysaccharides (EPSs).

EPS	Linkage	Monomer Units	Branching	Charge	Reference
Dextran	α(1→6); α(1→3)	Glucose	Branched	Neutral	[13]
Reuteran	α(1→6); α(1→4)
Mutan	α(1→3)		Linear
Alternan	α(1→6); α(1→3)
β-D-glucan	α(1→3)
Levan	β(2→6); β(2→1)	Fructose	Branched	Neutral
Kefiran		Glucose, galactose	Branched	Neutral
BC-25 EPS from Lactobacillus plantarum BC-25	(1→2)-linked Man, (1→2,6)-linked Glc, (2→6)-linked Man, and (2→6)-linked Gal	Mannose, galactose, glucose	Branched	ND	[25]
LgEPS from Lactobacillus gasseri FR4	1,6 linked-α-D-Glcp; 1,4 linked-α-D-Galp, 1,3,4 linked-α-D-Manp, 1,3 linked-α-L-Rhap, 1,4 linked-α-L-Fucp, 1,4 linked-β-D-Glcp, and β-D-Galp-1	Glucose, galactose, mannose, rhamnose, fucose	Branched	ND	[26]
R-17-EPS from Lactobacillus pentosus LZ-R-17	→2)-α-D-Galp-(1→4)-β-D-Glcp-(1→4)-β-D-Glcp-(1→4)-β-D-Glcp-(1→	Galactose, glucose	Linear	ND	[27]
LPE-1 EPS from L. plantarum AR307	Backbone: 1,4-β-D-Glcp, 1,4-α-D-Glcp, and 1,4,6-β-D-Galp; branched 1,6-β-D-Galp	Glucose, galactose	Branched	ND	[28]
EPS-T1 from L. plantarum T1	1,4-linked Glcp and 1,6-linked Galp	Glucose, galactose	Branched	ND	[29]

ND: Not determined.

Hetero-EPSs display complex repeating units consisting of 3–8 monosaccharides, including the following: (a) neutral sugars (glucose, galactose, mannose); (b) deoxy sugars (rhamnose, fucose); (c) acidic sugars (glucuronic acid); and/or (d) amino sugars (N-acetylglucosamine). Some hetero-EPSs may also contain non-carbohydrate substituents, such as acetyl, phosphate, or pyruvate groups, which influence solubility, charge, and biological interactions [4,5]. The production of hetero-EPS is more energy-intensive and occurs via the Wzx/Wzy-dependent pathway. It involves intracellular synthesis of sugar nucleotide precursors (e.g., UDP-glucose, dTDP-rhamnose). The assembly of repeating units is carried out on a lipid carrier (undecaprenyl phosphate) by glycosyltransferases. Then, the translocation across the cytoplasmic membrane is mediated by the Wzx flippase. Finally, polymerization is carried out by Wzy polymerase and exported via membrane transport proteins (Figure 1). This pathway is encoded by eps operons; genes located at the 5′ end of the cluster are involved in the modulation and construction machinery of EPS biosynthesis and exhibit the highest level of overall conservation. Their structure can vary among LAB strains, but typically includes gene clusters for regulation (epsA), initiation (epsB/C), polymerization and transport (epsD–G), and glycosyltransferases (epsH–J, a variable region, which includes the polymerase wzy, the flippase Wzx, one or more glucosyltransferases, and/or other polymer-modifying genes) [19]. In W. cibaria, the analysis of biological subsystems in the genome recorded five genes encoding for the EPS biosynthesis (epsB, epsC, epsD, GLT2, and epsE with functional roles of manganese-dependent protein-tyrosine phosphatase, tyrosine-protein kinase transmembrane modulator, tyrosine-protein kinase, glycosyl transferase, and undecaprenyl-phosphate galactosephosphotransferase, respectively) [30]. Meanwhile, the metabolic map of the genome identified genes encoding for GTF (α-D-glucan production) and nucleotide sugar biosynthesis [30]. In Lc. lactis, the epsR gene was the first to be highly expressed after 3 h of culture and decayed after 6 h, and epsBDK was expressed after 6 h. After 21 h of growth, the key genes responsible for EPS biosynthesis (epsB, epsD, epsR, epsK) declined in their expression, probably due to nutrient depletion [31]. Detailed descriptions of EPS biosynthesis can be found in Zeidan et al. [19], Zhang et al. [32], Zhang et al. [33], Zang et al. [34], and Zhang et al. [35].

Figure 1 outlines the general metabolic pathway leading from sugar uptake to nucleotide sugar precursors, the action of EPS biosynthetic enzymes, and the secretion of polymers. EPS synthesis in LAB is encoded by large, variable gene clusters, commonly referred to as eps or cps operons. These clusters typically contain 15–30 genes arranged in modules responsible for precursor biosynthesis, polymer assembly, chain length determination, and export. The pathway begins with carbohydrate uptake (glucose, sucrose, lactose, fructose), followed by conversion into nucleotide sugar precursors such as UDP-glucose, UDP-galactose, dTDP-rhamnose, and GDP-mannose. These activated sugars form the building blocks for EPS assembly. Glycosyltransferases sequentially transfer nucleotide sugars onto a lipid carrier embedded in the cytoplasmic membrane, generating repeat units. Polymerization is then catalyzed by a Wzy-dependent system, in which the Wzy polymerase elongates the chain and the Wzz protein regulates chain length distribution. Export across the membrane involves dedicated proteins such as EpsB, EpsC, and EpsD, as well as flippase-like proteins that translocate lipid-linked intermediates. Finally, the fully synthesized polymers are secreted into the extracellular space as homopolysaccharides (such as dextran, β-glucan, and levan) or heteropolysaccharides with diverse monosaccharide compositions.

2.3. EPS Yield, Culture Conditions, and Composition

The production of EPS is highly strain-dependent, with significant variability in yield, composition, molecular weight, and branching even within the same species [13,14,36]. The expression of eps genes and the activity of enzymes involved in EPS synthesis are regulated by carbon source availability (sucrose, glucose, lactose), pH, temperature, presence of metal ions (Ca²⁺, Mg²⁺), oxygen levels, and redox status, and strain adaptation to stress (osmotic or acid stress). In general, sucrose often leads to homo-EPS production, whereas glucose, lactose, or galactose may stimulate hetero-EPS synthesis. Fermentation conditions, including temperature, pH, and oxygen availability, can modulate EPS yield and molecular characteristics. For selected strains, acidic stress on cells could inhibit bacterial growth but stimulate EPS production [37]. Regarding the growth phase, EPS biosynthesis may be growth-associated or occur predominantly in the stationary phase, depending on the strain. For most LAB, static or anaerobic conditions increase EPS production; however, for specific strains, aerobic conditions favor EPS synthesis [38].

2.3.1. Effect of Carbon Sources

EPSs are produced at exponential and stationary growth phases (~30 h) when bacteria are metabolically most active; subsequently, and in some cases, they hydrolyze with fermentation time. An overproduction of EPS occurs when abundant sugar is present, thus providing carbon and energy [3]. In general, sugars stimulate EPS production in most bacteria in a dose-dependent manner. However, each sugar can reach a saturation point that promotes the production of other metabolites and is competitive with EPS formation. Therefore, the supplementation of sugars into the culture medium to produce EPS should be evaluated carefully. Based on the vast number of studies on EPS production by several LAB, it is relatively easy to find a range for sugar testing to increase or optimize EPS production. Despite sucrose improving or promoting the EPS production of most LAB, selected strains do not utilize it as a carbon source for growth or EPS production. For instance, L. sanfranciscensis Ls-1001 isolated from sourdough fermented maltose rather than glucose, sucrose, fructose, and galactose [39].

In various studies, selected factors have been investigated to increase EPS yield production. Adesulu-Dahunsi et al. [40] evaluated the effect of carbon source, nitrogen source, pH, and fermentation time on dextran yield production from W. confusa OF126. The best carbon source (20 g/L) was sucrose, followed by galactose, glucose, and lactose. In contrast, yeast extract (2.5%) proved to be a more effective nitrogen source than tryptone, beef extract, and peptone. Various combinations of sucrose concentration (16–24 g/L, carbon source), yeast extract (2–2.5%, nitrogen source), pH (6.5–8.0), and cultivation time (45–48.13 h) reached EPS yields of 3 g/L. The best EPS yield (3 g/L) was obtained with 16 g/L of sucrose, 2.5% yeast extract, pH 6.50, and 45 h of cultivation [40]. For L. sanfranciscensis Ls-1001, the best carbon source was maltose (25 g/L), the nitrogen source was yeast peptone (10.24 g/L), and fresh yeast extract (12.9 mL/L) produced 249.3 mg/L, 30.99% more EPS than the unoptimized fermented medium [39]. In another study, the optimal level of sucrose (91.2 g/L), pH (5.8), and sodium acetate (NaAc, 1.2 g/L) increased the EPS (homo-EPS-L2) yield by 7.6-fold from Leuconostoc lactis L2 cultured in a defined broth, reaching 58.1 g/L EPS [41].

2.3.2. Stress Factors’ Impact

Stress factors could enhance EPS production to increase LAB’s resistance to stress [4]. In de Man, Rogosa, and Sharpe (MRS) broth, the addition of H₂O₂ (3 mM) and/or CaCl₂ (10 mM) increased the EPS production drastically (9.5-fold) from Lactobacillus rhamnosus ZY, with the combination being most effective [42]. The effect of H₂O₂ and CaCl₂ on the cells showed an over-expression of heat shock protein, energy metabolism proteins, and mitigation of H₂O₂ cytotoxicity [42]. An amount of 20 mg/L of CaCl₂ favored the hetero-EPS of L. plantarum K25 in semi-defined medium up to 238.6 mg/L after 24 h of fermentation at 37 °C compared with non-CaCl₂ (140 mg/L) and with amounts >20 mg/L [43]. The viability of L. plantarum and the pH were not modified by CaCl₂ addition. Ca²⁺ changes the glycometabolism of L. plantarum, increasing the expression level of β-glucosidase/β-galactosidase, phosphate transferase system IIA, and mannosidase. Long-chain fatty acid synthesis, membrane transporters, amino acid, and UMP biosynthesis were also upregulated when Ca²⁺ was supplemented [43].

2.3.3. Co-Culture Effects on EPS Production

Co-culture of LAB with Saccharomyces cerevisiae enhances EPS production through contact with components on the yeast cell surface [4]. L. rhamnosus ATCC 9595 and RW-9595M increased by 49% (224 mg/L) and 42% (568 mg/L), respectively, the EPS production in co-culture with yeast, compared with monoculture after 48 h in whey permeate medium [44]. In depth, the genetic analysis revealed that co-culture significantly influenced the sugar metabolism (over-expressing genes during the first 12 h of fermentation), the EPS operons (over-expressing), amino acid biosynthesis and lipid metabolism, stress and cellular response, and other functions (microbial metabolism in diverse environments) [44]. The co-cultivation of LAB and yeast impacts metabolism and increases EPS production in a strain-dependent manner.

2.3.4. Temperature Effect on EPS Production

Temperature is another key factor in EPS production; the ex situ EPS production allows control and adjustment of the optimal temperature. Adesulu-Dahunsi et al. [40] investigated the effect of temperature (20, 25, 30, 37, and 40 °C), pH (6, 6.5, 4.5, 8), and cultivation time (12, 24, 36, 48, 60, 72, 84, and 96 h) on EPS yield from W. confusa OF12. The best conditions to increase EPS yield production (~2.1 g/L) were pH = 7.0, 30 °C, and 48 h. Three levels of pH (4.8, 5.6, and 6.2), sucrose (2, 4, and 6%), temperature (22, 30, and 37 °C), and fermentation time (12, 24, and 36 h) were evaluated and optimized to allow maximum hetero-EPS production for W. cibaria 27 in MRS broth [45]. The best EPS yield (25.6 g/L) was obtained at pH of 6.2, 22 °C, 6% sucrose, and 24 h. In addition, great EPS productivity (17.8 mg/L/h) and the increasing expression fold of drs gene (dextransucrase, DSRA) were observed by 2.5- and 2.7-fold at 40 g/L and 60 g/L sucrose, respectively [45]. In another study, 20 °C was the best temperature for EPS production of W. cibaria CH2 in MRS broth supplemented with sucrose (5%), resulting in an 81.3% increase [30]. The optimal EPS production happens below the optimal growth temperature of the bacteria [30,46]. Pediococcus acidilactici M76 enhanced its EPS production up to 1.62 g/L at pH 4.0 (tested range 4–9), 25 °C (25–30 °C), after 3 days (3–15 days) in black raspberry beverage with 30 °Brix (from 20, 30, or 40 °Brix), and longer fermentation times degraded the EPS produced [47]. The quantity of EPS obtained from the beverage is sufficient to provide potential health benefits, as indicated by the in vitro analysis. In fermented milk, EPS production by Lactobacillus paracasei H9 was improved at 40 °C, an 8-h fermentation time, and a 14% inoculum size, reaching a value of 991.5 mg/L [48]. In situ EPS synthesis requires testing at food manufacturing or production temperatures, such as refrigeration (2–15 °C). EPS production by two LAB (L. plantarum 162 R and Ln. mesenteroides N6) was evaluated in sucuk (Turkish-type fermented sausage) at three temperatures (14, 16, and 18 °C) and three times (8, 12, and 16 days) [49]. Increasing temperature and time enhanced EPS production between 43 and 51% for both bacteria and their mixture at the same ripening time (16 days) for the three temperatures. Likewise, time also favored the EPS production when sucuk was maintained at the same temperature. An interesting finding was that Ln. mesenteorides N6 formed more EPS alone than mixed with L. plantarum. In another study, Hilbig et al. [50] investigated the in situ EPS production from four LAB (homo-EPS L. curvatus 1.624 and L. sakei 1.411, and hetero-EPS by L. plantarum 1.1478 and 1.25) in pork meat for cooked ham (supplementation of brine with 0.5% sucrose or dextrose improves EPS production) during tumbling at two temperatures (2 and 15 °C). They found that hetero-EPS-forming strain produces significantly (p < 0.05) higher amounts than homo-EPS-producing strains. The optimization of EPS production has been extensively investigated due to its functional and health importance in food and culture media. Therefore, various studies have tested the effect of the key factors previously mentioned in a wide variety of LAB species. As previously mentioned, the EPS amounts obtained at tested conditions are widely variable, from marginal (0.08–1.21 g/L) [51] to great yields (97.5 g/L) [52].

According to Table 2, the screening, initial production, characterization, quantification, and optimization of EPS from LAB producers are frequently conducted in MRS broth. MRS broth, replaced with a carbon source (glucose by another sugar, such as sucrose, maltose, fructose, or lactose), or supplemented with sucrose (2–20%), has been investigated to identify EPS and optimize its production in liquid media (Table 2). Minimum liquid medium and brain heart infusion were also used as LAB medium for EPS production. An interesting approach for EPS production is the utilization of food by-products, such as fruit pomaces, bagasse, and cereal residues, which are rich in carbon, nitrogen, and minerals. Most studies have utilized a liquid medium over a solid one for EPS production; however, only one study used MRS agar as the culture medium. The authors stated some advantages of EPS production on solid medium, such as easy handling and extraction, since no culture medium is mixed. On the other hand, for EPS applications in foods (in situ), commonly due to its techno-functional or health properties, EPS production, identification, and quantification are analyzed directly in the food matrix. As Table 2 shows, whey, milk, sausage (sucuk), legume dough/suspension, wheat dough, and juice beverage (black raspberry) have been used in EPS production and quantification. The EPS yields varied widely as the strain and culture medium were varied; the range is from 2.5 to 97,500 mg/L or kg. The better EPS producers are L. acidophilus, L. reuteri, L. gasseri, L. plantarum, L. kefiranofaciens, W. confusa, W. cibaria, P. acidilactici, Ln. pseudomesenteroides, and Ln. citreum with levels >1000 mg/L (Table 2).

Table 2. Lactic acid bacteria production of exopolysaccharides (EPS), culture conditions, and yield.

Lactic Acid Bacteria	EPS Type	Culture Conditions	Yield	Reference
Lactobacillus rhamnosus RW-9595M	Hetero-EPS (glucose, galactose, and rhamnose)	Supplemented whey permeate (5% (w/w) whey permeate, MgSO4·7H2O, MnSO4·H2O, Tween 80, corn steep liquor, and yeast extract) at 37 °C for 7 h and 200 rpm	2350 mg/L	[53]
Weissella cibaria WC4 and Lactobacillus plantarum PL9	Glucan (homo EPS constituted of glucose)	MRS broth supplemented with 292 mM sucrose incubated at 30 °C for 1 day Wheat flour (312 g), water (137.5 mL), sucrose (50 g), and cellular suspension (50 mL) incubated at 30 °C for 24 h	3.88 and 3.14 mg/mL, respectively, in MRS broth W. cibaria produces 2500 mg/kg glucan in dough	[54]
Streptococcus thermophilus NIZO0131, NIZO2104, and Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842, NCIMB 702074, and DGCC 291	Hetero-EPS NIZO0131: galactose/rhamnose NIZO2104: galactose/ribose/N-acetyl-galactosamine ATCC 11842: galactose/glucose NCIMB 702074: galactose/glucose DGCC 291: galactose/glucose	Reconstituted skim milk at 12% incubated at 37 °C for 16 h For S. thermophilus strains, a subcultivation at 42 °C for 6 h was applied	NIZO0131: 78 mg/L NIZO2104: 45 mg/L ATCC 11842: 60 mg/L NCIMB 702074: 36 mg/L DGCC 291: 38 mg/L	[55]
W. cibaria MG1	Dextran (homo-EPS constituted of glucose)	MRS broth added with sucrose (10%), incubated at 30 °C for 72 h Wort (9% final extract content) added with 5 or 10% sucrose, incubated at 30 °C for 72 h	36,400 mg powder/L in MRS+Suc, 8600 mg powder/L in wort + 5% Suc, and 14,400 mg powder/L in wort with 10% Suc	[56]
L. plantarum 162 R, Leuconostoc mesenteroides N6, and the mixture	Not reported	Sucuk sausage (beef meat, tail fat, salt, garlic, red pepper, powdered black pepper, cumin, allspice) incubated at 18 °C for 12 days	9.79, 18.60, and 17.56 mg/kg dry matter for L. plantarum, Ln. mesenteroides, and the mixture, respectively. Sucuk fermented spontaneously did not detect EPS	[49]
Lactobacillus kefiranofaciens DN1	Hetero-EPS DN1 (rhamnose, arabinose, galactose, glucose, and mannose)	MRS broth (20 g/L glucose) and supplemented with glucose (40, 60, or 80 g/L)	1380 mg powder/L in MRS broth and 2260 mg/L for MRS + 60 g/L glucose	[57]
L. rhamnosus ZY	Hetero-EPS ZY (fructose, galactose, glucose, fucose, rhamnose, and mannose)	MRS broth supplemented with H2H2 or/and CaCl2 at 37 °C under anaerobic conditions	342.8 mg powder/L in MRS broth at 24 h, 567 mg powder/L in MRS broth + 3 mM H2O2 after 24 h, 2203.5 mg powder/L in MRS broth + 10 mM CaCl2 after 12 h, and 2498.5 mg powder/L in MRS broth + 3 mM H2O2 + 10 mM CaCl2 and 12 h	[42]
L. plantarum BR2	Hetero-EPS (glucose and mannose)	EPS production medium (yeast extract 4, lactose 4, Tween 80 0.1, sodium acetate 0.5, and ammonium sulfate 0.5 g/100 mL), incubate at 37 °C for 72 h	2800 mg powder/L	[58]
L. plantarum NR 104573.1 and Pediococcus pentosaceus NR 042058.1 from wheat bran sourdough	----	MRS broth supplemented with 10% glucose at 37 °C for 24 h	408 and 263 mg/L	[59]
Weissella confusa OF126	Dextran (homo-EPS constituted of glucose)	MRS broth + 10 g/L sucrose incubated at 30 °C for 24 h and 170 rpm	2000 mg/L	[40]
Leuconostoc citreum B-2	Highly branched dextran (homo-EPS constituted of glucose)	MRS broth with 75 g/L sucrose incubated at 30 °C for 48 h and 80 rpm/min	28,300 mg powder/L	[24]
Lactobacillus gasseri FR4	Hetero LgEPS (glucose, mannose, galactose, rhamnose, and fucose)	MRS broth (glucose substituted by sucrose) added with 2% sucrose	7200 mg powder/L	[26]
W. confusa PP29	Dextran (homo-EPS constituted of glucose)	MRS I: MRS broth added with fructose (40 g/L) and glucose (40 g/L) MRS II: MRS broth plus 80 g/L sucrose MRS III: MRS broth and 80 g/L sucrose dissolved in UHT milk Culture media were incubated at 33 °C for 48 h under agitation at 100 rpm	2800 mg powder/L MRS I, 5180 mg powder/L MRS II, and 17,400 mg powder/L MRS III	[60]
W. cibaria SJ14	Hetero-EPS (mannose, glucose, galactose, arabinose, xylose, and rhamnose)	Semi-defined medium (MRS modified) incubated at 30 °C for 34 h	331.47 mg/L	[61]
Lactobacillus sanfranciscensis Ls-1001	Glucan (homo EPS constituted of glucose)	MRS broth, carbon source replaced by maltose, incubated at 30 °C for 24 h	190.3 mg/L	[39]
Fructilactobacillus sanfranciscensis Ls5	Hetero-EPS (glucose and mannose)	MRS broth, carbon source replaced by maltose, incubated at 30 °C for 24 h	202.3 mg/L	[62]
L. rhamnosus EM1107, Lactobacillus mucosae CNPC007, L. plantarum CNPC003	For L. plantarum: Hetero-EPS (mannose, glucose, and galactose), the composition was similar despite the carbon source	MRS broth or MRS broth carbon source replaced by fructooligosaccharide (FOS, Orafti®), incubated at 37 °C for 24 h	In MRS broth, EPS production was 167.6, 153.2, and 378 mg/L, respectively. In MRS containing FOS, EPS was 356.8, 345.7, and 568.4 mg/L, respectively	[63]
Leuconostoc pseudomesenteroides JF17	Dextran (homo EPS constituted of glucose)	MRS broth added with 18% sucrose, pH 7.3 at 20 °C for 48 h	53,770 mg/L	[64]
Ln. pseudomesenteroides DSM 20193 and W. confusa Ck15	Dextran (homo EPS constituted of glucose)	Chickpea flour (28 g), sucrose (2 g), water (70 mL), incubated at 30 °C for 24 h	1.18% and 1.49%, respectively	[65]
W. confusa C19	Dextran	MRS agar and cereal (rice, oat, wheat, and maize) extract (ratio cereal and water 1:10) in proportion 1:1 was added with sucrose (5%), incubated at 37 °C for 3 days	21,900 mg/L, 20,900 mg/L, 19,100 mg/L, 18,500 mg/L for rice, wheat, maize, and oat medium, respectively	[66]
Lactobacillus reuteri E81	Glucan (homo EPS constituted of glucose)	Wheat dough yield of 200 added with 15% sucrose, incubated for 24 h	15,200 mg/kgdry sourdough	[67]
Lactobacillus paracasei H9	Hetero-EPS (mannose, glucose, galactose)	Milk incubated for 8 h, at 37 °C, and inoculum size 14%	932 mg/L	[48]
Lactobacillus fermentum S1	Hetero-EPS S1 (glucose, galactose, mannose, arabinose)	Liquid medium (glucose (20 g), ammonium citrate (5 g), soya peptone (10 g), yeast extract (6 g), MnSO4 (0.05 g), FeSO4 (0.04 g), MgSO4 (0.2 g), and Tween 80 (1 mL)) incubated at 33 °C for 24 h	668 mg/L	[68]
Lactobacillus pentosus LZ-R-17	Hetero-R-17-EPS (galactose and glucose)	Milk incubated at 37 °C for 24 h	185.2 mg/L	[27]
P. acidilactici M76	----	Black raspberry beverage (30 °Brix) incubated at 25–30 °C for 3–15 days	1620 mg/L (at 25 °C and 3 days)	[47]
W. confusa QS813	Dextran (homo EPS constituted of glucose)	Red bean dough yield of 250 added with 10% sucrose, incubated at 30 °C for 24 h	18,680 mg/kgsourdough	[69]
W. confusa XG-3	Dextran (homo EPS constituted of glucose)	Optimized medium (sucrose 80.1 g/L, beef extract 8 g/L, casein peptone 5 g/L, yeast extract 10 g/L, sodium acetate 3.7 g/L, ammonium citrate 3 g/L, K2HPO4 4 g/L, and Tween 80 2 mL/L adjusted to pH 5.8), incubated at 30 °C for 72 h and 120 rpm	97,500 mg powder/L	[52]
Lactobacillus curvatus SJTUF 62116	Hetero-EPS 1 (glucose and mannose)	MRS broth cultivated at 30 °C for 24 h	283.5 mg powder/L,	[70]
Lactiplantibacillus plantarum T1, CL80, CSK, S-1A	Hetero-EPS T1, -EPS CL80, -EPS CSK, and EPS S-1A (mannose, rhamnose, glucose, and galactose)	Inoculate in milk at 108 CFU/mL and incubated at 37 °C	385 mg powder/L, 336 mg powder/L, 157 mg powder/L, and 98 mg powder/L	[71]
Lpb. plantarum T1	Hetero-EPS T1 (glucose and galactose)	MRS broth at 37 °C for 30 h	249 mg powder/L	[29]
Enterococcus sp. BE11	Hetero-EPS BE11 (L-rhamnopyranose, D-arabinose, D-galactopyranose, D-glucuronic acid, D-glucopyranose)	MRS broth supplemented with 1% sucrose at 37 °C for 48 h	173 mg powder/L	[72]
Lactococcus lactis subsp. diacetylactis RBL 37	----	Modified MRS broth replaced carbon source with 20% sucrose, the cells were grown until DO600 0.5–2.0	274.3 mg/L	[73]
Lactobacillus acidophilus LAC-1	----	Whey and whey supplemented with 2% lactose incubated for 48–72 h under anaerobic conditions	2172 and 2168 mgdry EPS/L	[38]
Lpb. plantarum ITD-ZM-101 and Lc. lactis ITD-ZM-106	----	Brain heart infusion broth containing 15 g/L of dried agave bagasse or agave leaves, incubated at 37 °C, 120 rpm for 120 h	147.2 and 130 mg/L for agave leaves	[74]

Table 2. Lactic acid bacteria production of exopolysaccharides (EPS), culture conditions, and yield.

Lactic Acid Bacteria	EPS Type	Culture Conditions	Yield	Reference
Lactobacillus rhamnosus RW-9595M	Hetero-EPS (glucose, galactose, and rhamnose)	Supplemented whey permeate (5% (w/w) whey permeate, MgSO4·7H2O, MnSO4·H2O, Tween 80, corn steep liquor, and yeast extract) at 37 °C for 7 h and 200 rpm	2350 mg/L	[53]
Weissella cibaria WC4 and Lactobacillus plantarum PL9	Glucan (homo EPS constituted of glucose)	MRS broth supplemented with 292 mM sucrose incubated at 30 °C for 1 day Wheat flour (312 g), water (137.5 mL), sucrose (50 g), and cellular suspension (50 mL) incubated at 30 °C for 24 h	3.88 and 3.14 mg/mL, respectively, in MRS broth W. cibaria produces 2500 mg/kg glucan in dough	[54]
Streptococcus thermophilus NIZO0131, NIZO2104, and Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842, NCIMB 702074, and DGCC 291	Hetero-EPS NIZO0131: galactose/rhamnose NIZO2104: galactose/ribose/N-acetyl-galactosamine ATCC 11842: galactose/glucose NCIMB 702074: galactose/glucose DGCC 291: galactose/glucose	Reconstituted skim milk at 12% incubated at 37 °C for 16 h For S. thermophilus strains, a subcultivation at 42 °C for 6 h was applied	NIZO0131: 78 mg/L NIZO2104: 45 mg/L ATCC 11842: 60 mg/L NCIMB 702074: 36 mg/L DGCC 291: 38 mg/L	[55]
W. cibaria MG1	Dextran (homo-EPS constituted of glucose)	MRS broth added with sucrose (10%), incubated at 30 °C for 72 h Wort (9% final extract content) added with 5 or 10% sucrose, incubated at 30 °C for 72 h	36,400 mg powder/L in MRS+Suc, 8600 mg powder/L in wort + 5% Suc, and 14,400 mg powder/L in wort with 10% Suc	[56]
L. plantarum 162 R, Leuconostoc mesenteroides N6, and the mixture	Not reported	Sucuk sausage (beef meat, tail fat, salt, garlic, red pepper, powdered black pepper, cumin, allspice) incubated at 18 °C for 12 days	9.79, 18.60, and 17.56 mg/kg dry matter for L. plantarum, Ln. mesenteroides, and the mixture, respectively. Sucuk fermented spontaneously did not detect EPS	[49]
Lactobacillus kefiranofaciens DN1	Hetero-EPS DN1 (rhamnose, arabinose, galactose, glucose, and mannose)	MRS broth (20 g/L glucose) and supplemented with glucose (40, 60, or 80 g/L)	1380 mg powder/L in MRS broth and 2260 mg/L for MRS + 60 g/L glucose	[57]
L. rhamnosus ZY	Hetero-EPS ZY (fructose, galactose, glucose, fucose, rhamnose, and mannose)	MRS broth supplemented with H2H2 or/and CaCl2 at 37 °C under anaerobic conditions	342.8 mg powder/L in MRS broth at 24 h, 567 mg powder/L in MRS broth + 3 mM H2O2 after 24 h, 2203.5 mg powder/L in MRS broth + 10 mM CaCl2 after 12 h, and 2498.5 mg powder/L in MRS broth + 3 mM H2O2 + 10 mM CaCl2 and 12 h	[42]
L. plantarum BR2	Hetero-EPS (glucose and mannose)	EPS production medium (yeast extract 4, lactose 4, Tween 80 0.1, sodium acetate 0.5, and ammonium sulfate 0.5 g/100 mL), incubate at 37 °C for 72 h	2800 mg powder/L	[58]
L. plantarum NR 104573.1 and Pediococcus pentosaceus NR 042058.1 from wheat bran sourdough	----	MRS broth supplemented with 10% glucose at 37 °C for 24 h	408 and 263 mg/L	[59]
Weissella confusa OF126	Dextran (homo-EPS constituted of glucose)	MRS broth + 10 g/L sucrose incubated at 30 °C for 24 h and 170 rpm	2000 mg/L	[40]
Leuconostoc citreum B-2	Highly branched dextran (homo-EPS constituted of glucose)	MRS broth with 75 g/L sucrose incubated at 30 °C for 48 h and 80 rpm/min	28,300 mg powder/L	[24]
Lactobacillus gasseri FR4	Hetero LgEPS (glucose, mannose, galactose, rhamnose, and fucose)	MRS broth (glucose substituted by sucrose) added with 2% sucrose	7200 mg powder/L	[26]
W. confusa PP29	Dextran (homo-EPS constituted of glucose)	MRS I: MRS broth added with fructose (40 g/L) and glucose (40 g/L) MRS II: MRS broth plus 80 g/L sucrose MRS III: MRS broth and 80 g/L sucrose dissolved in UHT milk Culture media were incubated at 33 °C for 48 h under agitation at 100 rpm	2800 mg powder/L MRS I, 5180 mg powder/L MRS II, and 17,400 mg powder/L MRS III	[60]
W. cibaria SJ14	Hetero-EPS (mannose, glucose, galactose, arabinose, xylose, and rhamnose)	Semi-defined medium (MRS modified) incubated at 30 °C for 34 h	331.47 mg/L	[61]
Lactobacillus sanfranciscensis Ls-1001	Glucan (homo EPS constituted of glucose)	MRS broth, carbon source replaced by maltose, incubated at 30 °C for 24 h	190.3 mg/L	[39]
Fructilactobacillus sanfranciscensis Ls5	Hetero-EPS (glucose and mannose)	MRS broth, carbon source replaced by maltose, incubated at 30 °C for 24 h	202.3 mg/L	[62]
L. rhamnosus EM1107, Lactobacillus mucosae CNPC007, L. plantarum CNPC003	For L. plantarum: Hetero-EPS (mannose, glucose, and galactose), the composition was similar despite the carbon source	MRS broth or MRS broth carbon source replaced by fructooligosaccharide (FOS, Orafti®), incubated at 37 °C for 24 h	In MRS broth, EPS production was 167.6, 153.2, and 378 mg/L, respectively. In MRS containing FOS, EPS was 356.8, 345.7, and 568.4 mg/L, respectively	[63]
Leuconostoc pseudomesenteroides JF17	Dextran (homo EPS constituted of glucose)	MRS broth added with 18% sucrose, pH 7.3 at 20 °C for 48 h	53,770 mg/L	[64]
Ln. pseudomesenteroides DSM 20193 and W. confusa Ck15	Dextran (homo EPS constituted of glucose)	Chickpea flour (28 g), sucrose (2 g), water (70 mL), incubated at 30 °C for 24 h	1.18% and 1.49%, respectively	[65]
W. confusa C19	Dextran	MRS agar and cereal (rice, oat, wheat, and maize) extract (ratio cereal and water 1:10) in proportion 1:1 was added with sucrose (5%), incubated at 37 °C for 3 days	21,900 mg/L, 20,900 mg/L, 19,100 mg/L, 18,500 mg/L for rice, wheat, maize, and oat medium, respectively	[66]
Lactobacillus reuteri E81	Glucan (homo EPS constituted of glucose)	Wheat dough yield of 200 added with 15% sucrose, incubated for 24 h	15,200 mg/kgdry sourdough	[67]
Lactobacillus paracasei H9	Hetero-EPS (mannose, glucose, galactose)	Milk incubated for 8 h, at 37 °C, and inoculum size 14%	932 mg/L	[48]
Lactobacillus fermentum S1	Hetero-EPS S1 (glucose, galactose, mannose, arabinose)	Liquid medium (glucose (20 g), ammonium citrate (5 g), soya peptone (10 g), yeast extract (6 g), MnSO4 (0.05 g), FeSO4 (0.04 g), MgSO4 (0.2 g), and Tween 80 (1 mL)) incubated at 33 °C for 24 h	668 mg/L	[68]
Lactobacillus pentosus LZ-R-17	Hetero-R-17-EPS (galactose and glucose)	Milk incubated at 37 °C for 24 h	185.2 mg/L	[27]
P. acidilactici M76	----	Black raspberry beverage (30 °Brix) incubated at 25–30 °C for 3–15 days	1620 mg/L (at 25 °C and 3 days)	[47]
W. confusa QS813	Dextran (homo EPS constituted of glucose)	Red bean dough yield of 250 added with 10% sucrose, incubated at 30 °C for 24 h	18,680 mg/kgsourdough	[69]
W. confusa XG-3	Dextran (homo EPS constituted of glucose)	Optimized medium (sucrose 80.1 g/L, beef extract 8 g/L, casein peptone 5 g/L, yeast extract 10 g/L, sodium acetate 3.7 g/L, ammonium citrate 3 g/L, K2HPO4 4 g/L, and Tween 80 2 mL/L adjusted to pH 5.8), incubated at 30 °C for 72 h and 120 rpm	97,500 mg powder/L	[52]
Lactobacillus curvatus SJTUF 62116	Hetero-EPS 1 (glucose and mannose)	MRS broth cultivated at 30 °C for 24 h	283.5 mg powder/L,	[70]
Lactiplantibacillus plantarum T1, CL80, CSK, S-1A	Hetero-EPS T1, -EPS CL80, -EPS CSK, and EPS S-1A (mannose, rhamnose, glucose, and galactose)	Inoculate in milk at 108 CFU/mL and incubated at 37 °C	385 mg powder/L, 336 mg powder/L, 157 mg powder/L, and 98 mg powder/L	[71]
Lpb. plantarum T1	Hetero-EPS T1 (glucose and galactose)	MRS broth at 37 °C for 30 h	249 mg powder/L	[29]
Enterococcus sp. BE11	Hetero-EPS BE11 (L-rhamnopyranose, D-arabinose, D-galactopyranose, D-glucuronic acid, D-glucopyranose)	MRS broth supplemented with 1% sucrose at 37 °C for 48 h	173 mg powder/L	[72]
Lactococcus lactis subsp. diacetylactis RBL 37	----	Modified MRS broth replaced carbon source with 20% sucrose, the cells were grown until DO600 0.5–2.0	274.3 mg/L	[73]
Lactobacillus acidophilus LAC-1	----	Whey and whey supplemented with 2% lactose incubated for 48–72 h under anaerobic conditions	2172 and 2168 mgdry EPS/L	[38]
Lpb. plantarum ITD-ZM-101 and Lc. lactis ITD-ZM-106	----	Brain heart infusion broth containing 15 g/L of dried agave bagasse or agave leaves, incubated at 37 °C, 120 rpm for 120 h	147.2 and 130 mg/L for agave leaves	[74]

3. Techno-Functional Properties of LAB Exopolysaccharides in Food Systems

EPSs synthesized by LAB contribute significantly to the techno-functional properties of a wide range of fermented and formulated food products. These polymers improve texture, enhance mouthfeel, increase water retention, and stabilize emulsions, all of which are essential attributes in modern food design, especially in low-fat, plant-based, and gluten-free matrices [1,2,8,9]. The diverse molecular structures of EPS—ranging in size, branching, and monomer composition—result in unique interactions with food biopolymers such as proteins and starches, which in turn modulate their rheological behavior and stability under processing conditions [4,6,7,14]. EPSs impart desirable rheological changes in foods, such as increased viscosity, reduced syneresis, improved texture, and emulsifying, thickening, and stabilizing properties. EPSs can range from 10 kDa to over 1000 kDa, with hetero-EPS generally exhibiting lower molecular weights than homo-EPS. Molecular weight influences viscosity, water-holding capacity, resistance to enzymatic breakdown, and interactions with proteins and lipids in food matrices. Branching patterns, especially in dextrans and hetero-EPS, can significantly influence the three-dimensional conformation of the polymer and its structural–functional properties. For instance, highly branched dextrans may form compact or coiled structures, contributing to high viscosity at low concentrations. Linear EPSs (e.g., levan) may form extended chains, influencing gelation and film-forming abilities.

Table 3 presents selected EPSs produced by LAB and their techno-functional properties in foods. According to Table 3, most structural–functional properties of foods include enhanced texture, reduced viscosity, reduced syneresis, improved water retention, and increased stability during storage or temperature changes, as well as gel formation in foods such as cereal and legume doughs/breads, cereal beverages, soymilk, fermented milk, and meat products.

Table 3. Techno-functional properties of exopolysaccharides (EPSs) in foods produced by lactic acid bacteria.

Lactic Acid Bacteria	EPS Type	Food Product	Techno-Functional Properties	Quality Improvement	Reference
Lactobacillus sanfranciscensis TMW 1.392	Levan (homo-EPS)	Dough/bread	Water absorption, bread volume, and crumb firmness	Bread texture and quality	[75]
Streptococcus thermophilus NIZO2104 and Lactobacillus delbrueckii subsp. bulgaricus DGCC 291	Hetero-EPS NIZO2104: galactose/ribose/N-acetyl-galactosamine DGCC 291: galactose/glucose	Fermented milk	Gel formation, viscosity, and water-holding capacity	Firmness, apparent viscosity, and reduced syneresis	[55]
Lactobacillus curvatus TMW 1.624	Dextran (homo-EPS)	Gluten-free bread	Increase viscosity, gas retention, and water-holding capacity	Bread texture and volume Shelf life by retarding bread staling	[76]
Weissella cibaria MG1	Dextran (homo-EPS)	Malt fermented beverage	Viscosity improver	Beverage stability Desirable body	[56]
Lactobacillus plantarum 162 R, Leuconostoc mesenteroides N6, and the mixture.	Not reported	Sucuk (Turkish-type fermented sausage)	Gel formation and retention water	Texture	[49]
Leuconostoc citreum B-2 Leuconostoc pseudomesenteriodes JF17	Highly branched dextran (homo-EPS) Dextran (homo-EPS)	----	Water-holding capacity	Binding and stabilizing agent of water	[24,64]
Lactobacillus reuteri E81	Glucan (homo EPS constituted of glucose)	Dough/wheat bread	Dough viscoelasticity and retention water	Hardness of fresh bread	[67]
Leuconostoc lactis L2	EPS-L2 (homo EPS constituted of glucose)	Fermented milk	Gel formation, viscosity	Gel stability, texture	[41]
Weissella confusa QS813	Dextran (homo EPS constituted of glucose)	Red bean sourdough and gluten-red bean dough	Water binding capacity and reduced water distribution Cryoprotective on gluten protein matrix	Quality of frozen gluten-red bean dough during freeze–thaw cycles	[69]
Lactiplantibacillus plantarum T1 and CL80	Hetero-EPS T1 and hetero-EPS S-1A	Fermented milk	Viscosity enhancer Water-holding capacity	Gel stability Texture (less hardness and increasing cohesiveness and gumminess)	[71]
Lpb. plantarum CSK	Hetero-EPS CSK (glucose and galactose)	Soymilk fermented	Gel formation, viscosity, water-holding capacity	Gel stability, texture, shelf life, and reducing syneresis	[18]

Table 3. Techno-functional properties of exopolysaccharides (EPSs) in foods produced by lactic acid bacteria.

Lactic Acid Bacteria	EPS Type	Food Product	Techno-Functional Properties	Quality Improvement	Reference
Lactobacillus sanfranciscensis TMW 1.392	Levan (homo-EPS)	Dough/bread	Water absorption, bread volume, and crumb firmness	Bread texture and quality	[75]
Streptococcus thermophilus NIZO2104 and Lactobacillus delbrueckii subsp. bulgaricus DGCC 291	Hetero-EPS NIZO2104: galactose/ribose/N-acetyl-galactosamine DGCC 291: galactose/glucose	Fermented milk	Gel formation, viscosity, and water-holding capacity	Firmness, apparent viscosity, and reduced syneresis	[55]
Lactobacillus curvatus TMW 1.624	Dextran (homo-EPS)	Gluten-free bread	Increase viscosity, gas retention, and water-holding capacity	Bread texture and volume Shelf life by retarding bread staling	[76]
Weissella cibaria MG1	Dextran (homo-EPS)	Malt fermented beverage	Viscosity improver	Beverage stability Desirable body	[56]
Lactobacillus plantarum 162 R, Leuconostoc mesenteroides N6, and the mixture.	Not reported	Sucuk (Turkish-type fermented sausage)	Gel formation and retention water	Texture	[49]
Leuconostoc citreum B-2 Leuconostoc pseudomesenteriodes JF17	Highly branched dextran (homo-EPS) Dextran (homo-EPS)	----	Water-holding capacity	Binding and stabilizing agent of water	[24,64]
Lactobacillus reuteri E81	Glucan (homo EPS constituted of glucose)	Dough/wheat bread	Dough viscoelasticity and retention water	Hardness of fresh bread	[67]
Leuconostoc lactis L2	EPS-L2 (homo EPS constituted of glucose)	Fermented milk	Gel formation, viscosity	Gel stability, texture	[41]
Weissella confusa QS813	Dextran (homo EPS constituted of glucose)	Red bean sourdough and gluten-red bean dough	Water binding capacity and reduced water distribution Cryoprotective on gluten protein matrix	Quality of frozen gluten-red bean dough during freeze–thaw cycles	[69]
Lactiplantibacillus plantarum T1 and CL80	Hetero-EPS T1 and hetero-EPS S-1A	Fermented milk	Viscosity enhancer Water-holding capacity	Gel stability Texture (less hardness and increasing cohesiveness and gumminess)	[71]
Lpb. plantarum CSK	Hetero-EPS CSK (glucose and galactose)	Soymilk fermented	Gel formation, viscosity, water-holding capacity	Gel stability, texture, shelf life, and reducing syneresis	[18]

EPSs play a central role in modifying viscosity and texture in fermented dairy and non-dairy products. Rheologically, EPS often impart pseudoplastic behavior, a desirable property in products like yogurt, sauces, or plant-based creams, where viscosity decreases under shear (during pouring or spooning) but recovers once at rest. Capsular or slime-type EPS produced during fermentation with strains like S. thermophilus or L. bulgaricus are responsible for the high viscosity and creaminess of yogurt and fermented milks [3,14,77]. In fermented milk, neutral EPSs interfere with and prevent protein–protein interactions, thereby weakening the gel’s stiffness. Anionic EPSs interact with positive charges of milk proteins (caseinates), contributing to reinforcing the casein network, leading to a firm gel and greater apparent viscosity [48]. EPSs with high molecular weight, a stiff chain, and little branching enhanced the apparent viscosity, firmness, and whey retention of fermented milk [55]. Their ability to form entangled polymer networks results in shear-thinning behavior and an improved mouthfeel. In non-dairy applications, such as oat- or almond-based fermented beverages, EPS can mimic the body and texture of conventional dairy through similar water-holding and stabilizing effects. The increasing viscosity of milk or plant-based milk alternatives using EPS is caused by the enhancement of protein steric interactions and entanglement [78]. Despite EPS production in liquid media resulting in a viscous suspension, the viscosifying effect depends on its shape (degree of branching and flexibility of the backbone), the molecular weight, and charge [55]. EPSs from Lpb. plantarum CSK lead to decline in shear strain rate, indicating greater elasticity and recovery ability of soymilk gel, achieving better resistance to shear stress and restoration, giving a robust and strengthening gel [78].

The EPS concentration in aqueous suspensions determines the rheological behavior of the suspension. Viscosity is positively correlated with EPS secretion in situ [48]. In general, aqueous suspensions (20–40 mg/mL) present liquid forms that exhibit weak gelling properties and solid-like behavior with increasing EPS concentration, with elastic features acting as a thickener [29,79]. Salt ions, temperature, and the pH of the suspension influence the rheological properties. High temperatures (≥55 °C) increase EPS solubility by increasing the space between molecules and reducing the intermolecular binding forces, which leads to weakening of the apparent viscosity. In addition, high temperatures could promote (electrostatic and hydrogen bonds) the hydrolysis of selected monosaccharides (galactose), favoring the apparent viscosity decrease. The pH conditions impact the relative molecular conformation via the binding force and hydrogen bond strength that form the polysaccharide gel [29]. Anionic EPSs are affected by acidic pHs that induce protonation of charged groups, resulting in a decrease in the electrostatic repulsions, which leads to an increase in EPS chain flexibility and consequently to a reduction in viscosity. In contrast, viscosity at alkaline pH (9.0) remains stable compared with neutral pH (7.0) [79]. Hetero-EPS from Lc. lactis IMAU11823 improved its viscosity with pH levels of 5.0 and 6.0 [31]. The food matrix (nutrients available for bacterial growth and EPS production) also determined the EPS’s rheological properties. For instance, W. confusa C19, a dextran producer strain, increased the viscosity when EPS was produced in maize medium compared with wheat, oat, and rice [66]. In fermented milk, viscosity rose exponentially with the EPS production, which coincides with the optimal EPS production conditions in milk by L. paracasei H9 [48]. Also, fermented milk texture (firmness, cohesiveness) was associated with the release of EPS. EPS viscosity could be modified by the presence of salts (NaCl and CaCl₂), particularly in sauces, salad dressings, gravies, soups, cottage cheese, and processed cheese, which contain high-salt levels. In these foods, the use of EPS should be evaluated before use as an ingredient. The EPS-BMS, an EPS produced by Ln. citreum BMS, maintained its viscosity when 0.05 or 1.00 M of NaCl and 0.05 M CaCl₂ were added into aqueous solution (9%), but increased at 1.00 M CaCl₂, probably due to intermolecular associations caused by Ca²⁺ bridges [79]. The same behavior was recorded for hetero-EPS from Lc. lactis IMAU11823 when metal ions (Na⁺ and Mg²⁺) were added at 10 mg/mL [31].

EPS significantly enhances the water-holding capacity (WHC) of food matrices by binding free water within their molecular structure. This is especially valuable in fermented dairy products such as yogurt, where high WHC reduces syneresis (whey separation) and improves product stability during storage. The efficiency of WHC depends on the molecular weight and branching of the EPS, as well as interactions with milk proteins, primarily casein micelles [1,20,29].

Although not all EPSs possess strong emulsifying abilities, some hetero-EPSs exhibit moderate emulsion-stabilizing capacity through steric stabilization and interfacial activity [4,5]. This is useful in the formulation of dressings, beverages, and plant-based emulsions, where EPSs can improve the dispersion of oil droplets and prevent phase separation. EPS-BMS (from Ln. citreum BMS) at 1.5% efficiently emulsified sunflower oil (emulsion 70.1% after 24 h at 25 °C), and pH level (7 or 3) did not significantly affect the emulsifying activity up to 15 days [79]. Emulsifying stability of sunflower oil and whey protein isolate using xanthan gum (0.5%) at pH ~7 had similar emulsion stability >70% after 24 h at 30 °C [80]. In addition, the interfacial tension of the emulsion formulated using EPS was not affected by acidic pH, making it suitable for use in sauces and salad dressings. At an EPS-BMS concentration of 1%, the emulsion stability was comparable to that of a commercial xanthan emulsion at 3.5 mg/mL [79]. The emulsifying capability results from the thickening properties and the macromolecular barrier that forms between the aqueous medium and the dispersed droplets. EPSs also function as stabilizers in foam-based systems, helping maintain structure and volume in whipped or aerated products.

EPS can interact synergistically or antagonistically with other macromolecules in the food matrix. In dairy systems, EPS–protein interactions can enhance gel strength and consistency by modifying the gel network formed during fermentation or renneting. In cereal-based systems like sourdough, EPS–starch interactions have been shown to improve crumb softness, delay staling, and affect dough rheology, particularly in gluten-free formulations [6,7,14]. The dextran (EPS) is capable of forming hydrogen bonds and steric interactions with S-S bonds and gluten proteins in sourdough (gluten-legume (red bean) dough), resulting in a compact structure that supports distortions caused by ice recrystallization during frozen dough storage or freeze–thaw cycles [69]. Moreover, the incorporation of dextran as an ingredient (ex situ) or in situ production in gluten-red bean dough inhibited ice recrystallization and controlled ice crystal size after five freeze–thaw cycles. Thus, EPS maintained good and stable gluten integrity of dough, preserving the gluten network of frozen gluten-red bean dough that could enhance baking quality parameters of frozen dough [69]. Likewise, in wheat sourdough, α-glucan (in situ produced by L. reuteri E81) interacted with starch granules to form a film-like structure with positive effects on the rheological properties of dough, resulting in a more elastic dough [67]. Supplementation of α-glucan as an ingredient improved the wheat dough viscoelasticity and reduced the dough strength, similar to commercial hydrocolloids (xanthan or modified cellulose) during breadmaking. Furthermore, bread added with 0.09% or 0.19% of α-glucan exhibited lower hardness for 8 days compared with bread containing the in situ α-glucan produced during sourdough [67]. In fermented meat products (fermented sausages), the in situ EPS production promotes a harder gel and water retention, which affects the final texture. The EPS resulted in network structures due to EPS interaction with proteins in the sausage, while a compact structure was observed without EPS strains [49].

4. Health-Promoting Potential of LAB Exopolysaccharides

EPSs produced by LAB are increasingly recognized not only for their technological benefits in food systems but also for their bioactive functions that may contribute to human health. These health-promoting properties are mainly dependent on their structural features, including molecular weight, charge, degree of branching, and the presence of specific monosaccharides or functional groups [4,5]. As natural biopolymers of microbial origin, EPSs are often non-digestible and may exert prebiotic effects, modulate immune responses, reduce oxidative stress, and even improve metabolic health [11,13,71]. The in vitro biological activity of EPS from LAB has been widely reported; in vivo, animal models are also commonly used.

EPS from LAB may act as prebiotic substrates, selectively stimulating the growth of beneficial gut microorganisms such as Bifidobacterium and Lactobacillus species [11]. Unlike digestible carbohydrates, EPSs resist hydrolysis in the upper gastrointestinal tract and reach the colon, where specific microbial populations ferment them. For instance, EPS from Lpb. plantarum stimulated short-chain fatty acid (SCFA) production, particularly butyrate and propionate, in fecal fermentation models [14,77]. Dextran and levan-type EPSs have been shown to increase bifidobacterial abundance and microbial diversity, potentially enhancing colon barrier function and reducing inflammation [10,11,52].

Several EPSs exhibit immunomodulatory effects, including stimulation of macrophages, dendritic cells, and natural killer (NK) cells. These effects are often mediated through pattern recognition receptors (e.g., TLR2, TLR4), triggering cytokine release and enhancing innate and adaptive immunity [13,36]. An intervention (clinical trial) to evaluate the effect of yogurt containing EPS on the immune function against influenza of women healthcare workers showed no significant effect on cytokines (IL-2, IL-4, IL-5, IL-10, IL-12, IL-13, TNF-α) [81]. EPS from L. rhamnosus and L. helveticus induced expression of IL-10 and TNF-α in murine macrophages. Hetero-EPS with uronic acids or phosphate groups showed stronger immunomodulatory effects by enhancing phagocytic activity [4,5]. The immunological benefits may be especially valuable in fermented foods designed to improve gut barrier function, alleviate allergies, or boost immune resilience during infections.

EPSs have demonstrated antioxidant activity via radical-scavenging capacity, metal ion chelation, and inhibition of lipid peroxidation. These properties can help reduce oxidative stress, a known contributor to chronic diseases such as cardiovascular disorders, diabetes, and cancer [2,8,9]. Structural elements such as sulfate groups, phenolic residues (from co-metabolites), or high uronic acid content were associated with greater antioxidant capacity [10,12]. In animal models, antioxidant EPSs have been shown to reduce oxidative biomarkers and improve liver function parameters, suggesting therapeutic potential. Certain EPSs inhibit the growth or biofilm formation of pathogenic bacteria and fungi. The mechanisms include (a) competitive exclusion on surfaces; (b) EPS–pathogen interaction disrupting adhesion; and (c) synergistic effects with organic acids or bacteriocins. Some EPSs are implicated in cholesterol-lowering effects, likely via binding and precipitating bile salts in the intestine. Also, the increasing SCFA production (particularly propionate) inhibits hepatic cholesterol synthesis. Animal studies with EPS-producing Lactobacillus strains have reported reductions in plasma total cholesterol, LDL, and triglycerides, as well as improvements in insulin sensitivity and lipid metabolism [13,14,36].

Table 4 shows selected clinical trials of EPS from LAB and its health benefits. Some findings from in vitro or animal models of the health benefits of EPS have been demonstrated in humans, such as immunomodulatory, antioxidant, anti-inflammatory, hypotriglyceridemic, and dietary fiber effects (Figure 2). Most studies use yogurt, fermented milk, or kefir formulated with LAB producers of EPS, which have previously been investigated for their health benefits in in vitro and/or animal models. Another option is to provide fermented food containing EPS in a dried form (capsules). Japan is a leading country in conducting and publishing clinical trials. The intervention times ranged from 2 to 16 weeks. A limited number of strains have been investigated, and L. delbrueckii subsp. bulgaricus OLL1073R-1 is the most studied, with multiple health benefits associated. S. thermophilus OLS3059, L. paracasei IJH-SONE68, and W. confusa VP30 are also reported. In most studies, the identification and precise mechanisms of action of EPSs on each health benefit are still missing or scarcely understood; thus, further investigations are needed.

Table 4. Health benefits of exopolysaccharides (EPS) from lactic acid bacteria in humans.

Intervention Type and Time	Lactic Acid Bacteria or Food Intake	Primary Effect of EPS	Main Findings	Reference
Randomized controlled study. Simultaneous comparative study in men (>40 from Funagata or >60 years from Arita) for 8 weeks for Funagata and 12 weeks for Arita. The effect on immune system parameters in the elderly and preventive effects against respiratory tract infections (common cold and influenza virus) were investigated	Lactobacillus delbrueckii subsp. bulgaricus OLL1073R-1 produced immunostimulatory EPS and Streptococcus thermophilus OLS3059 Yogurt containing (36.5–68 mg/kg EPS) or milk group 90 g yogurt or 100 mL milk per day	Immunomodulatory activity	The risk of catching the common cold or influenza virus infection was lower in yogurt groups from both places. Lymphocyte blastoid transformation induced by Con A increased in yogurt group from Funagata. Natural killer activity in the low-activity subjects improved to normal values in subjects’ intake yogurt from both places. Thus, yogurt reduced the risk of respiratory infections. The score for eye/nose/throat was higher for yogurt group. Also, yogurt improved the quality-of-life score of the elderly.	[82]
Randomized control pre-test–post-test design on diabetes mellitus (DM) outpatients in various hospitals to investigate the biomolecular nature of the glycemic status of Type 2 DM 30 days	Clear kefir and control group 200 mL/day	Antioxidant activity	HbA1c was significantly reduced in delta level, and insulin was reduced in the groups that consumed clear kefir.	[83]
Randomized, double-blind, controlled trial in men for 12 weeks to investigate the effect of summer heat fatigue	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus OLS3059 Yogurt containing (2.9 mg/100 mL EPS) or placebo (acidified yogurt) 100 mL per day	Antioxidant activity and radical-scavenging activity	The visual analogue scale (VAS) scores for “general malaise”, “feeling languid”, “fatigue”, and “psychological stress” were significantly lower after 12 weeks in the study group. The blood pressure was reduced in the yogurt + EPS group after 4 weeks. The autonomic nervous system balance was better maintained in yogurt + EPS group, relieving the physical and mental disorders induced by seasonal changes.	[84]
Randomized controlled open-label study in women healthcare workers for 16 weeks The effects of influenza infection during winter were studied	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus 112 mL of yogurt per day	Immunomodulatory activity	The production of IFN-γ (immunobiological market) increased in the intervention group. The influenza A or common cold cumulative incidence rate was similar in both groups.	[81]
Randomized, double-blind, parallel, and placebo-controlled in the elderly with limited activity and a mostly sedentary life, resident in nursing homes 12 weeks of intervention to evaluate the saliva flow rate, the total amount of salivary IgA, and the amount of the influenza virus-bound salivary IgA	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS Yogurt with EPS and yogurt fermented with L. bulgaricus OLL1256 (placebo) 100 g yogurt containing EPS or placebo yogurt daily	Immunomodulatory activity	Influenza virus A subtype H3N2-bound IgA in saliva was higher in yogurt EPS compared with placebo.	[85]
Randomized crossover method Adult males participated in the study to evaluate the carotene absorption during co-ingestion with yogurt 2 weeks	L. bulgaricus OLL1256 and S. thermophilus OLS3059 EPS produced strains 100 g yogurt containing (90 μg/g) + 100 g carrot juice concentrate/tomato paste/spinach paste or 100 g water + 100 g carrot juice concentrate/tomato paste/spinach paste	Diffusion mechanisms (emulsifier, dispersion stabilizer, and prolong carotenoid contact with adsorbing membranes)	Higher β-carotene, α-carotene, lycopene and incremental area under the concentration-time curve for β-carotene, α-carotene, and retinyl palmitate, and lycopene concentration for the plasma triacylglycerol-rich lipoprotein fraction at 4, 6, and 8 h were recorded when pastes were intake with yogurt. Lutein concentration increased in total plasma after 2, 4, 6, and 8 h when spinach paste was consumed with yogurt. Thus, yogurt enhanced the bioavailability and absorption of dietary carotenoids in humans.	[86]
Randomized controlled study in women healthcare workers for 16 weeks The impact on psychological quality was assessed	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus 112 mL of yogurt per day	Antioxidant activity	The scores of Pittsburgh Sleep Quality Index, the General Health and Vitality from the eight-item Short Form Health Survey (subjective quality of life), and the constipation of the Gastrointestinal Symptom Rating Scale were improved after 16 weeks of yogurt intake.	[87]
Randomized, double-blind, placebo-controlled study in adults with perennial allergy symptoms to study allergic conditions 12 weeks	Capsules containing 260 mg of dried pineapple juice fermented with Lactobacillus paracasei IJH-SONE68 heat treated, and dextrin or capsules of dextrin as a placebo Four capsules per day	Anti-inflammatory	Head dullness, watery eyes, frequency of nose-blowing, and sneezing symptoms decreased in the study group. Also, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and cholinesterase (serum liver function indices) in serum decline in the study group.	[88]
Randomized, double-blind, placebo-controlled trial in overweight adults to investigate the effect on obesity indices, anti-inflammatory, and other obesity-related factors 12 weeks	Capsules containing 260 mg of dried pineapple juice fermented with L. paracasei IJH-SONE68 heat treated, and dextrin or capsules of dextrin as placebo Four capsules per day	Anti-inflammatory and hypotriglyceridemia	Serum triglyceride and serum liver function indices (aspartate aminotransferase and alanine aminotransferase) levels reduced in the study group. Anaerostipes genus increased while Veillonella decreased from human microbiota in the study group.	[89]
Trial 1: Randomized, double-blind, placebo-controlled, vaccinated male university students Trial 2: Randomized, double-blind, placebo-controlled vaccinated healthy 25- to 59-year-old adults	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus OLS3059 Yogurt containing (3.3 mg EPS) or placebo (acidified yogurt) 112 mL per day	Immunomodulatory activity (act as a B-cell mitogen)	The daily intake of yogurt + EPS augmented the serum antibody titers against the seasonal influenza vaccine. Trial 1: The geometric mean titer (GMT) of the H3N2 and B viruses were significantly higher in the yogurt group fulfilled the EMA criteria of seroprotection, improving the vaccine immunogenicity leading to enhance protection against influenza infection. Trial 2: The GMT of the H1N1 and B viruses was significantly higher in yogurt group, indicating yogurt intake improved the vaccine immunogenicity via serum antibodies production.	[90]
Randomized, double-blind, placebo-controlled human study for 4 weeks to evaluate the effect on functional constipation	Weissella confusa VP30 Pasteurized fermented milk containing 3.52 EPS g/L (control) or 39.2 g/L EPS 200 mL of fermented milk was intake daily	Dietary fiber	Defecation frequency and fecal volume increased while stool hardness and the score sum of symptoms (difficulty, flatulence, pain, bloating, severity) reduced when fermented milk with EPS was consumed for 4 weeks. Regarding laboratory analysis, fecal water content increased in fermented milk + EPS. Weight loss or reduction was also observed in fermented milk + EPS.	[91]

Table 4. Health benefits of exopolysaccharides (EPS) from lactic acid bacteria in humans.

Intervention Type and Time	Lactic Acid Bacteria or Food Intake	Primary Effect of EPS	Main Findings	Reference
Randomized controlled study. Simultaneous comparative study in men (>40 from Funagata or >60 years from Arita) for 8 weeks for Funagata and 12 weeks for Arita. The effect on immune system parameters in the elderly and preventive effects against respiratory tract infections (common cold and influenza virus) were investigated	Lactobacillus delbrueckii subsp. bulgaricus OLL1073R-1 produced immunostimulatory EPS and Streptococcus thermophilus OLS3059 Yogurt containing (36.5–68 mg/kg EPS) or milk group 90 g yogurt or 100 mL milk per day	Immunomodulatory activity	The risk of catching the common cold or influenza virus infection was lower in yogurt groups from both places. Lymphocyte blastoid transformation induced by Con A increased in yogurt group from Funagata. Natural killer activity in the low-activity subjects improved to normal values in subjects’ intake yogurt from both places. Thus, yogurt reduced the risk of respiratory infections. The score for eye/nose/throat was higher for yogurt group. Also, yogurt improved the quality-of-life score of the elderly.	[82]
Randomized control pre-test–post-test design on diabetes mellitus (DM) outpatients in various hospitals to investigate the biomolecular nature of the glycemic status of Type 2 DM 30 days	Clear kefir and control group 200 mL/day	Antioxidant activity	HbA1c was significantly reduced in delta level, and insulin was reduced in the groups that consumed clear kefir.	[83]
Randomized, double-blind, controlled trial in men for 12 weeks to investigate the effect of summer heat fatigue	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus OLS3059 Yogurt containing (2.9 mg/100 mL EPS) or placebo (acidified yogurt) 100 mL per day	Antioxidant activity and radical-scavenging activity	The visual analogue scale (VAS) scores for “general malaise”, “feeling languid”, “fatigue”, and “psychological stress” were significantly lower after 12 weeks in the study group. The blood pressure was reduced in the yogurt + EPS group after 4 weeks. The autonomic nervous system balance was better maintained in yogurt + EPS group, relieving the physical and mental disorders induced by seasonal changes.	[84]
Randomized controlled open-label study in women healthcare workers for 16 weeks The effects of influenza infection during winter were studied	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus 112 mL of yogurt per day	Immunomodulatory activity	The production of IFN-γ (immunobiological market) increased in the intervention group. The influenza A or common cold cumulative incidence rate was similar in both groups.	[81]
Randomized, double-blind, parallel, and placebo-controlled in the elderly with limited activity and a mostly sedentary life, resident in nursing homes 12 weeks of intervention to evaluate the saliva flow rate, the total amount of salivary IgA, and the amount of the influenza virus-bound salivary IgA	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS Yogurt with EPS and yogurt fermented with L. bulgaricus OLL1256 (placebo) 100 g yogurt containing EPS or placebo yogurt daily	Immunomodulatory activity	Influenza virus A subtype H3N2-bound IgA in saliva was higher in yogurt EPS compared with placebo.	[85]
Randomized crossover method Adult males participated in the study to evaluate the carotene absorption during co-ingestion with yogurt 2 weeks	L. bulgaricus OLL1256 and S. thermophilus OLS3059 EPS produced strains 100 g yogurt containing (90 μg/g) + 100 g carrot juice concentrate/tomato paste/spinach paste or 100 g water + 100 g carrot juice concentrate/tomato paste/spinach paste	Diffusion mechanisms (emulsifier, dispersion stabilizer, and prolong carotenoid contact with adsorbing membranes)	Higher β-carotene, α-carotene, lycopene and incremental area under the concentration-time curve for β-carotene, α-carotene, and retinyl palmitate, and lycopene concentration for the plasma triacylglycerol-rich lipoprotein fraction at 4, 6, and 8 h were recorded when pastes were intake with yogurt. Lutein concentration increased in total plasma after 2, 4, 6, and 8 h when spinach paste was consumed with yogurt. Thus, yogurt enhanced the bioavailability and absorption of dietary carotenoids in humans.	[86]
Randomized controlled study in women healthcare workers for 16 weeks The impact on psychological quality was assessed	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus 112 mL of yogurt per day	Antioxidant activity	The scores of Pittsburgh Sleep Quality Index, the General Health and Vitality from the eight-item Short Form Health Survey (subjective quality of life), and the constipation of the Gastrointestinal Symptom Rating Scale were improved after 16 weeks of yogurt intake.	[87]
Randomized, double-blind, placebo-controlled study in adults with perennial allergy symptoms to study allergic conditions 12 weeks	Capsules containing 260 mg of dried pineapple juice fermented with Lactobacillus paracasei IJH-SONE68 heat treated, and dextrin or capsules of dextrin as a placebo Four capsules per day	Anti-inflammatory	Head dullness, watery eyes, frequency of nose-blowing, and sneezing symptoms decreased in the study group. Also, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and cholinesterase (serum liver function indices) in serum decline in the study group.	[88]
Randomized, double-blind, placebo-controlled trial in overweight adults to investigate the effect on obesity indices, anti-inflammatory, and other obesity-related factors 12 weeks	Capsules containing 260 mg of dried pineapple juice fermented with L. paracasei IJH-SONE68 heat treated, and dextrin or capsules of dextrin as placebo Four capsules per day	Anti-inflammatory and hypotriglyceridemia	Serum triglyceride and serum liver function indices (aspartate aminotransferase and alanine aminotransferase) levels reduced in the study group. Anaerostipes genus increased while Veillonella decreased from human microbiota in the study group.	[89]
Trial 1: Randomized, double-blind, placebo-controlled, vaccinated male university students Trial 2: Randomized, double-blind, placebo-controlled vaccinated healthy 25- to 59-year-old adults	L. bulgaricus OLL1073R-1 produced immunostimulatory EPS and S. thermophilus OLS3059 Yogurt containing (3.3 mg EPS) or placebo (acidified yogurt) 112 mL per day	Immunomodulatory activity (act as a B-cell mitogen)	The daily intake of yogurt + EPS augmented the serum antibody titers against the seasonal influenza vaccine. Trial 1: The geometric mean titer (GMT) of the H3N2 and B viruses were significantly higher in the yogurt group fulfilled the EMA criteria of seroprotection, improving the vaccine immunogenicity leading to enhance protection against influenza infection. Trial 2: The GMT of the H1N1 and B viruses was significantly higher in yogurt group, indicating yogurt intake improved the vaccine immunogenicity via serum antibodies production.	[90]
Randomized, double-blind, placebo-controlled human study for 4 weeks to evaluate the effect on functional constipation	Weissella confusa VP30 Pasteurized fermented milk containing 3.52 EPS g/L (control) or 39.2 g/L EPS 200 mL of fermented milk was intake daily	Dietary fiber	Defecation frequency and fecal volume increased while stool hardness and the score sum of symptoms (difficulty, flatulence, pain, bloating, severity) reduced when fermented milk with EPS was consumed for 4 weeks. Regarding laboratory analysis, fecal water content increased in fermented milk + EPS. Weight loss or reduction was also observed in fermented milk + EPS.	[91]

The primary effect of EPS against functional constipation is providing dietary fiber. EPSs from W. confusa VP30 have a water-holding capacity between 500 and 2500%, thus increasing the water content of feces in the intestine, resulting in relief of constipation symptoms by improving bowel movement, supporting colonic motility, and resulting in shorter intestinal transit time by minimizing water reabsorption in the colon [91]. In other studies, the specific effects and the mechanisms of EPS need to be investigated, such as in immunomodulatory effects on influenza vaccines, while in summer heat fatigue, the inclusion of indicators associated with antibody production and dysfunction of the immune system are required to understand the EPS effects [87,90]. EPSs (L. bulgaricus OLL1073-R-1) probably induce production of high-affinity IgA specific to influenza virus via the pathway of T cell dependency that upregulates TGF-β1, IL-21, and retinoic acid or IFN-γ cytokine to reduce respiratory infections [81,85]. To enhance carotenoid bioavailability and absorption, the EPS and milk proteins in yogurt together could enhance intestinal membrane permeability and prolong the contact between carotenoids and membranes by EPS adhesive properties [86]. EPS’s psychological effects (improved sleep quality) could reflect in reduced fatigue; however, the mechanisms of EPS on the quality of sleep were not fully understood [87].

EPS’s health effects on clinical trials have been explored for ~15 years, and findings highlight their benefits. However, much further research is needed to elucidate the mechanisms of action of EPS to understand how to stimulate or regulate the immune system against viral respiratory infections. Immunomodulatory activity against bacterial infections has not been studied yet. The antioxidant activity, hypoglycemic, anti-inflammatory, and hypolipidemic effects of EPS, as well as their impact on nutrient bioavailability, have been less investigated. First, strong relationships and evidence of these effects should be demonstrated and verified. Afterwards, the mechanisms of action should be studied. Other health benefits reported in vitro and/or in animals, such as prebiotic, antitumor, anti-obesity, hepatoprotective, and antimicrobial effects, require investigation in clinical trials. In addition, clinical trials with different populations, various fermented foods, isolated EPS, and EPS presentations (powders, tablets) are needed worldwide.

Critical factors in intervention studies are dose and time, as health effects depend directly on them. The adequate EPS doses in clinical trials are important to ensure the expected health effects. Sometimes, for easy handling, dried preparations are administered but do not contain an adequate dose of bioactive compound; for instance, Danshiitsoodol et al. [89] tested the effects of EPS through dried fermented pineapple juice + dextrin on overweight adults to evaluate weight gain and visceral fat. However, the doses administered in the clinical trial were insufficient to demonstrate previously found results in animals; the tested dose was less than 1/10. In addition, the intervention time is critical for observing the health effects, as certain studies have found that 16 weeks is too short a time.

5. Food Applications of LAB Exopolysaccharides

Figure 3 provides an overview of the main food applications of LAB-derived exopolysaccharides, distinguishing between in situ production during fermentation and ex situ incorporation of purified EPSs. While the former approach exploits microbial activity directly within the food matrix to improve texture, stability, and functionality, the latter allows the targeted use of EPS as natural additives with thickening, gelling, or film-forming properties. Together, these strategies highlight the versatility of LAB exopolysaccharides as multifunctional ingredients in diverse food systems.

5.1. Direct Addition of Purified EPS to Food Products

Purified EPS from LAB are gaining momentum as clean-label alternatives to commercial hydrocolloids such as carrageenan, pectin, xanthan, gellan, dextran, and curdlan [92]. Their high molecular weight (0.1–3 MDa), hydroxyl group density, and, in some cases, charged residues, allow them to bind water and adsorb at oil– or air–water interfaces. These properties contribute to improved viscosity, emulsion stability, and water retention in diverse food systems, including dairy, bakery, and beverages. EPSs can be introduced into formulations as purified ingredients via direct addition (ex situ), spray drying, or co-dissolution into food matrices. This method enables precise dosing, independent of fermentation variables, offering advantages over in situ production. At concentrations as low as 0.2–1% (w/w), branched α-glucans (e.g., dextran, reuteran), β-glucans, kefiran, and hetero-EPS from genera such as Weissella, Leuconostoc, and Lactobacillus can exhibit shear-thinning behavior comparable to commercial gums, while remaining fully biodegradable [14,78].

A summary of functional applications of purified EPSs from LAB across food systems is presented in Table 5. In bakery products, they improve dough rheology, moisture retention, and staling resistance; in dairy products, they enhance creaminess, reduce syneresis, and stabilize emulsions; in frozen desserts, they inhibit ice recrystallization and improve melt resistance. Additionally, EPSs can act as natural emulsifiers. Zammouri et al. [93] demonstrated that dextran-rich fractions from W. cibaria and Lc. lactis reduced interfacial tension by ~40% and stabilized oil-in-water emulsions for over 30 days, thanks to their anionic backbones (ζ-potential: −12 to −24 mV), which prevented droplet flocculation. In dough systems, EPS reinforce the gluten–starch matrix. For instance, a 759 kDa dextran from W. cibaria FAFU821 improved water-holding and reduced crumb hardness when incorporated at 0.5% (flour basis), owing to a three-dimensional α-(1→6)-linked network [94]. Similar effects were observed with kefiran gels (1–3% w/w), which increased water absorption and dough development time while reducing softening, leading to softer, more extensible loaves without altering pH or flavor [95,96,97,98]. From a production perspective, for EPSs to be viable as functional ingredients, yields should reach 10–15 g/L [9]. However, many LAB yielded lower EPS titers compared with other microorganisms, and scaling up production without compromising molecular consistency remains a challenge.

Table 5. Summary of applications of exopolysaccharides (EPS) from lactic acid bacteria (LAB) purified in food systems.

Application Area	Function of EPS	Food Examples	Relevant EPS Types/Sources	Reference
Baked Goods (wheat-based)	Improve dough rheology, moisture retention, and softness	Bread, rolls, cakes	Dextran (Weissella cibaria), kefiran	[94,95,96]
Gluten-Free Products	Mimic gluten structure, enhance volume, delay staling	Gluten-free bread, muffins, pizza bases	Dextran, β-glucans, heteropolysaccharides	[14,99]
Fermented Beverages	Improve mouthfeel, suspension, viscosity	Soy yogurt, oat drinks, kefir	Kefiran, dextran	[34,78]
Dairy Products	Enhance creaminess, reduce syneresis, stabilize emulsions	Yogurt, cream cheese, dairy emulsions	EPS from Leuconostoc pseudomesenteroides, Leuconostoc mesenteroides F27, Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, Weissella confusa	[13,99,100]
Emulsified Foods	Stabilize oil droplets, reduce interfacial tension	Salad dressings, emulsifier for low-fat mayonnaise	Heteropolysaccharides from L. plantarum, Leuconostoc lactis GW-6	[101,102,103]
Frozen Desserts	Inhibit ice recrystallization, improve texture	Low-fat ice cream, frozen yogurt	Dextran, kefiran, EPS from Leuconostoc citreum-BMS	[79,99,104]
Edible Films and Coatings	Moisture/O2 barrier, antimicrobial or antioxidant carrier	Fresh produce, cheese slices, minimally processed foods	Kefiran, dextran, composite blends with proteins/lipids	[105,106,107,108]

Table 5. Summary of applications of exopolysaccharides (EPS) from lactic acid bacteria (LAB) purified in food systems.

Application Area	Function of EPS	Food Examples	Relevant EPS Types/Sources	Reference
Baked Goods (wheat-based)	Improve dough rheology, moisture retention, and softness	Bread, rolls, cakes	Dextran (Weissella cibaria), kefiran	[94,95,96]
Gluten-Free Products	Mimic gluten structure, enhance volume, delay staling	Gluten-free bread, muffins, pizza bases	Dextran, β-glucans, heteropolysaccharides	[14,99]
Fermented Beverages	Improve mouthfeel, suspension, viscosity	Soy yogurt, oat drinks, kefir	Kefiran, dextran	[34,78]
Dairy Products	Enhance creaminess, reduce syneresis, stabilize emulsions	Yogurt, cream cheese, dairy emulsions	EPS from Leuconostoc pseudomesenteroides, Leuconostoc mesenteroides F27, Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, Weissella confusa	[13,99,100]
Emulsified Foods	Stabilize oil droplets, reduce interfacial tension	Salad dressings, emulsifier for low-fat mayonnaise	Heteropolysaccharides from L. plantarum, Leuconostoc lactis GW-6	[101,102,103]
Frozen Desserts	Inhibit ice recrystallization, improve texture	Low-fat ice cream, frozen yogurt	Dextran, kefiran, EPS from Leuconostoc citreum-BMS	[79,99,104]
Edible Films and Coatings	Moisture/O2 barrier, antimicrobial or antioxidant carrier	Fresh produce, cheese slices, minimally processed foods	Kefiran, dextran, composite blends with proteins/lipids	[105,106,107,108]

5.1.1. Milk Products

In dairy applications, such as yogurts, cream cheese, and milk-based beverages, purified EPSs enhance creaminess, reduce syneresis, and stabilize emulsions. These polymers interact with milk proteins to form cohesive gel networks, contributing to improved body and spoonability [2,8,9,109]. EPSs from S. thermophilus, L. bulgaricus, and W. confusa have shown efficacy in post-fermentation enrichment, especially in low-fat or plant-based variants [11,13,18,100]. Additionally, particular EPSs act as cryoprotectants in frozen dairy desserts. Their high hydration capacity reduces ice recrystallization and improves melt resistance in low-fat ice creams and frozen yogurts, thereby contributing to sensory quality and shelf life [99,104].

5.1.2. Bakery and Gluten-Free Products

In conventional and gluten-free (GF) baked goods, EPSs improve water retention, increase loaf volume, and delay staling. Dextrans and β-glucans mimic gluten structures through hydrogen bonding and formation of pseudo-networks with starch and proteins [14,99]. Their inclusion at 1–2% in GF batters significantly enhance specific volume and textural properties [6]. Similarly, Rühmkorf et al. [76] reported that dextran from L. curvatus offered superior water retention and crumb softness in GF breads compared to other EPS and hydroxypropyl methylcellulose (HPMC), highlighting the importance of EPS structure—specifically branching and molecular weight—for optimal functionality [110].

5.1.3. Beverages and Emulsions

In beverages such as oat and soy drinks, kefir, and smoothies, EPSs improve mouthfeel and suspension stability by forming entangled networks at low concentrations (0.1–0.5% w/v) [99,111]. In emulsified systems, EPSs serve dual roles: as thickening agents and as bio-emulsifiers. Hetero-EPSs from L. plantarum effectively stabilize oil-in-water emulsions with droplet sizes below 5 µm and shelf stability exceeding 30 days, resembling lecithin [101,103]. However, excessive dosing can lead to undesirable viscosity or mouth-coating sensations, necessitating careful optimization depending on the application [4,5].

5.1.4. Films and Coatings

Due to their biocompatibility, film-forming ability, and barrier properties, EPSs are excellent candidates for edible films and active coatings. Polymers like kefiran and dextran can form transparent, flexible films with good mechanical and barrier properties [107,112]. These coatings, when applied to fresh produce or cheeses, help preserve firmness, reduce weight loss, and maintain color and antioxidant levels [108]. Moreover, composite films incorporating proteins or lipids exhibit enhanced moisture and oxygen barrier properties, making them suitable for semi-hard cheese or deli meats [105,108]. EPS films also serve as carriers for probiotics, enzymes, or antimicrobials, enabling innovative packaging applications [113,114].

5.2. In Situ Production of EPS in Fermented Foods

In recent years, the in situ production of EPS has gathered increasing interest. This approach provides a cost-effective alternative to the direct addition of purified EPS; however, it may also lead to unintended alterations in sensory attributes, including taste and flavor. Table 6 summarizes recent studies on in situ EPS production by LAB across various food matrices, emphasizing key findings and significant changes in product quality attributes.

5.2.1. Dairy Products

In situ EPS production has been extensively studied in dairy products, mainly due to their inherent compatibility with lactic acid fermentation processes. Research has primarily focused on yogurt, aiming to enhance its physical stability during storage. Several studies have demonstrated that the use of EPS-producing starter cultures, such as Levilactobacillus brevis, Lc. lactis, and Ln. mesenteroides or dairy-native strains identified as EPS producers (L. bulgaricus 210R and S. thermophilus NIZO 2104), improves yogurt stability without adversely affecting sensory characteristics [7,115,116,117]. Moreover, Ramos et al. [115] demonstrated that in situ EPS production in low-fat yogurt offers a promising strategy to eliminate the need for hydrocolloids, contributing to the development of clean-label products.

The in situ formation of EPS in various types of cheeses to enhance product stability has been studied in recent years [118,119,120]. Generally, researchers have observed that EPS production improves the quality characteristics of different cheese varieties. Similarly, the nature of the EPS synthesized during cheese fermentation has a positive influence on specific quality attributes of the cheese. For example, in cream cheese, ropy EPS enhanced yield stress and creaminess, while capsular EPS improved serum retention after curd homogenization. Furthermore, L. plantarum JLK0142, when applied in low-fat Cheddar cheese fermentation, improved ripening characteristics, moisture retention, and sensory attributes, including appearance, flavor, and overall acceptability. These findings highlight the functional potential of in situ EPS production for enhancing texture, yield, and sensory quality in low-fat dairy matrices.

5.2.2. Meat Products

In meat products, LAB, together with other starter cultures, play a dual role by modulating sensory properties, such as texture and flavor, while also ensuring the microbial safety of the final products [34]. In fermented meat products, such as sausages, L. sakei and P. acidilactici are frequently employed as starter cultures due to their ability to produce organic acids and bacteriocins, which effectively suppress the growth of Listeria monocytogenes and Clostridium botulinum [34]. Accordingly, in recent years, the incorporation of EPS-producing LAB as starter cultures in meat products has been increasingly investigated. Hilbing et al. [50] evaluated the ability of various EPS-producing LAB strains to grow in brine during the production and storage of ham, as well as the impact of EPS synthesis on product characteristics. The researchers observed that homo-EPS-producing strains L. sakei 1.411 and L. curvatus 1.624, together with hetero-EPS-producing strains L. plantarum 1.1478 and 1.25, significantly increased the water-holding capacity of the product, a key factor in producing high-quality hams. This capability is particularly relevant for the tumbling stage in the production of cooked ham. However, the study did not include a detailed assessment of the quality or sensory properties of the cooked ham.

In fermented sausages, the addition of EPS-producing LAB, such as L. plantarum TMW 1.1478, resulted in products with a softer and less firm texture, which is atypical for this type of product [121]. Similarly, Hilbing et al. [122] demonstrated that the use of a homo-EPS-producing LAB (L. curvatus TMW 1.1928 and L. sakei TMW 1.411) as starter cultures in reduced-fat spreadable fermented sausage (Teewurst) improved the texture and spreadability without significantly altering other product characteristics. The researchers confirmed that these textural improvements were directly linked to homo-EPS formation. The application of EPS-producing LAB in low-fat spreadable meat products represents a promising alternative to reduce the need for hydrocolloids in this type of food.

5.2.3. Bakery Products

Lactic acid fermentations in sourdough have been extensively studied. In general, the incorporation of sourdough improves bread quality attributes such as texture, aroma, flavor, and shelf life and delays the staling process [123]. Moreover, the ability of EPSs to retain water makes them an interesting compound for in situ synthesis by LAB in sourdough, allowing their integration into the formulation to enhance bread characteristics. For example, Zhang et al. [94] used sourdoughs fermented with W. cibaria FAFU821 and observed improved bread moisture retention and reduced crumb hardness, thus extending the shelf life of the product. On the other hand, İspirli et al. [67] reported different results. Although α-glucan formation in sourdoughs fermented with L. reuteri E81 improved the dough’s rheological properties, the final bread texture was not affected. While wheat flour provides a suitable medium for the growth of EPS-producing LAB, the addition of specific sugars to the sourdough formulation can further enhance EPS production. For instance, the addition of sucrose to sourdough fermented with W. confusa QS813 (an EPS-producing strain) has been shown to significantly increase EPS synthesis, thereby improving the quality attributes of sourdough bread [124].

Due to the recent rise in demand for gluten-free breads, the food industry has explored the use of various hydrocolloids to improve the texture of these products. The in situ production of EPS in sourdough represents a promising alternative to achieve desirable gluten-free baked goods without more additives. Montemurro et al. [125] used W. cibaria P9 in type-II sourdoughs with sucrose to stimulate EPS synthesis, producing gluten-free breads with favorable protein digestibility and low sugar/fat content. In general, LAB from the genus Weissella have been shown to enhance the texture, overall acceptability, and quality of gluten-free breads [65,126,127].

5.2.4. Beverage Products

The production of EPSs in plant-based beverages has garnered significant attention due to their potential to improve product texture and confer functional benefits. Recent studies have demonstrated successful applications of EPS-producing LAB in non-dairy matrices, offering promising alternatives to traditional dairy products. Goveas et al. [128] developed a coconut water-based functional beverage fermented with L. plantarum SVP2, which was enriched with EPS and exhibited probiotic properties. During seven days of refrigerated storage, the EPS content, pH, and bacterial viability remained stable, indicating good product shelf life. Sensory evaluation revealed moderate consumer acceptance, with texture and sweet–sour flavor receiving favorable feedback. Similarly, Huang et al. [129] investigated the fermentation of soymilk using either wild-type W. confusa or its mutant strain. Both strains improved the water-holding capacity and viscosity of the fermented soymilk, highlighting their potential to enhance the textural properties of plant-based dairy alternatives. The in situ production of EPS in such matrices represents a viable strategy to achieve texture profiles comparable to conventional dairy products.

Table 6. Summary of applications of exopolysaccharides (EPS) from lactic acid bacteria (LAB) produced in situ food systems.

Type of Food	LAB Strain	Improvements in the Product Due to EPS	Reference
Dairy products
Reduced-fat yogurts	Levilactobacillus brevis UCLM-Lb47, Leuconostoc mesenteroides subsp. mesenteroides 6F6-12 and Ln. mesenteroides subsp. mesenteroides 2F6-9	Increased water-holding capacity. Higher EPS levels. Greater mouthfeel viscosity. A possible alternative to the use of hydrocolloids or gums in reduced-fat yogurts.	[115]
Fermented milk (yogurt type)	Lactobacillus helveticus LH18	Increases the product’s consistency. Enhances water-holding capacity. Reduces syneresis. Improves the overall texture of the product.	[7]
Fermented milk (yogurt type)	Streptococcus thermophilus (capsular exopolysaccharide producer) and Lactococcus lactis (non-capsular exopolysaccharide producer)	The combination of both LAB strains improves the protein network structure, resulting in smaller pores, reduced syneresis, and enhanced gel stability.	[116]
Dairy model system simulating yogurt conditions	Lactobacillus delbrueckii subsp. bulgaricus 210R and S. thermophilus NIZO 2104 S. thermophilus HC15 and L. bulgaricus DGCC 291	Linear, stiff, and negatively charged EPS likely enhanced gel stiffness (elastic modulus) through electrostatic interactions with caseins and contributed to increased viscosity. Neutral and stiff EPS increased viscosity by promoting water retention and increasing the bulk volume; however, their effect on gel stiffness was likely limited due to thermodynamic incompatibility.	[117]
Requeson-Type Cheese	L. bulgaricus NCFB 2772 and S. thermophilus SY-102	The co-culture exhibited the highest EPS production compared to the monocultures. Enhanced water retention in the co-culture cheese, resulting in increased cohesiveness and reduced hardness of the product. Fermentation with LAB and the production of EPS nearly doubled the cheese yield.	[118]
Cream cheese	Lc. lactis LL-1 (ropy EPS producer) Lc. lactis LL-2A (capsular EPS producer) Lc. lactis LL-2 (non-ropy EPS producer)	The presence of ropy EPS resulted in higher yield stress and creaminess. Capsular EPS presumably leads to higher serum retention of cheese after curd homogenization at 0.05 and 15 MPa. Higher firmness and serum retention after curd homogenization at higher pressure. Suggested conditions to obtain a cream cheese with greater creaminess, firmness, and whey retention were 0.05/15 MPa for ropy and capsular EPS; 15/30 MPa for non-ropy EPS.	[119]
Low-fat Cheddar cheese	Lactobacillus plantarum JLK0142	Enhancement of the ripening properties. Improved moisture retention. Improved textural and sensory properties (appearance, flavor, and overall acceptance).	[120]
Meat products
Cooked ham model systems	Homopolysaccharides producer (Lactobacillus curvatus TMW 1.624 and Lactobacillus sakei TMW 1.411) Heteropolysaccharides producer (L. plantarum TMW 1.1478 and TMW 1.25)	Homo-EPS-producing strains L. sakei 1.411 and L. curvatus 1.624, along with hetero-PS-producing strains L. plantarum 1.1478 and 1.25, were able to synthesize EPS not only at 15 °C but also at 2 °C within the initial 10–24 h of storage, a feature crucial for the tumbling stage in cooked ham manufacturing. Enhanced water retention in the ham. The study did not include an in-depth evaluation of the cooked ham’s quality or sensory attributes.	[50]
Fermented sausages (salami)	L. plantarum TMW 1.1478	The EPS were predominantly formed during the first 72 h of fermentation at 24 °C. The sausage fermented with the EPS-producing LAB exhibited a softer texture, which is atypical for this type of product. No alterations or adverse effects were detected in the flavor of the final product. Therefore, the application of this LAB strain could represent a potential alternative for the development of spreadable fermented meat products.	[121]
Fat-reduced raw fermented sausages (Teewurst)	Homopolysaccharides producer (L. curvatus TMW 1.1928 and L. sakei TMW 1.411) Heteropolysaccharides producer (L. plantarum TMW 1.1478)	The homopolysaccharide-producing strains reduced the hardness of the fat-reduced sausages. Homopolysaccharides LAB were rated softer and more spreadable than the corresponding control samples. The presence of EPS from LAB did not negatively influence the taste of the products.	[122]
Bakery products
Sourdough bread	Weissella cibaria FAFU821	Enhanced viscoelasticity of sourdough. Improved bread moisture retention by increasing the water-holding capacity. Reduces the hardness of bread. Increased the volatile profile of bread, including linoleic acid ethyl ester and acetic acid.	[94]
Sourdough bread	Lactobacillus reuteri E81	In situ α-glucan production enhanced dough elasticity. No significant changes were observed in the bread’s textural characteristics.	[67]
Sorghum sourdough bread (gluten-free)	Dextran-forming W. cibaria MG1 Reuteran producing L. reuteri VIP Fructan-forming L. reuteri Y2	The three types of EPS generated during sourdough fermentation contributed to a softer crumb in both fresh and stored sorghum bread. Dextran demonstrated the most significant effect on extending shelf life, reducing firmness in bread. All three strains synthesized oligosaccharides during sorghum sourdough fermentation, enhancing the nutritional value of gluten-free sorghum bread.	[127]
Chickpea sourdough	Weissella confusa Ck15	The production of EPS increased the dough viscosity. The researchers did not perform texture analysis on the bread.	[65]
Buckwheat bread	W. cibaria NC516.11	Improve the rheological properties and viscoelastic properties of sourdough. W. cibaria NC516.11 significantly improved the texture of the bread and reduced the hardness and moisture loss during storage.	[126]
Chinese steamed bread	W. confusa QS813 + sucrose addition.	The overall quality of the bread improved with the addition of the LAB strain and sucrose. The presence of EPS positively influenced dough behavior and bread quality.	[124]
Beverage products
Coconut water-based beverage	L. plantarum SVP2.	A non-dairy functional beverage enriched with exopolysaccharides and exhibiting probiotic benefits was successfully developed. During 7 days of refrigerated storage, the EPS content, pH, and bacterial viability remained nearly unchanged. The beverage exhibited moderate acceptance, with texture and flavor (sweet–sour) receiving favorable evaluations.	[128]
Fermented soymilk	W. confusa wild-type or sac mutant	Fermentation of soymilk with either the W. confusa wild-type or its sac mutant resulted in notable improvements in water-holding capacity and viscosity, indicating their potential. The production of EPS in fermented plant-based alternatives represents a promising strategy for achieving textural properties comparable to those of conventional dairy products.	[129]

Table 6. Summary of applications of exopolysaccharides (EPS) from lactic acid bacteria (LAB) produced in situ food systems.

Type of Food	LAB Strain	Improvements in the Product Due to EPS	Reference
Dairy products
Reduced-fat yogurts	Levilactobacillus brevis UCLM-Lb47, Leuconostoc mesenteroides subsp. mesenteroides 6F6-12 and Ln. mesenteroides subsp. mesenteroides 2F6-9	Increased water-holding capacity. Higher EPS levels. Greater mouthfeel viscosity. A possible alternative to the use of hydrocolloids or gums in reduced-fat yogurts.	[115]
Fermented milk (yogurt type)	Lactobacillus helveticus LH18	Increases the product’s consistency. Enhances water-holding capacity. Reduces syneresis. Improves the overall texture of the product.	[7]
Fermented milk (yogurt type)	Streptococcus thermophilus (capsular exopolysaccharide producer) and Lactococcus lactis (non-capsular exopolysaccharide producer)	The combination of both LAB strains improves the protein network structure, resulting in smaller pores, reduced syneresis, and enhanced gel stability.	[116]
Dairy model system simulating yogurt conditions	Lactobacillus delbrueckii subsp. bulgaricus 210R and S. thermophilus NIZO 2104 S. thermophilus HC15 and L. bulgaricus DGCC 291	Linear, stiff, and negatively charged EPS likely enhanced gel stiffness (elastic modulus) through electrostatic interactions with caseins and contributed to increased viscosity. Neutral and stiff EPS increased viscosity by promoting water retention and increasing the bulk volume; however, their effect on gel stiffness was likely limited due to thermodynamic incompatibility.	[117]
Requeson-Type Cheese	L. bulgaricus NCFB 2772 and S. thermophilus SY-102	The co-culture exhibited the highest EPS production compared to the monocultures. Enhanced water retention in the co-culture cheese, resulting in increased cohesiveness and reduced hardness of the product. Fermentation with LAB and the production of EPS nearly doubled the cheese yield.	[118]
Cream cheese	Lc. lactis LL-1 (ropy EPS producer) Lc. lactis LL-2A (capsular EPS producer) Lc. lactis LL-2 (non-ropy EPS producer)	The presence of ropy EPS resulted in higher yield stress and creaminess. Capsular EPS presumably leads to higher serum retention of cheese after curd homogenization at 0.05 and 15 MPa. Higher firmness and serum retention after curd homogenization at higher pressure. Suggested conditions to obtain a cream cheese with greater creaminess, firmness, and whey retention were 0.05/15 MPa for ropy and capsular EPS; 15/30 MPa for non-ropy EPS.	[119]
Low-fat Cheddar cheese	Lactobacillus plantarum JLK0142	Enhancement of the ripening properties. Improved moisture retention. Improved textural and sensory properties (appearance, flavor, and overall acceptance).	[120]
Meat products
Cooked ham model systems	Homopolysaccharides producer (Lactobacillus curvatus TMW 1.624 and Lactobacillus sakei TMW 1.411) Heteropolysaccharides producer (L. plantarum TMW 1.1478 and TMW 1.25)	Homo-EPS-producing strains L. sakei 1.411 and L. curvatus 1.624, along with hetero-PS-producing strains L. plantarum 1.1478 and 1.25, were able to synthesize EPS not only at 15 °C but also at 2 °C within the initial 10–24 h of storage, a feature crucial for the tumbling stage in cooked ham manufacturing. Enhanced water retention in the ham. The study did not include an in-depth evaluation of the cooked ham’s quality or sensory attributes.	[50]
Fermented sausages (salami)	L. plantarum TMW 1.1478	The EPS were predominantly formed during the first 72 h of fermentation at 24 °C. The sausage fermented with the EPS-producing LAB exhibited a softer texture, which is atypical for this type of product. No alterations or adverse effects were detected in the flavor of the final product. Therefore, the application of this LAB strain could represent a potential alternative for the development of spreadable fermented meat products.	[121]
Fat-reduced raw fermented sausages (Teewurst)	Homopolysaccharides producer (L. curvatus TMW 1.1928 and L. sakei TMW 1.411) Heteropolysaccharides producer (L. plantarum TMW 1.1478)	The homopolysaccharide-producing strains reduced the hardness of the fat-reduced sausages. Homopolysaccharides LAB were rated softer and more spreadable than the corresponding control samples. The presence of EPS from LAB did not negatively influence the taste of the products.	[122]
Bakery products
Sourdough bread	Weissella cibaria FAFU821	Enhanced viscoelasticity of sourdough. Improved bread moisture retention by increasing the water-holding capacity. Reduces the hardness of bread. Increased the volatile profile of bread, including linoleic acid ethyl ester and acetic acid.	[94]
Sourdough bread	Lactobacillus reuteri E81	In situ α-glucan production enhanced dough elasticity. No significant changes were observed in the bread’s textural characteristics.	[67]
Sorghum sourdough bread (gluten-free)	Dextran-forming W. cibaria MG1 Reuteran producing L. reuteri VIP Fructan-forming L. reuteri Y2	The three types of EPS generated during sourdough fermentation contributed to a softer crumb in both fresh and stored sorghum bread. Dextran demonstrated the most significant effect on extending shelf life, reducing firmness in bread. All three strains synthesized oligosaccharides during sorghum sourdough fermentation, enhancing the nutritional value of gluten-free sorghum bread.	[127]
Chickpea sourdough	Weissella confusa Ck15	The production of EPS increased the dough viscosity. The researchers did not perform texture analysis on the bread.	[65]
Buckwheat bread	W. cibaria NC516.11	Improve the rheological properties and viscoelastic properties of sourdough. W. cibaria NC516.11 significantly improved the texture of the bread and reduced the hardness and moisture loss during storage.	[126]
Chinese steamed bread	W. confusa QS813 + sucrose addition.	The overall quality of the bread improved with the addition of the LAB strain and sucrose. The presence of EPS positively influenced dough behavior and bread quality.	[124]
Beverage products
Coconut water-based beverage	L. plantarum SVP2.	A non-dairy functional beverage enriched with exopolysaccharides and exhibiting probiotic benefits was successfully developed. During 7 days of refrigerated storage, the EPS content, pH, and bacterial viability remained nearly unchanged. The beverage exhibited moderate acceptance, with texture and flavor (sweet–sour) receiving favorable evaluations.	[128]
Fermented soymilk	W. confusa wild-type or sac mutant	Fermentation of soymilk with either the W. confusa wild-type or its sac mutant resulted in notable improvements in water-holding capacity and viscosity, indicating their potential. The production of EPS in fermented plant-based alternatives represents a promising strategy for achieving textural properties comparable to those of conventional dairy products.	[129]

6. Challenges and Limitations in the Application of LAB Exopolysaccharides

Despite the promising technological and health-promoting properties of EPS produced by LAB, their broader application in food systems is constrained by a series of scientific, technological, and regulatory hurdles. Understanding these constraints is essential for developing effective strategies that enable the standardized and sustainable integration of EPS into diverse food matrices [3,11,14,77,130].

6.1. Low and Variable Yields

One of the main bottlenecks is the low and highly variable EPS yield produced by LAB under standard fermentation conditions [9]. EPSs are typically secreted in amounts below 0.1% w/v, insufficient for economically viable applications, especially when compared to high-yield microbial polysaccharides like xanthan or gellan gums [2]. This limitation could be more pronounced in in situ applications, where EPS must contribute to food texture or bioactivity without precise control over concentration [131].

EPS production is highly strain-specific and influenced by factors such as carbon source, pH, temperature, oxygen availability, and fermentation time [132,133,134]. For example, some Lpb. plantarum strains may produce negligible levels of EPSs in dairy substrates but significantly more in sucrose-rich plant matrices, while Leuconostoc spp. require sucrose feeding for optimal EPS synthesis [135,136,137]. Such complexity complicates scale-up and reproducibility. To improve yields, approaches like metabolic engineering, adaptive evolution, and co-culture systems have been explored [4,5,103]. Gene editing techniques, including CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats), have been employed to enhance the expression of key biosynthetic enzymes [138,139,140,141]. Although promising, these methods face challenges related to regulation, cost, and stability that hinder their widespread adoption in industry. A more feasible strategy may lie in combining precision strain selection and traditional fermentation optimization with multi-omics tools to guide targeted improvements in both purified and in situ applications [10,11,43,142].

6.2. Structural Heterogeneity and Lack of Standardization

The structural complexity and variability of EPS represent another barrier. Unlike standardized polysaccharides like pectin or carrageenan, hetero-EPSs are composed of glucose, galactose, rhamnose, mannose, and uronic acids, with differences in chain length, branching, and glycosidic bonds [7,14,143,144]. These subtle variations have a significant impact on functional attributes, including viscosity, fermentability, and bioactivity [12,29,71,145]. Structural elucidation typically requires advanced techniques such as NMR, FTIR, HPAEC-PAD (high-performance anion-exchange chromatography/pulsed amperometric detection), and size-exclusion chromatography [137,146]. Still, these are resource-intensive and not widely accessible. Moreover, differences in extraction, purification, and analysis protocols across laboratories hinder cross-study comparisons [100,110,147,148,149].

LAB strains also tend to produce mixtures of EPS, including both capsular and slime forms, as well as homo-EPS and hetero-EPS, which complicates structure–function correlations. This ambiguity impacts both product reproducibility and regulatory classification, since the designation of EPS as a fiber, prebiotic, or processing aid varies by jurisdiction and lacks consensus [19]. To overcome this, integrating genomics and metabolomics with carbohydrate structural analysis is essential for developing robust structure–function models [150]. Standardized reporting guidelines and EPS-specific databases would also support regulatory submissions and product development [13,14].

6.3. Extraction and Purification Challenges

When used as a purified ingredient, EPS must be extracted from complex fermentation matrices, often rich in proteins and other macromolecules. EPS recovery is challenging, particularly for capsular types that are tightly associated with cell surfaces [3,9,136,151]. Standard purification methods, such as centrifugation, ethanol precipitation, dialysis, and lyophilization, are not only resource-intensive but may also compromise the structure and functionality of EPSs [3,13,77]. Further complications arise from the co-precipitation of non-target proteins and polysaccharides, especially in dairy or cereal-based media. Proteolytic or enzymatic treatments can enhance selectivity but increase process complexity and cost [146,152,153]. The need for strain- and matrix-specific protocols limits standardization and scalability [132,134].

Emerging strategies, such as membrane separation, affinity chromatography, and the use of green solvents (including ionic liquids), offer potential but remain largely at the experimental stage, raising concerns regarding regulatory acceptance and food-grade status [36,100,154]. Developing mild, scalable, and cost-effective purification methods are essential for broader commercialization, especially in biofunctional applications. Integrating upstream fermentation control with automation and process analytical technologies can help overcome these bottlenecks [11,34,111,155].

6.4. Functional Variability in Food Systems

The functional performance of EPS varies widely depending on the food matrix and processing conditions. Factors such as pH, protein content, ionic strength, and shear forces significantly influence EPS behavior [6,7,14]. An EPS that improves viscosity and diminishes syneresis in yogurt, for example, may underperform in low-protein plant-based beverages or high-shear bakery systems [2,34,111,155]. EPS functionality may also be compromised during thermal processing or in acidic environments, leading to structural degradation or altered conformation [19]. Moreover, interactions with native polysaccharides or proteins can enhance or hinder performance, often unpredictably [100,110,148]. Overcoming this requires targeted optimization for each application, supported by rational screening, in situ fermentation strategies, and multivariate modeling to predict functional outcomes [12,137].

6.5. Regulatory Status, Labeling Barrier, and Limited Clinical Evidence for Health Claims

The regulatory landscape for EPS produced by LAB is currently fragmented. LAB strains used in food fermentations—such as Lactococcus lactis, Lactobacillus, Leuconostoc, and related genera—are widely considered Generally Recognized as Safe (GRAS) in the U.S., owing to historical use and expert consensus [156]. In the European Union, many of these species are also covered under EFSA’s Qualified Presumption of Safety (QPS) framework, a pre-assessed safety status for intentional use of microbial taxa in food and feed [157]. However, such designations relate to the organisms themselves and do not extend to their excreted or purified EPSs. When considering EPSs as isolated food additives, regulatory regimes demand independent safety evaluations. In the U.S., this typically means conducting a formal GRAS determination—whether self-affirmed or submitted through FDA notification—requiring rigorous evidence of identity, manufacturing process, exposure levels, and toxicology [158]. In the EU, purified EPS falls under the Novel Food Regulation (EU) 2015/2283. Approval of a novel food ingredient requires a comprehensive dossier that covers compositional analysis, proposed uses, intake estimates, nutritional effects, and safety data. No purified EPS from LAB is currently authorized under this regime. However, some microbial-derived polysaccharides like xanthan gum and β-glucans have been approved by EFSA [9]. EPSs produced in situ during traditional fermentations such as yogurt, sourdough, and kefir are accepted without further review, owing to the safety of LAB. Nonetheless, ex situ uses involving extraction or purification for other food applications face additional regulatory hurdles [159]. Future needs include well-designed toxicological studies and harmonized guidelines to assess the safety and functionality of EPS as novel food ingredients or additives.

Labeling requirements also vary. In situ EPS may be left undeclared as part of the fermentation process, whereas purified EPS must often be labeled as “fiber” or “hydrocolloid,” depending on the jurisdiction [100,160]. This poses challenges for clean-label claims and may cause consumer confusion. Moreover, regulatory approval for health claims is hindered by the limited availability of clinical data. Without well-characterized EPS and validated health endpoints, claims related to prebiotic or immunomodulatory effects are rarely accepted [143,161,162]. Harmonizing regulatory frameworks and establishing compositional and functional standards for EPSs will be key to enabling commercial uptake [14,153].

Although numerous in vitro and animal studies suggest health benefits, ranging from prebiotic activity to immunomodulation, human clinical trials are limited and sometimes lack specificity regarding EPS as the active component. The structural heterogeneity of EPS complicates standardization of test materials and dose–response studies [14,18,99]. With limited well-controlled clinical trials using structurally defined EPS and without regulatory recognition, consumer trust will remain limited [159,163,164]. More investigation efforts should focus on linking specific EPS structures and strains to quantifiable health outcomes, supported by multi-omics and biomarker validation.

6.6. Knowledge Gaps and Research Needs

Despite significant advances in the understanding of LAB-derived EPSs, including their structure, biosynthesis, and functional properties, critical gaps persist that hinder their full integration into food and health applications (Table 7). These gaps range from basic microbial physiology and metabolic regulation to applied formulation science and clinical validation, underscoring the need for interdisciplinary research that bridges microbiology, food science, biotechnology, and nutrition.

Table 7. Major knowledge gaps in exopolysaccharides (EPS) from lactic acid bacteria (LAB) research and proposed solutions.

Knowledge Gap	Description	Impact on EPS Application	Proposed Research/Technological Approaches	References
Structure–function relationships	Limited understanding of how the EPS molecular structure affects techno-functional and bioactive properties	Inconsistent functionality in foods and limited health applications	Integrate glycomics with functional assays; develop predictive models	[1,138,165]
Low and variable EPS yields	EPS production is strain- and condition-dependent; yields are often too low for industrial scale	Limits commercial feasibility and product consistency	Metabolic engineering, precision fermentation, adaptive evolution	[19,166]
Matrix interactions	Poor understanding of EPS behavior in complex food matrices, especially non-dairy	Reduced efficacy and unpredictable sensory effects	Systematic studies in diverse food matrices; multi-omics integration	[43,167]
Limited clinical evidence	Few human trials validating the health benefits of specific EPS	Restricts regulatory approval and consumer trust	Biomarker development	[1]
Regulatory fragmentation	Disparate regulations hinder global commercialization	Delays market entry and innovation	Harmonize standards and definitions; foster collaboration among agencies	[1,159,163]

Table 7. Major knowledge gaps in exopolysaccharides (EPS) from lactic acid bacteria (LAB) research and proposed solutions.

Knowledge Gap	Description	Impact on EPS Application	Proposed Research/Technological Approaches	References
Structure–function relationships	Limited understanding of how the EPS molecular structure affects techno-functional and bioactive properties	Inconsistent functionality in foods and limited health applications	Integrate glycomics with functional assays; develop predictive models	[1,138,165]
Low and variable EPS yields	EPS production is strain- and condition-dependent; yields are often too low for industrial scale	Limits commercial feasibility and product consistency	Metabolic engineering, precision fermentation, adaptive evolution	[19,166]
Matrix interactions	Poor understanding of EPS behavior in complex food matrices, especially non-dairy	Reduced efficacy and unpredictable sensory effects	Systematic studies in diverse food matrices; multi-omics integration	[43,167]
Limited clinical evidence	Few human trials validating the health benefits of specific EPS	Restricts regulatory approval and consumer trust	Biomarker development	[1]
Regulatory fragmentation	Disparate regulations hinder global commercialization	Delays market entry and innovation	Harmonize standards and definitions; foster collaboration among agencies	[1,159,163]

A major unresolved issue is the incomplete understanding of structure–function relationships. Although the structural diversity of EPS is well documented, there remains limited predictive capacity regarding how specific features, such as monosaccharide composition, glycosidic linkages, molecular weight, or branching patterns, govern techno-functional properties like viscosity and emulsification, or bioactive effects such as immunomodulation and prebiotic potential [100,110,148,149,168,169]. Advancing this area will require the integration of high-resolution carbohydrate analytics with functional assays, supported by modern glycomics and computational modeling tools that remain underutilized in EPS research.

Another key gap lies in the absence of high-throughput screening systems to identify and characterize LAB strains with desirable EPS profiles efficiently [11]. Most current workflows rely on laborious, small-scale fermentations followed by retrospective characterization. Innovative platforms leveraging biosensors, omics-based signatures, or phenotypic markers could accelerate the discovery and functional assessment of promising strains, particularly for applications targeting specific food environments [19].

From a formulation perspective, knowledge is still limited regarding the behavior of EPS in real food matrices. Most functional evaluations are conducted in simplified model systems that do not accurately capture the complexity of actual processing and storage conditions, such as pH shifts, heat treatments, or interactions with competing biopolymers. This is particularly true for plant-based and allergen-free systems, which are growing segments for clean-label functional ingredients [7,14,143,144]. Systematic studies are needed to characterize EPS performance in these contexts and identify formulation strategies that optimize their contribution to product quality and stability.

Bridging these gaps will require collaborative, cross-sector initiatives involving academia, industry, and regulatory agencies. Priorities include the development of standardized analytical and functional evaluation methods, the creation of open-access EPS structure–activity databases, and the promotion of interdisciplinary research programs with translational and clinical components. Only through such coordinated efforts can EPSs move beyond experimental promise to become reliable, scalable tools for food innovation and personalized health solutions.

7. Future Perspectives

The multifunctionality of EPSs derived from LAB positions them as valuable tools in addressing current trends in food innovation. As consumer preferences increasingly favor natural, clean-label, health-promoting, and personalized products, EPSs stand out for their ability to enhance texture, improve shelf life, support gut health, and modulate immune responses [3,11,77,130,170].

7.1. Precision Fermentation and Strain Engineering

Recent advances in synthetic biology, metabolic engineering, and systems biology offer promising avenues to overcome persistent limitations in EPS production (Table 8). Traditional fermentation processes often yield low and variable EPS quantities, which hinder industrial applications. However, by using genome sequencing, CRISPR-Cas tools, and transcriptomic data, researchers can now accurately manipulate genes involved in EPS biosynthesis—such as those encoding glycosyltransferases or controlling chain length—to enhance both yield and functionality [4,5,18,19,166].

Table 8. Biotechnological strategies for enhancing exopolysaccharides (EPS) from lactic acid bacteria production and functionality.

Strategy/Technology	Key Features	Advantages	Challenges	References
Metabolic engineering	Genetic modification of EPS biosynthetic pathways (e.g., glycosyltransferases)	Improved yield; customized EPS structure	Regulatory hurdles; strain stability	[19,166]
Adaptive laboratory evolution	Non-GMO selection under stress to improve traits	Regulatory-friendly, natural adaptations	Slow process; unpredictable results	[1]
CRISPR-Cas genome editing	Precise gene knock-ins/knockouts without off-targets	Targeted control, high specificity	Consumer perception; GMO classification issues	[171,172,173]
Synthetic biology and modular design	Integration of novel or heterologous EPS gene clusters	Novel EPS production; programmable structures	Requires deep pathway knowledge	[166,171]
Precision fermentation platforms	Real-time control of culture conditions and feeding strategies	Scalable, consistent production; structural tuning	High capital and technical complexity	[34,111,155,174,175]

Table 8. Biotechnological strategies for enhancing exopolysaccharides (EPS) from lactic acid bacteria production and functionality.

Strategy/Technology	Key Features	Advantages	Challenges	References
Metabolic engineering	Genetic modification of EPS biosynthetic pathways (e.g., glycosyltransferases)	Improved yield; customized EPS structure	Regulatory hurdles; strain stability	[19,166]
Adaptive laboratory evolution	Non-GMO selection under stress to improve traits	Regulatory-friendly, natural adaptations	Slow process; unpredictable results	[1]
CRISPR-Cas genome editing	Precise gene knock-ins/knockouts without off-targets	Targeted control, high specificity	Consumer perception; GMO classification issues	[171,172,173]
Synthetic biology and modular design	Integration of novel or heterologous EPS gene clusters	Novel EPS production; programmable structures	Requires deep pathway knowledge	[166,171]
Precision fermentation platforms	Real-time control of culture conditions and feeding strategies	Scalable, consistent production; structural tuning	High capital and technical complexity	[34,111,155,174,175]

These technologies enable the development of customized LAB strains capable of producing EPS with specific structural features, such as specific molecular weights or glycosidic linkages, linked to functional traits like viscosity or prebiotic benefits. Synthetic biology approaches even allow for the modular insertion of biosynthetic pathways from other organisms, enabling the production of novel EPSs with enhanced bioactivity or rheological properties [171,172,173].

Precision fermentation platforms, featuring real-time bioreactor control, fed-batch optimization, and online metabolic monitoring, further contribute to reproducible and scalable EPS production for specific food or health applications [34,111,155,174,175]. Nevertheless, regulatory and consumer concerns about GMOs necessitate exploring non-GMO strategies, such as adaptive laboratory evolution or CRISPR techniques, that avoid the integration of foreign DNA [100,110,148,149]. Ultimately, integrating multi-omics data with machine learning will accelerate the rational design of LAB strains for EPS production, thereby facilitating the development of customized solutions for various food matrices and health applications.

7.2. Multi-Omics for Structure–Function Insights

Understanding how the EPS structure relates to functionality remains a critical barrier to their broader application. Multi-omics approaches, including genomics, transcriptomics, proteomics, metabolomics, and glycomics, are providing valuable insights into the biosynthetic mechanisms of EPS and their techno-functional and bioactive profiles, as shown in

Table 9. Multi-omics tools for decoding exopolysaccharides (EPS) from lactic acid bacteria (LAB) structure–function relationships.

Omics Discipline	Primary Focus	Contribution to EPS Research	Limitations
Genomics	Identification of EPS gene clusters (eps operons)	Enables strain screening and rational pathway targeting	Does not capture regulation or dynamic behavior
Transcriptomics	Expression profiling under specific conditions	Reveals regulatory cues and fermentation triggers	Snapshot view: affected by culture conditions
Proteomics	Detection of biosynthetic enzymes and transporters	Connects genotype to active biosynthetic machinery	Challenges in analyzing membrane-bound proteins
Metabolomics	Monitoring of sugar fluxes and fermentation by-products	Identifies metabolic bottlenecks and EPS precursors	Requires careful interpretation; complex sample handling
Glycomics	EPS structure mapping (e.g., linkages, branching)	Elucidates functional motifs influencing techno- and bioactivity	Technically complex; limited standardization

[1,2].

Genomic and transcriptomic analyses enable the identification of EPS biosynthesis gene clusters and the regulatory networks that control their expression under various environmental conditions [19,140]. Proteomics and metabolomics complement these data by revealing the enzymes and metabolic pathways involved in sugar transport and precursor synthesis, as well as the fluxes that determine EPS yield and quality [166].

Glycomics, although underutilized, is essential for the detailed characterization of EPS structure, linkage type, branching, monosaccharide composition, and molecular size, all of which impact functional properties such as solubility, viscosity, and fermentability [138,165]. Integrating these datasets enables the development of predictive models for EPS functionality, guiding strain selection and fermentation optimization [166,176]. This system-level understanding is key for enhancing product consistency and substantiating regulatory claims, particularly for EPS with health-promoting properties.

7.3. Functional EPS in Symbiotic and Personalized Nutrition

In personalized nutrition, EPS-producing LAB strains are increasingly recognized for their dual function as probiotics and producers of bioactive polysaccharides (

Table 10. Functional roles of exopolysaccharides (EPS) from lactic acid bacteria in personalized and symbiotic nutrition.

Functional Role of EPS	Mechanism/Benefit	Target Population/Use Case
Prebiotic activity	Selective stimulation of beneficial gut microbiota	Individuals with dysbiosis, the elderly
Immunomodulation	Modulation of immune signaling pathways	Metabolic syndrome, Inflammatory Bowel Disease, and aging
Protective matrix for probiotics	Enhances viability and stability during digestion	Symbiotic formulations in functional foods
Synergistic formulations	Combination with fibers/polyphenols for precision effects	Personalized nutrition interventions

). EPSs can boost the survival and activity of probiotics while acting as prebiotics that selectively promote beneficial gut microbes [13,14,36].

Their immunomodulatory, anti-inflammatory, and gut barrier-supporting properties make EPSs promising candidates for targeted interventions in aging, metabolic disorders, and gastrointestinal diseases [8,177,178]. However, the strain- and structure-dependent nature of these effects requires careful matching of EPS profiles to specific health goals and host microbiomes [100,110,148,149].

Multi-omics tools facilitate the identification of EPS types that support key microbial groups or modulate immune signaling [179,180]. These insights can guide the development of personalized symbiotic products that combine EPS with specific probiotics, fibers, or bioactive compounds tailored to individual needs [181]. To advance this field, solid clinical studies are necessary to establish a link between EPS structures and proven health benefits. Transparent communication and education will also be crucial for fostering consumer trust and promoting the adoption of precision nutrition strategies.

7.4. Regulatory Harmonization and Clean-Label Strategies

For EPSs to gain widespread commercial use, clear regulations and harmonization are crucial. Currently, EPSs are classified differently as food additives, novel foods, or dietary fibers, depending on the jurisdiction, which complicates approval and labeling [1,159,163,182]. Differences in definitions, safety standards, and health claim criteria create challenges for global commercialization. In the EU, EFSA demands extensive safety and efficacy data for functional claims, whereas the U.S. FDA uses its criteria for dietary fiber classification and GRAS status. Additionally, the absence of standardized protocols for characterizing EPS—encompassing purity, structure, and bioactivity—complicates regulatory approval and affects market consistency [100,110,148,149].

Simultaneously, the clean-label movement presents both opportunities and challenges. EPSs align with clean-label principles as natural fermentation products, yet the use of processing aids, purification steps, or genetically modified strains can raise concerns [183,184]. To mitigate this, in situ production strategies, where EPSs are synthesized directly during food fermentation, are gaining traction as a low-intervention, consumer-friendly alternative [7,14,143,144]. Moving forward, coordinated efforts across academia, industry, and regulatory bodies will be essential to establish shared standards, facilitate labeling, and support the adoption of EPS in both conventional and emerging markets.

8. Final Remarks

EPSs produced by LAB represent a versatile and underutilized class of natural biopolymers with significant potential in modern food systems. Whether added as purified ingredients or produced in situ during fermentation, EPSs can enhance the texture, viscosity, gel formation, stability, and shelf life of foods while simultaneously contributing to health-promoting functions such as immune modulation, antioxidant activity, and dietary fiber. As the food industry evolves toward more sustainable, functional, and personalized products, EPSs offer a unique opportunity to bridge technological innovation with human health, positioning them as key ingredients in the next generation of fermented and functional foods.

The application of ex situ EPS as a food ingredient during the process allows the addition of specific amounts at specific times; thus, it is more controllable than in situ production, which depends on fermentation conditions. The addition of ex situ EPS as an ingredient can be utilized in unfermented foods, conferring the EPS properties without changes in food physicochemical and sensory properties associated with fermentation. However, as an ingredient, ex situ EPS should be declared as an additive that negatively impacts the product and may conflict with clean-label consumer preferences. In addition, production costs increase due to isolation and purification processes.

Despite their promise, broader industrial application of EPSs remains limited by challenges including low yields, structural heterogeneity, regulatory ambiguity, and limited clinical evidence. Nevertheless, advances in strain development, multi-omics, and bioprocess optimization are rapidly addressing these barriers.