Abstract
Probiotics, as beneficial microorganisms, are critical to host health. However, their viability is often compromised during processing, storage, and gastrointestinal transit, significantly compromises their colonization efficacy and therapeutic potential. Polysaccharides have emerged as pivotal materials for probiotic encapsulation due to their excellent biocompatibility, biodegradability, and unique functional properties. This review systematically examines traditional polysaccharide-based encapsulation technologies, such as embedding and coating techniques, highlighting limitations of single-polysaccharide systems, including excessive porosity, inadequate mechanical strength, suboptimal encapsulation efficiency, and poor targeted release precision. In contrast to previous research focused on single polysaccharides, this review focuses on composite polysaccharide encapsulation systems, particularly polysaccharide-polysaccharide hybrids and polysaccharide-protein complexes, which effectively address the limitations of single-polysaccharide systems in probiotic encapsulation while significantly enhancing encapsulation performance. Furthermore, it investigates the advantages of prebiotic incorporation in promoting probiotic proliferation and suppressing pathogenic microorganisms, providing novel optimization strategies for delivery systems. These findings establish critical theoretical and technical foundations for translating these advancements into functional foods and oral pharmaceutical formulations.
Keywords: Probiotics, Encapsulation, Biopolymer, Polysaccharide-protein complexes, Composite delivery systems, Prebiotic synergy
Graphical abstract
Highlights
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Composite polysaccharides enhance probiotic encapsulation efficiency.
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Polysaccharide-protein hybrids improve thermal stability and acid resistance.
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Prebiotic co-encapsulation boosts probiotic viability and suppresses pathogens.
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Composite polysaccharides are targeted for colon release by pH/enzyme response.
1. Introduction
The World Health Organization (WHO) and the Food and Agriculture Organization of the United Nations (FAO) have clearly defined probiotics as live microorganisms (Nezamdoost-Sani et al., 2023). These microorganisms are widely colonized in the intestinal ecosystems of humans and animals. Based on their morphological characteristics, physiological and metabolic mechanisms, and biochemical properties, they can mainly be classified into two categories: bacterial genera (e.g., Lactobacillus, Bifidobacterium) and fungal species (e.g., Saccharomyces spp.). Research indicates that the intestinal microbiota, as the “second genome” of the human body, plays a core role in maintaining the host's health homeostasis and regulating pathological processes (Mousavi Ghahfarrokhi et al., 2024). When consumed in sufficient quantities (≥106–107 CFU/g) (Nezamdoost-Sani et al., 2023), probiotics can enhance intestinal barrier function and restore microbiota ecological balance by producing short-chain fatty acids, bacteriocins, and other bioactive substances under specific microenvironmental conditions. Due to their beneficial properties, the term “beneficial microorganisms” is commonly used (Vivek et al., 2023). Notably, excessive intake may lead to dysbiosis.
In recent years, the significance of probiotics in health sciences has become increasingly prominent, with their multifaceted physiological functions extensively studied and validated. Clinical studies demonstrate that probiotics serve not only as effective therapeutic adjuvants but also play crucial roles in preventing and managing neurological disorders, allergic diseases, and malignancies such as colorectal cancer (Chen et al., 2024, Chen et al., 2024, Chen et al., 2024). Of particular interest is the bidirectional regulatory mechanism mediated by the microbiota-gut-brain axis. Probiotics have been shown to remodel gut microbiota composition, modulate signaling pathways along the gut-brain axis, and influence neurological functions through immune system regulation, thereby demonstrating unique advantages in the prevention and treatment of neuropsychiatric disorders (Dey et al., 2020). In metabolic regulation, probiotics have been confirmed to exert significant lipid metabolism-modulating functions, a critical attribute for maintaining systemic metabolic homeostasis. Notably, in preventing and intervening in metabolic diseases like diabetes and obesity, probiotics exhibit substantial clinical potential through structural modulation of the gut microbiome (Wlodarska et al., 2015). Furthermore, the antimicrobial activity of probiotics not only effectively inhibits pathogen proliferation to maintain intestinal microecological equilibrium, but their metabolites can synergistically exert extensive physiological regulatory effects (Zendeboodi et al., 2020). With advancing scientific exploration, probiotics have emerged as critical biotherapeutic agents in human health preservation, with increasingly sophisticated clinical applications (Fentie et al., 2024), underscoring their undeniable importance in maintaining human health and well-being. Despite their therapeutic potential, practical application is hindered by significant viability challenges during oral delivery.
Oral administration represents a convenient daily supplementation approach. However, probiotic viability is susceptible to external environmental factors and internal physiological challenges. Externally, processing conditions, storage parameters, and interactions with food components can compromise probiotics survival (Chen et al., 2024; Zhao et al., 2024). Internally, the gastrointestinal tract presents multiple hostile barriers including oral cavity lysozymes, gastric mucosal-bicarbonate barriers, intestinal digestive enzymes (Lou et al., 2023), strongly acidic gastric juice, bile salts, and pancreatic secretions. To achieve targeted delivery to the colon, probiotics must survive these harsh conditions during transit, which significantly reduces their colonization capacity and metabolic activity below the therapeutic threshold required for health benefits. Fig. 1 depicts external and internal factors affecting probiotic viability and delivery efficacy (Fig. 1).
Fig. 1.
Factors influencing probiotic viability.
Maintaining adequate viable cell counts remains critical for probiotics efficacy. The FAO/WHO mandates a minimum viable count of 106 colony-forming units (CFU) per gram in probiotics products to ensure functional benefits (Gao et al., 2023). Current research priorities therefore focus on developing innovative strategies to enhance probiotic viability during product shelf-life and gastrointestinal transit while ensuring sufficient colonic colonization density for therapeutic effectiveness.
In recent years, numerous researchers developed encapsulation systems to protect probiotics from inactivation during processing, storage, and digestion, while enabling their targeted delivery to the colon and other critical sites to exert specific functions. Encapsulation was demonstrated to enhance probiotics viability (Azeem et al., 2023). To construct such probiotics delivery models, researchers employed various encapsulation techniques (Luo et al., 2022; Yao et al., 2020), including extrusion, emulsification, electrospinning, and electrohydrodynamic processes (Wang et al., 2024, Wang et al., 2024, Wang et al., 2024; Xie et al., 2023, Xie et al., 2023; Xu et al., 2023, Xu et al., 2023), embedding probiotics within protective matrices. These strategies aim to ensure probiotic functionality at target sites by mitigating environmental stressors and optimizing controlled release properties.
If encapsulation technology serves as the fortress wall in probiotics protection and functional implementation, then encapsulation materials constitute the cornerstone of this fortress, whose physicochemical properties directly determine protective efficacy and functional performance. High-quality, compatible materials can establish seamless integration with probiotics surfaces through electrostatic interactions, covalent/non-covalent bonding, thereby forming stable protective barriers. These barriers effectively maintaine probiotics viability and functional integrity under food processing stresses and gastrointestinal challenges (Zhao et al., 2024). Furthermore, diverse encapsulation materials can confer unique functionalities to probiotics, including targeted delivery, stimuli-responsive behavior, and enhanced mucosal adhesion (Xu et al., 2022). Such innovations expand probiotics applications across therapeutic formulations and functional foods. Therefore, in advancing the development of probiotics encapsulation technologies, the in-depth investigation and strategic selection of encapsulation materials have constituted a critical and indispensable component. In recent years, food-grade biopolymers have been increasingly employed to construct delivery systems for bioactive compounds, demonstrating exceptional protective efficacy. When selecting encapsulation materials, critical considerations include oxygen barrier properties, acid resistance in gastric environments, bile salt tolerance, encapsulation efficiency, and colonic release characteristics. These parameters collectively determine the material's ability to maintain probiotics viability while ensuring targeted payload delivery.
A diverse array of biomaterials is utilized for probiotics encapsulation, primarily categorized into protein-based biopolymers, lipid-derived matrices, and polysaccharides (Phùng et al., 2025). These natural food-grade polymers are recognized as ideal materials for probiotics delivery systems due to their superior safety profiles, biodegradability, and biointerface self-assembly capabilities. Notably, polysaccharides have emerged as highly effective encapsulation materials, leveraging their diverse sources, non-toxic nature, biocompatibility, environmentally benign degradation properties, and functional versatility (Gheorghita et al., 2021). For example, sodium alginate, as a natural polysaccharide polymer, exhibits excellent gelation properties, stability, and water retention capacity. (Khoshdouni Farahani et al., 2022a; Khoshdouni Farahani et al., 2024c). It rapidly forms hydrogels through ionic crosslinking under mild conditions, enabling efficient production of encapsulation beads. Sodium alginate has been widely employed as an encapsulation material in extensive research (Khoshdouni Farahani et al., 2022b, 2023a). Despite the advantages of polysaccharides in probiotics encapsulation, these materials exhibit inherent limitations during the encapsulation process, including excessive porosity, low mechanical strength, and potential viability damage to probiotics (Jadav et al., 2023; Wang et al., 2022). Consequently, current research has prioritized the incorporation of supplementary biopolymers to enhance their functional performance through composite material engineering.
This review focuses on polysaccharide-based biopolymer encapsulation of probiotics. Initially, it systematically examines the application of polysaccharide materials and diverse technological methodologies for probiotic encapsulation, incorporating multidimensional analyses spanning material properties to technical implementations. Subsequently, critical inherent limitations of single-polysaccharide encapsulation systems were identified, including excessive porosity, inadequate mechanical strength, and compromised stability under extreme conditions. Building upon these constraints, the review extensively investigates composite polysaccharide encapsulation strategies, with particular emphasis on the operational mechanisms of polysaccharide-polysaccharide hybrids and polysaccharide-protein complexes. Ultimately, empirical analyses demonstrate functional enhancements achieved through multi-polysaccharide encapsulation approaches, such as improved encapsulation efficiency and enhanced targeted release precision, while delineating future developmental trajectories to guide subsequent research endeavors.
2. Food-grade delivery systems for probiotics
Current food-grade delivery systems for probiotics predominantly employ two distinct encapsulation technological designs.
Traditional encapsulation technologies are classified into embedding techniques and coating techniques. Fig. 2 illustrates the two probiotic encapsulation techniques, delineating the morphological features of the encapsulation carriers, technical classifications and mechanistic actions (Fig. 2). The embedding approach immobilizes probiotics within carrier matrices, encapsulating them in a tessellated architectural configuration biomimetically inspired by natural “cocoon” and “spiderweb” structural paradigms. “Cocoon"-inspired multicellular encapsulation systems include emulsions, liposomes, nanoparticles, microcapsules, and microspheres (Zang et al., 2025), all featuring protective shell architectures that immobilize probiotics. In contrast, the “spiderweb” methodology employs network-like configurations to safeguard viable bacteria from external perturbations. These systems typically incorporate gel-based matrices to effectively enhance and maintain probiotic viability. Spiderweb-inspired systems primarily consist of hydrogels, oleogels, microgels, and hybrid gels (bigels) (Gao et al., 2023; Yu et al., 2022). Current research predominantly concentrates on the microencapsulation of probiotics through embedding technologies, necessitating specialized processing operations such as spray-drying, extrusion, lyophilization, and emulsification (Xu et al., 2022). Studies have found that during spray-drying microcapsule preparation, increasing the glass transition temperature (Tg) of the embedding carrier improves thermal stability, thereby better protecting embedded active components from environmental damage (Khoshdouni Farahani et al., 2022c; Khoshdouni Farahani et al., 2024b). Polysaccharides with high Tg, such as gum arabic, maintain a glassy state during spray-drying, suppressing molecular mobility to preserve probiotic viability (Halahlah et al., 2023). Gum arabic, a complex heteropolysaccharide, demonstrates unique molecular architecture that facilitates synergistic interactions with complementary components, thereby modifying system physicochemical properties and enhancing Tg. In investigations evaluating plant-derived polysaccharide-pea protein isolate (PPI) composite carriers for spray-dried Lactobacillus casei powders, gum arabic -containing formulations achieved significantly increased Tg values up to 189.2 °C. Through spray-drying processing, this elevated Tg maintains powders in a glassy state during storage, substantially reducing molecular mobility to inhibit degradation pathways including lipid oxidation. Such physicochemical stabilization preserves powder integrity by preventing structural alterations and microbial habitat disruption, ultimately ensuring probiotics viability (Sharma et al., 2025). Conventional coating technology involves encapsulating probiotics cells with single or multiple polymeric layers, achieving robust integration between functional materials and probiotics through strategic exploitation of surface characteristics such as electrical charge and adhesion factors. For instance, electrostatic deposition enables layer-by-layer (LbL) assembly of oppositely charged polyelectrolytes onto probiotics surfaces (Luo et al., 2020), while covalent bonding between coating materials and surface functional groups establishes chemically stable outer coatings. Positioned as the outermost cellular barrier, these coatings enhance probiotics resistance to physicochemical stresses and improve mucoadhesive properties. Furthermore, they confer specialized functionalities, thereby creating novel application paradigms for probiotics delivery systems.
Fig. 2.
Two encapsulation technologies for food-grade probiotic delivery systems.
These two technologies are often synergistically employed to enhance probiotic protection. Coating techniques significantly optimize the performance of carrier matrices fabricated via embedding technologies, utilizing edible biopolymers like polysaccharides and proteins (Yao et al., 2020). These biopolymers form cohesive film matrices through hydrogen bonding, electrostatic interactions, hydrophobic associations, and/or non-covalent crosslinking. Critical process parameters including pH (modulating biopolymer charge states), temperature (influencing molecular mobility and interaction kinetics), ionic strength (regulating electrostatic shielding or enhancement), along with food-grade solvents, surfactants, and enzymes, collectively govern coating formation, structural integrity, and functional properties. For instance, LbL self-assembly technology has been applied to coat pre-fabricated microcapsules. In a study on the co-encapsulation of probiotics and polyphenols, microcapsules were successfully fabricated through LbL complexation of chitosan-whey protein isolate with pectin, sodium alginate, and Fuzhuan brick tea polysaccharides (FBTP). These microcapsules exhibit enhanced performance in probiotics protection, antioxidant capacity, anti-colitis efficacy, and yogurt application, with differential enhancement effects among various polysaccharides (Sun et al., 2024, Sun et al., 2024). Similarly, Liu et al. (2023) developed multilayered microcapsules by alternating deposition of zein nanoparticles and pectin for encapsulating Lactobacillus plantarum 550. Their research demonstrated that the outer matrix composition and coating layer number critically influence cellular viability during heating, storage, and gastrointestinal digestion, providing theoretical insights for probiotics encapsulation optimization (Liu et al., 2023). Furthermore, researchers effectively immobilized Lactobacillus rhamnosus 6133 using sodium alginate, hyaluronic acid, and gelatin combined with LbL technology. Their findings revealed that insufficient coating layers led to cell exposure due to alginate dissolution, whereas increased layers counteracted this issue through gelatin-hyaluronic acid protective synergy. The multilayered microcapsules significantly enhanced antioxidant properties and cellular extract viability under simulated gastric fluid (SGF) conditions (Wang et al., 2024). LbL technique encapsulates probiotics within multilayered polymeric membranes through electrostatic interactions between oppositely charged polymers. This approach enhances resistance to acidic and bile salt insults, promotes intestinal adhesion and proliferation, and elevates in vivo survivability. Compatible with diverse charged polyelectrolytes, proteins, polysaccharides, and probiotic strains, this technology establishes a robust foundation for advancing human health.
3. Advantages of polysaccharide-based encapsulation materials
The selection of materials for probiotics encapsulation necessitates a comprehensive evaluation of multifaceted factors. Primary consideration must be given to material safety and compliance with regulatory standards established by authorities such as the European Food Safety Authority (EFSA) and U.S. Food and Drug Administration (FDA), which constitute fundamental prerequisites for market authorization and consumer safety (Amiri et al., 2024). Additional critical parameters include encapsulation methodology, encapsulation efficiency, bacterial loading capacity, storage stability, controlled-release properties, and regulatory compliance - all essential for ensuring optimal encapsulation performance and product quality (Xu et al., 2024, Xu et al., 2024). Consequently, material selection for delivery systems must align with both probiotics requirements and functional objectives of final products. Commonly employed food-grade materials predominantly comprise polysaccharides, proteins, and lipids (Amiri et al., 2024; Xie et al., 2023).
Compared to proteins, polysaccharides possess distinct advantages. They exhibit diverse sourcing accessibility, obtainable from terrestrial plants to marine organisms (Sharma et al., 2023). While protein functionality critically depends on conformational integrity, environmental factors including temperature, pH, ionic strength, and enzymatic activity can disrupt their structural stability. For instance, elevated temperatures induce intensified molecular motion that disrupts protein spatial conformations, leading to denaturation (Ma et al., 2024, Ma et al., 2024). In contrast, polysaccharides demonstrate superior thermal stability and maintain structural integrity under extreme pH conditions, making them particularly suitable for gastric delivery systems requiring acid resistance.
Unlike lipids, polysaccharides can form typical hydrogels compatible with aqueous food systems, whereas lipids are predominantly employed in oil-based matrices. Although lipid architectures facilitate bioactive compound transport, their thermal sensitivity compromises probiotics viability retention during processing and restricts high-temperature food applications. Furthermore, some lipid systems require cholesterol for stabilization, raising potential health concerns (Amiri et al., 2024). Concurrently, polysaccharides inherently lack fat content while their molecular characteristics confer functional advantages including viscosity modulation, gel network formation, and enhanced water-holding capacity, thus enabling their utilization as fat replacers (Bourouis et al., 2023).
In summary, polysaccharides emerge as ideal candidates for constructing delivery systems due to their inherent biocompatibility, biodegradability, and multifunctionality, making them particularly suitable for diverse applications including probiotics delivery. Fig. 3 summarizes common polysaccharide and protein polymers used for encapsulating probiotics (Fig. 3). These natural polymers can be engineered into tailored delivery systems with optimized mechanical properties, environmental stability, and stimuli-responsive behavior, such as pH sensitivity, thus enabling precise adaptation to varied food matrices. Although these remarkable advantages render polysaccharides promising encapsulation materials, a comprehensive evaluation necessitates an objective consideration of their inherent limitations. Consequently, emerging research focuses on composite material strategies that combine polysaccharides with complementary ingredients, harnessing synergistic advantages while addressing individual material constraints, thereby advancing the development of high-performance composite architectures for application-specific requirements.
Fig. 3.
Polysaccharide- and protein-based biopolymers for probiotic encapsulation.
4. Single-polysaccharide polymer encapsulation of probiotics
This section provides an overview of polysaccharides commonly employed in the fabrication of delivery systems, including sodium alginate, chitosan, cellulose, starch, pectin, and others. Furthermore, it critically examines the distinct encapsulation performance of these polysaccharides-specifically their advantages and limitations in probiotics protection and delivery-based on their unique physicochemical properties and interaction mechanisms. To offer a comparative analysis, Table 1 summarizes the processing parameters, encapsulation efficiency, and viability retention rates of probiotics encapsulated using individual polysaccharides under diverse environmental conditions. The data collectively demonstrate the inherent limitations of single-polysaccharide-based probiotic encapsulation strategies.
Table 1.
Encapsulation efficiency of single-polysaccharide systems for probiotics.
| Strain | Encapsulation material | Encapsulation process | Encapsulation efficiency (%) | Simulates gastrointestinal stability |
Storage stability (SR) | Other stability (SR) | References | |
|---|---|---|---|---|---|---|---|---|
| Simulated gastric fluid (SR) | Simulated intestinal fluid (SR) | |||||||
| Pediococcus pentose | Sodium alginate | Gel-embedded + In situ biofilm culture | 50.36 % (pH 2.01, 2 h) | 74.28 % (pH 8.02, 4 h) | 88.6 % (14 d, 4 °C) | 1. 36.04 % (vacuum freeze-drying, 72 h) | Mgomi et al. (2024) | |
| 2. 30.7 % (acidic stress, 2 h) | ||||||||
| 3. 67.7 % (5 % NaCl, 4 h) | ||||||||
| Lactobacillus pentose | Chitosan | Layer-by-Layer self-assembled microcapsules | 49.18 % (pH 1.5, 2 h) | 72.78 % (pH 6.8, 3 h) | 87.67 % (30 d, 4 °C) | Wang et al., 2019 | ||
| Lactobacillus plantarum ATCC 13643 | Carboxymethyl cellulose | Cross-linked carboxymethyl cellulose | 74.2 | 41.2 % (pH 2, 2 h) | 43.5 % (30 d, 4 °C) | Dafe et al. (2017a) | ||
| Lactobacillus plantarum ACCC 11095 | Lotus seed starch | Crosslinker crosslinking + Freeze-drying | 48.8 | 73.56 % (pH 2, 1 h) | 71.87 % (pH 7, 2 h) | Li et al. (2024) | ||
| Lactobacillus casei DSM 20011 | Olive Flour (OLF) | Composite emulsification-crosslinking | 52.74 % (pH 2.0, 2 h) | 84.84 % (pH 7.0, 2 h) | 16.84 % (28 d, −24 °C) | Karkar et al. (2024) | ||
| Lactobacillus acidophilus DSM 20079 | Olive Flour (OLF) | Composite emulsification-crosslinking | 36.28 % (pH 2.0, 2 h) | 83.62 % (pH 7.0, 2 h) | 21.66 % (28 d, −24 °C) | Karkar et al. (2024) | ||
| Lactobacillus casei | High amylose-starch | Zinc chloride crosslinking | 88.8 | 95.74 % (pH 2.0, 2 h) | 71.28 % (pH 7.5, 3 h) | Zhang et al. (2021) | ||
| Lactobacillus rhamnosus ATCC 53103 | Citrus Pectin LC950 | Calcium chloride crosslinking + Freeze-drying | 99.9 | 97.10 % (pH 1.6, 2 h) | 94.78 % (pH 6.5, 2 h) | 10.2 Log CFU/mL (28 d) | 99.90 % (pH 2, 2 h) | Li et al., 2016 |
| Lactobacillus plantarum, GDMCC 1.140 | Pectin (Citrus) | In situ biofilm culture | 65.1 | 65.15 % (pH 2.0, 2 h) | 88.42 % (pH 6.8, 4 h) | 1. 99.68 % (45 d, −20 °C) | Chen et al. (2024) | |
| 2. 98.06 % (45 d, 4 °C) | ||||||||
| 3. 77.42 % (45 d, 25 °C) | ||||||||
| Lactobacillus rhamnosus | Pectin (Citrus) | In situ biofilm culture | 66.0 | 79.26 % (pH 2.0, 2 h) | 85.74 % (pH 6.8, 2 h) | 1. 99.89 % (45 d, −20 °C) | Chen et al. (2024) | |
| 2. 98.03 % (45 d, 4 °C) | ||||||||
| 3. 82.95 % (45 d, 25 °C) | ||||||||
| Bifidobacterium breve CICC 6182 | Low Methoxy Pectin | Ionic crosslinking + Freeze-drying | 91.04 % (pH 2, 2 h) | 83.04 % (pH 7.4, 2 h) | 1. 91.00 % (13 weeks, −20 °C) | Li et al. (2019) | ||
| 2. 86.40 % (13 weeks, 4 °C) | ||||||||
| 3. 68.49 % (13 weeks, 20 °C) | ||||||||
SR: Survival rate.
4.1. Sodium alginate
Among natural polymers, sodium alginate emerges as the material of choice. This anionic polysaccharide, extracted from brown algae or bacterial species such as Pseudomonas and Azotobacter, is structurally composed of β-D-mannuronic acid (M) and α-L-guluronic acid (G) units. Its distinctive architecture endows it with exceptional gelation capacity, biocompatibility, and biodegradability (Wang et al., 2023). Sodium alginate undergoes instantaneous gelation in the presence of polyvalent cations, forming particulate structures through ionic interactions (Panichikkal et al., 2021). As the most prevalent alginate derivative, sodium alginate exhibits intrinsic biodegradability and environmental non-toxicity (Panichikkal et al., 2021). Its gel-forming mechanism, termed ionic gelation, arises from interactions with divalent cations (Ca2+, Zn2+, Mn2+, Sr2+), fundamentally governed by ionotropic crosslinking mechanisms. Sodium alginate undergoes rapid ion exchange upon exposure to calcium ions, forming a thermally irreversible hydrogel through instantaneous crosslinking (Khoshdouni Farahani et al., 2023b). This process involves cooperative cation binding to the G-blocks of the alginate chain, creating a three-dimensional “egg-box” lattice structure that ensures mechanical stability while retaining bioactive compounds. The “egg-box” interaction constructs a hydrogel matrix with a three-dimensional network architecture, establishing the foundation for sodium alginate's food industry applications (Razavi et al., 2021) and, more critically, significantly enhancing probiotic resilience against environmental stressors. The cation-mediated “egg-box” complexation (Cheng et al., 2020) substantially expands its functional versatility. During this process, liquid droplets progressively undergo gelation, transitioning into microcapsules that encapsulate probiotics within the alginate matrix. Capitalizing on its thermal tolerance and lyoprotective properties (freeze-drying protective), Mgomi et al. (2024) engineered calcium alginate gel beads (CAGB) for encapsulating Pediococcus pentosaceus. Under refrigerated storage conditions, the CAGB-biofilm-situ (CAGB-bio-situ) formulation achieved 88.6 % viability retention, demonstrating statistically significant improvement compared to non-encapsulated counterparts. Fourier-transform infrared (FTIR) spectroscopy analysis confirmed the absence of novel chemical bond formation between sodium alginate and bacterial cells, elucidating the mechanistic stability of the CAGB encapsulation system (Mgomi et al., 2024).
Notably, the presence of carboxyl groups in sodium alginate confers pH-responsive release behavior to alginate hydrogels (Mulia et al., 2020). In acidic environments (pH 1.5–3.0), the conversion of carboxylate ions (-COO-) groups to carboxyl groups induces polymer chain contraction and pore size reduction through protonation. This structural adaptation effectively shields probiotics from gastric acid degradation by creating a stabilized microenvironment. Given the inherent vulnerability of probiotics to low-pH conditions, such contraction behavior significantly minimizes acid-induced cellular damage. Conversely, under alkaline conditions (intestinal pH > 6.5), deprotonation regenerates -COO- moieties. Intensified electrostatic repulsion triggers hydrogel swelling via molecular chain expansion and pore size enlargement. This pH-dependent swelling facilitates controlled probiotic release in intestinal environments, enabling targeted colonization and functionality (Mulia et al., 2020; Xu et al., 2022).
While sodium alginate functions as an effective probiotics carrier by enhancing viability retention, it exhibits inherent limitations including inadequate gastric protection under low-pH, suboptimal encapsulation efficiency due to microsphere porosity, premature degradation in acidic environments, and compromised intestinal transit efficacy (Wang et al., 2022). Studies demonstrate that sodium alginate hydrogels achieve only 50.36 ± 1.97 % viability in SGF, as evidenced by the CAGB system (Mgomi et al., 2024). To address these challenges, researchers are developing composite systems combining alginate with complementary polymers to improve gastric stability and achieve synchronized protection-release kinetics. Alginate can form polyelectrolyte complexes with oppositely charged polymers via electrostatic interactions, providing superior acid-resistant shielding. Notable examples include chitosan and protein composites. Specifically, alginate-chitosan complexes generate mechanically robust hydrogels with broad biomedical applicability (Razavi et al., 2021).
4.2. Chitosan
Chitosan, a deacetylated derivative of chitin, is structurally composed of β-1, 4-linked 2-amino-2-deoxy-β-D-glucopyranose and N-acetyl-D-glucosamine units (Tabatabaei et al., 2022). Distinct from other polysaccharides, chitosan possesses positively charged amino groups, rendering it a water-soluble cationic biopolymer (Maleki and Milani, 2020). This unique chemical architecture enables facile structural tailoring and mechanical property enhancement through molecular modifications, surpassing the functional plasticity of conventional polysaccharides.
However, chitosan's inherent antimicrobial activity prevents its direct use as a standalone encapsulation material for probiotics, as it would induce microbial lethality. Nevertheless, owing to its cationic behavior and acid-resistant properties, chitosan has become one of the most widely employed coating materials for protecting probiotics against harsh gastrointestinal conditions (Lin et al., 2024, Lin et al., 2024). In the microencapsulation strategy developed by Thinkohkaew et al. (2024), chitosan was utilized to coat alginate-gellan gum microcapsules. These microcapsules, containing anionic functional groups such as carboxyl, exhibit negative surface charges in solution. Under acidic conditions, chitosan undergoes protonation of its amino groups (-NH2 → -NH3+), generating positively charged ammonium ions. When the microcapsules are immersed in chitosan solution, electrostatic interactions between the cationic chitosan and anionic microcapsules enable tight adhesion of chitosan to the microcapsule surface. This interfacial assembly forms a uniform coating that modifies surface properties, significantly enhancing structural stability and probiotics protection efficacy (Thinkohkaew et al., 2024).
Moreover, repulsive forces between NH3+ groups in chitosan cause hydrogel swelling, leading to premature probiotic release in gastric environments and thus inadequate protection (Hoang et al., 2021). Even though chitosan remains insoluble in the small intestine, its tendency to swell and release cargo persists. This dual-phase release behavior in both acidic and alkaline environments disqualifies chitosan as an effective pH-responsive material (Xu et al., 2022). Notably, chitosan is selectively degraded by colonic polysaccharidases and microbiota, making it inherently suitable for colon-targeted delivery. Consequently, hybridizing chitosan with acid-resistant polysaccharides offers a superior strategy for developing oral, colon-specific probiotics delivery systems with pH- or enzyme-responsive functionalities (Hoang et al., 2021).
4.3. Cellulose
Cellulose, a linear high-molecular-weight polymer composed of β-D-glucopyranose units interconnected via β-1, 4-glycosidic bonds (Zainal et al., 2021), represents the most abundant natural polysaccharide, serving as a structural component in wood and plant biomass. Isolation from these sources employs physical, chemical, and enzymatic techniques (Huo et al., 2022). Nanocellulose (NC), a derivative of cellulose, exhibits exceptional potential as a drug delivery vehicle due to its unique properties. NC is categorized into three classes: cellulose nanofibrils (CNF), crystalline nanocellulose (CNC), and bacterial nanocellulose (BNC) (Huo et al., 2022).
These nanocellulose variants have been successfully implemented in advanced delivery systems. For instance, Luan et al. engineered pH-responsive macrocapsules by crosslinking CNF with Ca2+ to form a core, followed by electrostatic LbL assembly with chitosan hydrochloride and sodium alginate (Luan et al., 2023). Notably, the glucose units in cellulose adopt a chair conformation, conferring molecular rigidity and extensibility to the polymer chains. Each anhydroglucose unit possesses three hydroxyl (-OH) groups capable of forming hydrogen bonds and undergoing chemical modifications, providing structural versatility for probiotics encapsulation.
A study investigated the encapsulation effect of the composite system of soy cellulose nanofibers (SCNFs) and sodium alginate on Lactobacillus paracasei R23. Their study demonstrated that SCNF10-30 optimizes the multiscale architecture of sodium alginate microbeads through hydroxyl-mediated hydrogen bonding between SCNFs and sodium alginate chains. FTIR spectroscopy confirmed the reinforcement of hydrogen-bond networks within the alginate gel matrix. This structural enhancement improved physical barrier properties and moisture regulation capacity, achieving 89.7 ± 1.5 % gastrointestinal viability and controlled release kinetics, thereby establishing a novel delivery platform for lactic acid bacteria stabilization (Chen et al., 2024). Cellulose-based hydrogels can be engineered through hydrophobic interactions, electrostatic forces, and hydrogen bonding (Bhaladhare and Das, 2022). A study crosslinked methylcellulose (MSCC) with carboxymethylcellulose (MSCCMC) to form pH-responsive hydrogels. FT-IR spectroscopy revealed a blue shift in the -OH absorption peak, indicating intermolecular crosslinking via hydroxyl groups that established the hydrogel network. The hydrophilicity of MSCCMC enhanced the swelling ratio proportionally with its concentration. In acidic gastric fluid (pH 2.0), protonated -COOH groups on MSCCMC and stable hydrogen bonds formed by -OH groups on MSCC maintain structural integrity, limiting premature probiotic release. Conversely, in near-neutral intestinal fluid (pH 8.0), deprotonation generates -COO- groups, intensifying electrostatic repulsion, inducing pore size expansion, and facilitating controlled probiotic release into the intestinal environment, optimizing microbial survival and functionality (Li et al., 2023, Li et al., 2023).
Building upon these advancements, chemically modified celluloses are engineered from native cellulose to achieve tailored material properties. Carboxymethyl cellulose (CMC), a representative derivative, is an anionic linear polysaccharide with exceptional film-forming capacity, functioning as a foundational substrate for edible film fabrication (Khoshdouni Farahani et al., 2025). It derived through partial hydroxyl substitution with carboxymethyl groups (-CH2COO−) on cellulose chains (Pourmadadi et al., 2023). While retaining the fundamental cellulose backbone of β-D-glucopyranose units, this substitution introduces carboxyl functionalities and enhanced aqueous solubility, synergistically preserving cellulose's structural integrity while conferring novel chemical reactivity. The Degree of Substitution (DS), quantifying carboxymethyl substitution extent, significantly governs physicochemical properties. CMC with DS values of 0.0–0.4 is water-insoluble but exhibits controlled swelling, whereas CMC with DS ≥ 0.4 demonstrates full aqueous solubility (Yang et al., 2024). These properties are synergistically enhanced by CMC's inherent advantages, including biodegradability, biocompatibility, hydrophilicity, non-toxicity, and cost-effectiveness. A study innovatively engineered a novel carrier system based on a CMC and κ-carrageenan (κ-Carr) composite blend for the microencapsulation of Lactiplantibacillus plantarum ATCC 13643. In SGF (pH 2.0), free cells experienced near-total inactivation (<1 Log CFU/g), while cells encapsulated within the CMC/κ-Carr blend achieved a survival rate of 7.30 Log CFU/g, demonstrating superior gastrointestinal protection (Dafe et al., 2017b).
In probiotic nanoencapsulation, the standalone application of food-grade polysaccharides for preparing colon-targeting nanofibers remains exploratory. Although biocompatible, their standalone use presents challenges in constructing nanofibers with adequate mechanical properties and colon-targeting functionality. Consequently, current research predominantly focuses on combining different food-grade polysaccharides or polysaccharide-polymer composites, supplemented with spinning aids. Notably, cellulose-based bionanocomposites formed through hybridization demonstrate superior probiotic encapsulation performance. For instance, chitosan is frequently employed as a wall material to confer antibacterial properties, while sodium alginate enhances resistance against digestive fluid erosion (Xu et al., 2022).
4.4. Starch
Starch is typically derived from plant roots, seeds, leaves, stems, and immature fruits. Starch granules comprise amylose and amylopectin. Amylose primarily exists as a linear polymer of D-glucopyranose units linked by α-1,4-glycosidic bonds, with limited branching, while amylopectin forms highly branched clusters through α-1,6-glycosidic linkages, exhibiting high molecular weight and polydispersity. This structural disparity governs functional diversity (Salimi et al., 2023). Starch with high amylose content forms robust, flexible films highly suitable for probiotic microencapsulation (Pech-Canul et al., 2020). High-amylose starch films demonstrate exceptional barrier properties against oxygen and water vapor (Pech-Canul et al., 2020). High-amylose starch films demonstrate exceptional barrier properties against oxygen and water vapor (Lauer and Smith, 2020). Consequently, these films enhance probiotics stability in gastric fluids by reducing water ingress through the matrix and modulating the release kinetics of encapsulated probiotics. The 2,2,6,6-Tetramethylpiperidin-1-oxyl (TEMPO)-mediated oxidation was routinely employed to modify high-amylose starch, introducing substantial carboxyl groups into the polysaccharide structure. These carboxyl groups formed a dense three-dimensional network through strong coordination interactions with Zn2+ ions, enabling efficient probiotic entrapment. Studies implementing this methodology effectively minimized probiotic leakage during encapsulation, achieving an encapsulation efficiency of 88.8 % (Zhang et al., 2021).
Despite promising properties, starch's poor cold-water solubility and instability limit applications. Modifications through physical, chemical, or biological methods are routinely employed to enhance functionality (Ren et al., 2022; Tian et al., 2022). Physical modification alters structure and properties. Techniques like microwave irradiation, high-pressure homogenization, ultra-high-pressure treatment, and high-speed shear induce changes in crystalline structure and granule morphology, improving solubility, water-holding capacity, and viscosity (Punia, 2020).
Chemical modification optimizes functionality and expands applications by structurally altering starch molecules through oxidation, cross-linking, acid hydrolysis, and esterification (Lin et al., 2024). For instance, TEMPO-mediated oxidation significantly elevates carboxyl group content in starch. The introduction of Zn2+ ions further promote gelation by facilitating the formation of an “egg-box” structure through interactions with carboxyl groups. This modified gel system achieved an 88.8 % encapsulation efficiency for Lactobacillus paracasei while enhancing gastric acid resistance (Zhang et al., 2021). Cross-linking agents can also transform starch into resistant starch (Li et al., 2024), which is highly valued for its ability to bypass small intestinal digestion and selectively release probiotics in the colon. This property is particularly advantageous for oral probiotics delivery, ensuring probiotics viability and activity under diverse conditions (Razavi et al., 2021).
Enzymatic modification alters starch molecular structures through hydrolysis, debranching, and branching reactions, regulating functional properties while establishing favorable conditions for probiotic encapsulation. For instance, α-amylase hydrolyzes α-1,4-glycosidic bonds, reducing molecular weight. Branching enzyme cleaves α-1,4-glycosidic linkages and introduces α-1,6-glycosidic bonds, increasing resistant and slowly digestible starch content. Concurrently, it enhanced gelation properties, with the resulting gel network exhibiting high probiotic encapsulation efficiency. The viscoelastic nature of the gel buffered mechanical stress-induced damage to probiotics (Tian et al., 2022). Pullulanase specifically targets α-1, 6-glycosidic bonds, debranching starch molecules to improve molecular alignment and aggregation states. This process increased amylose content, relative crystallinity, and thermal stability. The modified starch formed a dense network structure that underwent slow intestinal degradation, enabling targeted probiotic release and enhancing survival rates in simulated gastrointestinal fluids (Kumari and Sit, 2023). The enzymatic modifications optimized the pore structure, gel strength, and degradation characteristics of starch, thereby providing efficient carriers for probiotic encapsulation. This enhancement improved the stability of probiotics during processing, storage, and gastrointestinal transit.
4.5. Pectin
Pectin, a natural anionic polysaccharide, is a nontoxic, multifunctional heteropolysaccharide derived from fruit cell walls, existing in relatively high contents in grapes, apples, cherries, and citrus fruits. It can be efficiently extracted from plant waste, seaweeds, and fruits (Khoshdouni Farahani, 2021). It primarily consists of homogalacturonan domains, with its main chain composed of α-(1 → 4)-linked D-galacturonic acid (D-GalA), which constitutes approximately 70 % of its structure. Additionally, it incorporates monosaccharides such as L-rhamnose (L-Rha), D-galactose (D-Gal), and L-arabinose (L-Ara), linked in diverse configurations and ratios to form its complex molecular architecture (Barrera-Chamorro et al., 2025; Xiang et al., 2024). This structural diversity underpins pectin's multifunctional properties.
Pectin can form gels through cross-linking with divalent cations such as Ca2+. Notably, low-esterified pectin (degree of esterification, DE < 50 %) generates denser and more stable gel structures when cross-linked with divalent ions, significantly enhancing probiotics protection. In recent studies, Rhamnogalacturonan I rich (RG-I-rich) pectin with low esterification degrees demonstrated superior gel network formation during microencapsulation, providing robust shielding for probiotics. In acidic environments, pectin exists as macromolecular aggregates resistant to proteases and amylases active in the upper gastrointestinal tract (Chen et al., 2024). However, it is selectively degraded by pectinases produced by colonic microbiota. Concomitantly, some research has employed low-methoxy pectin and Ca2+ to form a stable gel structure through ionic cross-linking as a carrier for embedding Bifidobacterium breve. The embedding rate exceeded 90 %, and in a simulated gastrointestinal environment, the probiotics only decreased by 1.76 log cfu/g, which was significantly lower than the 4.82 log cfu/g reduction in the unembedded group (Li et al., 2019).
Pectin exhibits intestinal mucoadhesive properties, which reportedly enhance probiotics adhesion to epithelial cells in vitro (Xu et al., 2023). For instance, the adhesion rate of Lactobacillus fermentum PCC to Caco-2 cells increased from 35 % to 73 % in the presence of pectin with 53.56 % esterification (Srimahaeak et al.). These attributes position pectin as an ideal candidate for colon-targeted delivery.
However, pectin-based delivery matrices are prone to acidic swelling, compromising probiotic viability in gastric fluids. Combining pectin with other biopolymers like alginate significantly enhances mechanical and chemical stability (Heumann et al., 2020).
5. Co-encapsulation of probiotics with multiple polymers
Although polysaccharides have been extensively utilized as encapsulation matrices for probiotics, they exhibit inherent limitations such as porosity, potential probiotics damage, and low mechanical strength (Gao et al., 2022; Lin et al., 2024). Researchers encapsulated Lactobacillus casei and Lactobacillus acidophilus using an emulsification method with oleaster flour. Although encapsulation efficiency was enhanced, scanning electron microscopy (SEM) observations revealed capsules with rough surfaces and porous structures. These morphological imperfections potentially created pathways for acid and enzyme permeation, thereby weakening the protective barrier and ultimately resulting in diminished probiotic viability under SGF conditions (Karkar et al., 2024). Significant advancements require overcoming these deficiencies. Researchers are developing novel strategies to enhance protective efficacy and controlled release through structural optimization, increasingly focusing on constructing robust, compact architectures by combining polysaccharides with other polymers (proteins or alternative polysaccharides). This approach addresses single-polysaccharide system limitations, substantially improving performance and providing a stronger theoretical and practical foundation. Concurrently, studies explore integrating prebiotics as supplementary optimization tools. Building on single-polysaccharide systems, this section examines multi-polymer system advantages, offering insights for next-generation delivery platforms.
5.1. Polysaccharide-polysaccharide composite systems for probiotics encapsulation
Combining two or more polysaccharides can significantly enhance probiotic stability. Researchers commonly employ composites with complementary or synergistic properties. Table 2 summarizes encapsulation performance improvements achieved through such synergies.
Table 2.
Encapsulation efficacy of polysaccharide-polysaccharide synergistic systems for probiotics.
| Strain | Encapsulation material | Encapsulation process | Encapsulation Efficiency (%) |
Simulates gastrointestinal stability |
Storage stability (SR) | Thermal stability (SR) | Otherv Stability (SR) | References | |
|---|---|---|---|---|---|---|---|---|---|
| Simulated gastric fluid (SR) | Simulated intestinal fluid (SR) | ||||||||
| Lactobacillus plantarum Lp90 | Sodium alginate, Chitosan | Ionic cross-linking + polyelectrolyte coating | 83.6 | 92.92 % (pH 2, 2 h) | 77.31 % (pH 8, 2 h) | 87.43 % (28 d, 4 °C) | 1. 61.71 % (68 °C, 30 min) 2. 85.12 % (30 min, 50 °C) |
1. 75.50 % (pH 2, 3 h) | Jike et al. (2024) |
| 2. 83.81 % (0.75 %bile salt, 3 h) | |||||||||
| 3. 95.95 % (centrifugation, 10,000×g, 20 min, 4 °C) | |||||||||
| Lactobacillus casei TISTR 1463 | Sodium alginate, Chitosan, Gellan Gum |
Ionic cross-linking + polyelectrolyte coating | 97.4 | 78.52 % (pH 2.5, 2 h) | 93.06 % (30 d, 4 °C) | The maximum degradation temperature was 270 °C | Thinkohkaew et al. (2024) | ||
| Bifidobacteria | Sodium alginate, Panax ginseng polysaccharide |
Ionic cross-linking | 99.4 | 80.30 % (pH 1.5, 1 h) | 73.66 % (pH 7.5, 2 h) | The zeta potential was −33.0 mV | Yang et al. (2022) | ||
| Lactobacillus plantarum ZJ316 | Chitosan, Pullulan |
Electrospinning technology | 98.7 | 87.24 % (pH 3, 2 h) | 79.71 % (pH 6.8, 3 h) | The thermal degradation temperature was 298.33 °C | Zhang et al. (2024) | ||
| Lactobacillus plantarum SHS01 | Sodium alginate, Chitosan, Lycium barbarum polysaccharide | Ionic cross-linking z + polyelectrolyte coating | 84.5 | 98.63 % (pH 2, 2 h) | 89.87 % (28 d, 4 °C) | Liu et al. (2024) | |||
| Lactobacillus paracasei R23 | Sodium alginate, Cellulose nanofibers | Ionic cross-linking | 92.59 % (pH 2, 2 h) | 75.76 % (pH 7.4, 2 h) | Chen et al. (2024) | ||||
| Lactobacillus plantarum CICC 6240 | Sodium alginate, Chitosan hydrochloride Cellulose nanofibers |
Ionic cross-linking + polyelectrolyte coating | 80.74 % (pH 2.0, 2 h) | 85.61 % (0.3 mg/mL bile salt, 2 h) | Luan et al. (2023) | ||||
SR: Survival rate.
Polysaccharide materials form tightly integrated structures through intermolecular interactions like electrostatic forces, hydrogen bonds, and van der Waals forces. These composite systems maintain stability across broader environmental ranges, reducing leakage or inactivation caused by external fluctuations. Multilayered architectures or composite networks significantly enhance encapsulation efficiency, whereas single systems often fail to sufficiently reduce pore size, leading to premature leakage. Sodium alginate and chitosan exemplify synergistic electrostatic cross-linking. During encapsulation, sodium alginate's carboxyl groups ionize to -COO- under specific conditions, while chitosan molecules contain -NH3+. Electrostatic interactions between -NH3+ and -COO- drive cross-linking, forming stable microcapsules. Jike et al. (2024) encapsulated Lactobacillus plantarum 90 using sodium alginate - chitosan as wall material. This electrostatic cross-linking reduced pore size, minimized probiotics leakage, and achieved an encapsulation efficiency of 83.61 ± 1.20 %. In a study, linear anionic polysaccharide gellan gum was incorporated with so sodium alginate. Ionic cross-linking, double-helix structures, and hydrogen bonding synergistically constructed a denser network. Additionally, chitosan was applied as a coating on the sodium alginate-gellan microcapsule surface via electrostatic interactions. The dual modification significantly enhanced microcapsule stability with reduced porosity (from 42.5 μm to 21.7 μm). This densified architecture effectively minimized probiotic leakage, achieving 97.35 % encapsulation efficiency while suppressing gastric acid infiltration, thereby enabling enhanced probiotic survival through intestinal fluid transit (Thinkohkaew et al., 2024). Similarly, Yang et al. (2022) demonstrated that microcapsules fabricated with Panax ginseng polysaccharide and sodium alginate achieved 99.39 % ± 2.42 encapsulation efficiency. Ginseng polysaccharide integration optimized internal architecture, minimizing leakage and enabling higher retention (Yang et al., 2022).
Composite polysaccharide systems further optimize protection by mimicking the natural intestinal microenvironment, providing comprehensive shielding with dual advantages: (1) supplying diverse nutrients or growth factors to enhance proliferation; (2) forming multilayered structures or specialized microenvironments to block external stressors like gastric acid, bile salts, and competitive inhibition by harmful gut bacteria. This synergy facilitates effective intestinal colonization and functionality. Researchers employed electrospinning technology to encapsulate Lactobacillus plantarum ZJ316 within a dried nanofibrous membrane composed of chitosan and pullulan (PUL). Hydrogen bonding facilitated uniform dispersion within nanofibers, forming stable structures that collectively protected probiotics. In simulated gastrointestinal environments, encapsulated ZJ316 exhibited significantly higher viability than free cells. After SGF, free ZJ316 survival was 78.93 ± 2.51 %, while encapsulated achieved 87.24 ± 2.54 %. Following SIF, survival increased from 68.92 ± 2.46 % (free) to 79.71 ± 1.84 % (encapsulated), demonstrating robust protective efficacy. Pullulan (PUL), a slowly digestible polymer, acts as a prebiotic, providing nutritional substrates. Following intestinal release, encapsulated probiotics utilize PUL as a carbon source, enhancing metabolic activity and facilitating colonization, amplifying beneficial effects on gut health (Zhang et al., 2024, Zhang et al., 2024). Similarly, probiotics gel encapsulated with chitosan, sodium alginate, and Lycium barbarum polysaccharide (LBP), designated SLCG, stored at 4 °C for 28 days, showed viable counts decreasing by only 1.072 log CFU/mL, compared to 1.657 log CFU/mL in sodium alginate gel and 1.393 log CFU/mL in sodium alginate-LBP gel (SLG). This was attributed to LBP promoting probiotic growth and maintaining activity, while its interaction optimized microcapsule structure, enhancing resistance to adverse environments and improving storage stability (Liu et al., 2024).
Furthermore, the study revealed that incorporating cellulose nanofibers (SCNF) into sodium alginate significantly improved the survival rate of probiotics in the gastrointestinal (GI) environment (Chen et al., 2024). Specifically, SCNF10-30 microcapsules exhibited only a 0.63 log CFU/g reduction in probiotics viability after 2 h of SGF digestion, whereas sodium alginate microcapsules showed a markedly higher inactivation exceeding 1.5 log CFU/g. Mechanistically, SCNF10-30 enhanced the immobilized water content in sodium alginate microcapsules from 72.23 % to 87.82 % through calcium ion coordination. This elevated immobilized water fraction stabilized the internal moisture microenvironment, mitigated moisture fluctuation-induced stress on probiotics, and ultimately created more favorable survival conditions.
Building on the foundation of ensuring high survival rates of probiotics encapsulated in composite systems under diverse environmental stresses, the subsequent and more critical challenge lies in achieving precise probiotics release to targeted intestinal regions while maintaining optimal release concentrations to maximize therapeutic efficacy. Polysaccharides can serve as targeted carriers to guide probiotics toward specific intestinal sites, enhancing their local concentration and functional performance (Wang et al., 2024). Earlier reviews, such as that by Cui et al. (2021) detailed how certain polysaccharides not only confer enzyme-responsive colon-targeting capabilities to nanocarriers but also synergize with therapeutics to alleviate inflammatory bowel disease (IBD), providing a theoretical basis for polysaccharide-mediated probiotics targeting. For instance, polysaccharide-based gels (PBGs) enable site-specific release in the intestine through pH- or enzyme-triggered responses (Zhang et al., 2024). Their mucoadhesive properties enhance retention, promote gut homeostasis, and modulate immune function (Xu et al., 2023; Zhang et al., 2024). A notable example is the alginate/chitosan multilayer-coated nanocellulose macrocapsule (Luan et al., 2023). The outer sodium alginate shell undergoes partial dissociation of carboxyl groups in weakly alkaline SIF (pH 7.0), weakening intermolecular interactions and enabling gradual dissolution. This slow degradation process facilitates sustained probiotics release over 8 h, ensuring sufficient viable bacteria reach the distal intestine. A study by D'Amico demonstrated that in the alkaline colonic environment (pH 7.2 ± 0.2), sodium alginate undergoes functional group ionization and swelling, followed by degradation via colonic enzymes. This dual mechanism enabled controlled probiotics release from microcapsules: 3.28 log CFU/mL (36.44 %) of encapsulated probiotics were released after 4 h, increasing to 7.75 log CFU/mL (86.11 %) by 24 h. The progressive degradation of sodium alginate facilitated sustained probiotics liberation, achieving targeted colonic delivery and enhancing their probiotics efficacy in the colon (D'Amico et al., 2024).
Moreover, the differential enzymatic susceptibility of polysaccharides can be strategically leveraged to engineer colonic delivery systems, where carrier matrices are selectively degraded by microbiota-derived specific enzymes (e.g., pectinases, amylases) to achieve site-specific probiotic release in the colon (Luo et al., 2022). Relevant studies demonstrated that when pectin-starch hydrogel microcapsules encapsulating Lactobacillus plantarum were exposed to SGF for 2 h, non-encapsulated free probiotics were almost completely inactivated (survival rate <1 %), while the encapsulated counterparts maintained viability at 6.67 log CFU/g. Upon reaching the colon, microbiota-derived pectinase degradation enabled sufficient viable probiotics to be released and transit to the distal colon (Dafe et al., 2017a).
Beyond direct protection, polysaccharides confer additional functionalities to encapsulation systems, amplifying therapeutic potential. Certain polysaccharides exhibit immunomodulatory properties that enhance host immune responses, creating synergy with probiotic activity. Given the critical interplay between gut microbiota and immune systems, polysaccharides further optimize efficacy by modulating microbial composition and improving microecological balance. The “polysaccharide-gut microbiota axis” shows significant promise in cancer prevention and treatment (Liu et al., 2019).
5.2. Polysaccharide-protein composite systems for probiotics encapsulation
As common encapsulation materials, proteins form suitable embedding walls around active substances, leveraging superior properties for optimal protection and controlled release (Ma et al., 2024). While proteins exhibit excellent film-forming properties, they are prone to aggregation in gastric fluids and highly sensitive to pepsin. Polysaccharides, with their acid resistance and gastric stability, enhance microcapsule compactness and enable targeted release. However, polysaccharides alone generally provide insufficient protection due to poor film-forming properties and large intermolecular pores. Consequently, protein-polysaccharide hybrid systems garner significant attention (Hu et al., 2023), ensuring higher encapsulation efficiency and enhanced protective efficacy. Under specific conditions, non-covalent or covalent interactions (Ke and Li, 2023) enable assembly of multi-scale structured delivery systems. Beyond these interactions, the intrinsic properties of proteins-such as gelation, emulsification, and amphiphilicity-play critical roles in constructing robust probiotics delivery models when combined with polysaccharides. The synergistic advantages of polysaccharide-protein composites in protecting probiotics are summarized in Table 3.
Table 3.
Encapsulation efficiency of polysaccharide-protein systems for probiotics.
| Strain | Encapsulation material | Encapsulation process | Encapsulation efficiency (%) | Simulates gastrointestinal stability |
Storage stability (SR) | Thermal stability (SR) | Other stability (SR) | References | |
|---|---|---|---|---|---|---|---|---|---|
| Simulated gastric fluid (SR) | Simulated gastric fluid (SR) | ||||||||
| Lactobacillus plantarum CECT 220 | Casein, Chitosan | Complex coacervation + spray drying | The load capacity was 11.0 Log CFU/g | 89.91 % (pH 2.5, 2 h) | 86.34 % (pH 6.8, 4 h) | 90.89 % (260 d, 25 °C) | Peñalva et al. (2023) | ||
| Lactobacillus plantarum ATCC 14917 | Casein, Chitosan | Layer-by-Layer self-assembly method | 83.21 % (pH 2, 2 h) | 86.31 % (pH 7.5, 2 h) | −26.39 mV (Zeta potential) | Mohamadzadeh et al. (2025) | |||
| Lactobacillus plantarum CICC 6002 | Casein, Condensing Glue | Water-in-oil emulsions | 89.9 | 69.00 % (pH 1.8, 2 h) | 61.43 % (pH 6.5, 4 h) | 96.45 % (21 d, 4 °C) | 77.21 % (15 s, 85 °C); 71.88 % (15min, 72 °C); 63.58 % (30min, 63 °C) | The zeta potential was −32.9 mV | Zhang et al. (2025) |
| Lactobacillus paracasei subsp. Paracasei | Whey Protein Isolate (WPI), Sodium Alginate |
Double-layer composite crosslinking | 84.5 | 83.76 % (pH 2.0, 2 h) | 89.38 % (pH 6.5, 4 h) | 95.74 % (10 min, 55 °C); 72.68 % (10 min, 75 °C); 89.93 % (10 min, 65 °C) | 41 % (freeze−drying, 36 h, −60 °C); 59.24 % (pH 1.8, 2 h); 80.69 % (pH 2.0, 2 h); 89.89 % (pH 2.5, 2 h) | Alfaro-Galarza et al. (2020) | |
| Lactobacillus acidophilus La5 | WPI, Bitter gourd, polysaccharide (MP) | Thermally cross-linked protein gel | 98.7 | 90.50 % (pH 3, 2 h) | 87.46 % (pH 7, 2 h) | 87.87 % (30 d, 4 °C) | −17.5 mV (Zeta potential) | Bora et al. (2023) | |
| Lactobacillus paraplantarum LR-1 | WPI, Pectin (PEC), Sodium alginate | Bio-induced polyelectrolyte complex gel | The load volume was 10.3 log CFU/g | 15.03 % (pH 2.5, 2 h) | 69.85 % (pH 8.0, 2 h) | 93.5 % (28 d, −20 °C) | 42.53 % (30 min, 63 °C); 69.55 % (15 s, 89 °C) | Rao et al. (2025) | |
| Lactobacillus reuteri | WPI, Sodium alginate, Chitosan |
Layer-by-layer assembly + Freeze-drying | 95.9 | 92.86 % (pH 2.0, 2 h) | 87.91 % (1 min, 95 °C) | 92.86 % (0.3 mg/ml bile salt, 2 h); 76.92 % (0.85 % NaCl2, 2 h, pH 11.0); 92.86 % (0.85 % NaCl2, 2 h, pH 2.0); 87.69 % (freeze-drying, −80 °C, 36 h) | Ding et al. (2023) | ||
| Lactobacillus plantarum 550 | Soy Protein Isolate (SPI), PEC | Complex coacervation + Spray drying | 96.9 | 98.72 % (pH 2.0, 2 h) | 96.65 % (pH 7.2, 4 h) | 99.79 % (28 d, 4 °C) | 99.89 % (10 min, 55 °C); 88.90 % (10 min, 65 °C); 72.79 % (10 min, 75 °C) | 99.68 % (ultraviolet light, 235.7 nm, 30 min) | Hu et al. (2023) |
| Lactobacillus plantarum 550 | SPI, Peach gum polysaccharide (PG) | Complex coacervation + Spray drying | 93.14 % (pH 2.0, 2 h) | 96.69 % (pH 7.2, 4 h) | 96 % (32 d, 4 °C) | 71.54 % (30 min, 65 °C); 79.54 % (10 min, 75 °C) | Yao et al. (2023) | ||
| Lactobacillus plantarum | SPI, Soybean hypocotyl polysaccharide (SHP) | High internal phase emulsions | 71.50 % (simulated colon fluid, pH 6.8, 2 h) | −57.68 mV (Zeta potential) | Sun et al. (2024) | ||||
| Bacillus subtilis 168 | Zein, Soy Soluble Polysaccharide (SSP) | Self-assembling polyelectrolyte complex coacervation | 81.4 | 95.87 % (pH2.0, 2 h) | 88.00 % (pH6.8, 2 h) | 83.40 % (56 d, 4 °C) | 95.89 % (30 min, 65 °C) | Cheng et al. (2024) | |
| Lactobacillus casei | Zein, Chitosan |
Polyelectrolyte complex coacervation | 68.4 | 87.5 % (pH 1.2, 2 h) | 93.85 % (pH 6.8, 4 h) | 96.48 % (28 d, 4 °C) | 47.5 % (5 min, 85 °C) | Ma et al. (2024) | |
| Lactobacillus plantarum LP23-1 | Zein, PEC | Water-in-oil-in-water double emulsions | 96.3 | 86.75 % (pH 2, 2 h) | 72.41 % (pH 7, 2 h) | −50.06 Mv (Zeta potential) | Xu et al. (2024) | ||
| Lactobacillus plantarum Lp90 | SPI, Reducing sugars | Maillard reaction-conjugated encapsulation matrices | 87.4 | 97.00 % (pH 2, 2 h) | 86.14 % (pH 8, 2 h) | 94.29 % (28 d, 4 °C) | 68.57 % (30 min, 68 °C); 91.57 % (30 min, 50 °C) | 96.29 % (pH 2, 3 h); 86.14 % (0.75 % bile salt, 3 h); 98.14 % (separation, 10,000 × 4 °C, 20min) | Jike et al. (2024) |
| Lactobacillus casei | WPI, SHP | Maillard reaction-conjugated encapsulation matrices | 92.68 % (pH 2, 2 h) | 76.86 % (pH 8, 3 h) | 76 % (28 d, 4 °C) | 1. 98.19 % (30 min, 60 °C) 2. 62.34 % (5 min, 80 °C) |
87.35 % (lyophilization, −80 °C, 24 h) | Song et al. (2024) | |
SR: Survival rate.
5.2.1. Polysaccharide-casein delivery system
Casein is a protein abundant in milk, accounting for approximately 80 % of the total milk protein. It is a phosphocalcium-binding protein, and its surface carries different charges under different pH conditions. When the pH is higher than its isoelectric point (about pH 4.6), the carboxyl groups in the casein molecules will dissociate, generating negatively charged carboxylate ions (-COO-), making the casein molecule negatively charged overall. This creates conditions for electrostatic interactions with positively charged substances. One of the most studied delivery systems is the casein-chitosan composite carrier. In the research by Allahverdi et al., based on electrostatic interactions, the LbL deposition method was used. Positively charged chitosan was first adsorbed onto the negatively charged surface of Lactobacillus plantarum, followed by the deposition of negatively charged casein, ultimately forming micro-particles casein-chitosan microparticles with a stable encapsulation structure (Allahverdi et al., 2024). In the study by Peñalva et al. (2023), the co-encapsulation of probiotics by casein and chitosan was achieved through the complex coacervation method. Although the two research teams used different embedding methods, both relied on the electrostatic interaction between chitosan and casein. This electrostatic interaction is the key to constructing a stable encapsulation structure, which helps improve the embedding effect and the stability of probiotics. The survival rate of probiotics in a simulated gastrointestinal environment exceeded 80 % in both studies (Peñalva et al., 2023).
Casein exhibits unique physicochemical properties due to its proline-rich peptide sequences, which disrupt α-helical and β-sheet conformations in its supramolecular structure, resulting in an open and flexible molecular configuration. This structural flexibility confers exceptional thermal stability, enabling casein to remain stable under high-temperature conditions. Under thermal stress, casein-gellan gum complexes form stable matrices that efficiently encapsulate Lactobacillus plantarum, minimizing heat-induced damage by reducing direct environmental exposure. This protective mechanism significantly enhances the thermal resilience of Lactobacillus plantarum, achieving a 77.21 % survival rate even after 15 s of extreme heat treatment at 85 °C (Zhang et al., 2025).
5.2.2. Polysaccharide-whey protein isolate delivery systems
Whey protein isolate (WPI), a highly digestible and nutritionally superior protein. In the food industry, it serves not only as a critical raw material for formulating functional foods but is also widely utilized for encapsulating probiotics and stabilizing bioactive compounds. (Khoshdouni Farahani et al., 2022d, Zhang et al., 2025). WPI primarily consists of β-lactoglobulin, α-lactalbumin, bovine serum albumin, and immunoglobulins (Xie et al., 2023). Under the extreme conditions of gastric digestion, these proteins generate localized high-pH microenvironments within the protein matrix of microcapsules. For instance, β-lactoglobulin in WPI exhibits resistance to acidic environments and pepsin, effectively shielding probiotics from gastric acid and enzymatic degradation. Additionally, β-lactoglobulin enhances the buffering capacity of the medium, stabilizing the internal microenvironment of microcapsules and mitigating acid-induced stress on probiotics. In SGF, microcapsules containing β-lactoglobulin maintained a significantly higher probiotics survival rate: free cells exhibited a reduction of 5.02 log CFU/g after 2 h of SGF digestion, while encapsulated cells showed only a 1.59 log CFU/g decline (Han et al., 2020).
In addition, the excellent gel-forming properties and amphiphilic characteristics of WPI have played a positive role in the construction of probiotics delivery models (Xie et al., 2023). In the combination of bitter gourd active polysaccharide (MP) and WPI, the two interact through electrostatic, hydrophobic and hydrogen bonding forces. The gel-forming ability of WPI provides a basic structural framework for the composite system. During the preparation of the composite gel, WPI and MP jointly form a three-dimensional network structure, and their synergistic effect enhances the texture properties of the gel, such as hardness, elasticity and cohesiveness, to form a stable composite gel network. Compared with the WPI gel alone, the MP-WPI composite gel encapsulates probiotics and significantly improves the encapsulation efficiency, reaching up to 98 %. The gel network formed by WPI provided a robust structural framework, while the MP enhanced the network density via hydrogen bonding, thereby improving the encapsulation efficiency. This stable gel structure provides a physical protective barrier for probiotics, making them more stable during storage and digestion (Bora et al., 2023).
5.2.3. Polysaccharide-soy protein isolate delivery systems
Soy protein isolate (SPI), a plant-based protein with broad application prospects, offers advantages including low cost, high nutritional value, abundant availability, and full biodegradability. SPI exhibits film-forming properties, enabling the creation of a foundational structure for probiotic encapsulation. Furthermore, it can form complexes with hydrocolloids as delivery vehicles, significantly enhancing the transport efficiency of bioactive compounds. Furthermore, SPI demonstrates inherent water retention capacity and emulsifying properties, with its moisture retention capability being substantially improved through polysaccharide complexation (Khoshdouni Farahani et al., 2024a). However, SPI microcapsules developed in the study by Hu et al. suffered from structural porosity, low encapsulation efficiency, and inadequate digestive resistance, providing limited protection to probiotics during spray drying and gastrointestinal transit. To address these limitations, pectin (PEC) was incorporated to mitigate thermal stress and dehydration-induced damage to cell membranes. The resulting SPI-PEC microcapsules achieved a probiotic survival rate of 50.52 % post-spray drying, significantly surpassing SPI-only capsules (26.19 %). Leveraging SPI's inherent acid-base buffering capacity synergized with PEC's ability to reduce microcapsule water solubility and resist enzymatic degradation, the hybrid system provided comprehensive probiotic protection. During 6 h of simulated gastrointestinal digestion, viable bacterial counts decreased by only 0.43 Log CFU/g. Thermal stability testing revealed a negligible reduction of 0.01 Log CFU/g after 10 min heat treatment at 55 °C. The microcapsules also demonstrated exceptional UV resistance, with a viability loss of merely 0.03 Log CFU/g following 30-min UV irradiation. Furthermore, probiotic survival rates remained stable without significant decline after 4-week storage at 4 °C (Hu et al., 2023).
This co-encapsulation strategy enhances probiotic stability and compensates for inherent material limitations. Studies have demonstrated that increasing polysaccharide content in composite systems improves the storage stability of encapsulated probiotics. For instance, in SPI microcapsules incorporating peach gum polysaccharide, probiotics exhibited higher survival rates. During 4 °C storage, polysaccharide addition significantly enhanced probiotic viability. As the polysaccharide concentration increased, the microcapsules showed reduced moisture content, elevated Tg, increased matrix viscosity, and suppressed molecular mobility, thereby inhibiting diffusion-controlled reactions such as oxidation. Furthermore, polysaccharide's free radical scavenging capacity strengthened proportionally with its content, enhancing the wall material's ability to neutralize storage-induced free radicals and further improving the storage stability of encapsulated probiotics (Zhang et al., 2025).
5.2.4. Polysaccharide-zein delivery system
Zein, a corn-derived storage protein, possesses a unique amphiphilic structure that enables spontaneous self-assembly into nanosphere configurations through antisolvent precipitation. This intrinsic property renders it particularly suitable for active ingredient encapsulation, targeted delivery systems, and controlled-release mechanisms, as demonstrated in numerous studies (Xie et al., 2023; Zhang et al., 2022, Zhang et al., 2022).
Zein contains acidic carboxyl groups and basic amino acids, conferring exceptional buffering capacity. In gastric acid environments, it modulates local pH by reversibly binding or releasing protons, creating a mild and survival-conducive microenvironment for probiotics. In a simulated gastrointestinal digestion study, free Lactobacillus plantarum 23-1 viability dropped below detectable levels, whereas Lactobacillus plantarum 23-1 encapsulated in water-in-oil-in-water (W/O/W) emulsions stabilized by zein-pectin complexes maintained high survival rates. Pectin formed robust interactions with zein through electrostatic interactions and hydrogen bonding, which not only compensated for the emulsion phase separation defect caused by the strong hydrophobicity of zein when used alone but also, when combined with zein's buffering effect in the acidic gastric environment, enabled the 1:1 zein-pectin composite to increase the probiotic survival rate to 73.36 %. (Xu et al., 2024).
Under thermal stress, zein can decelerate heat transfer to encapsulated probiotics. Its unique structural and physicochemical properties create a barrier that impedes heat penetration through the encapsulation layer, thereby reducing thermal damage to probiotics. Using LbL assembly technology, zein and pectin were co-encapsulated via charge-driven interactions. Cells encapsulated with (ZNP/P) 0.5 (1 layer of zein nanoparticles and 1 layer of pectin) exhibited a viability loss of 0.68 ± 0.02 log CFU/mL after 10 min of heating and 1.20 ± 0.03 log CFU/mL after 30 min. In contrast, unencapsulated cells lost nearly all viability within 10 min (Liu et al., 2023).
5.2.5. Polysaccharide-gelatin delivery systems
Gelatin (GL), a biodegradable protein derived from partial hydrolysis of collagen, is recognized as a commercially viable material for encapsulation (Li et al., 2022). Its excellent water solubility, emulsification, thickening capacity, and high cross-linking activity (due to primary amine groups) make it an ideal candidate for co-encapsulation with anionic polysaccharides, forming strong interactions with negatively charged materials (Zhang et al., 2022). Although gelatin has garnered interest for probiotics microencapsulation, its high solubility at physiological and aqueous temperatures limits standalone applications. Consequently, gelatin is often combined with other compounds to enhance probiotics protection (Paula et al., 2019).
For instance, Studies have employed LbL assembly to co-encapsulate Lactobacillus rhamnosus 6133 with gelatin, sodium alginate, and hyaluronic acid. The resulting multilayer structure, stabilized by electrostatic interactions and hydrogen bonding, demonstrated superior gastrointestinal resistance. After 90 min in SGF, free Lr-6133 and sodium alginate-encapsulated cells suffered a viability loss of ∼3 log CFU/mL, while gelatin-based microcapsules reduced this loss to 0.99 log CFU/mL. In SIF, Lr-6133 viability decreased by only 0.5 log CFU/g, with post-digestion survival rates remaining above 6 log CFU/mL, indicating effective gastric protection and controlled intestinal release (Wang et al., 2024). Similarly, recent studies have successfully fabricated probiotic microcapsules through co-encapsulation using gelatin combined with gum arabic and diacylglycerol. The resulting microcapsules exhibited a moisture content of 1.67 % and hygroscopicity of 9.89 g/100 g, data indicating enhanced storage stability with suppressed microbial growth and minimized chemical reactivity (Chen et al., 2023). Furthermore, the multiple functional groups in gelatin molecules enable synergistic interactions with sodium alginate, increasing both cross-linking density and structural complexity of the gel network. This structural enhancement significantly improved the stability of composite hydrogel beads. During storage and thermal stress experiments, the gelatin-containing hydrogel beads demonstrated superior probiotic protection compared to sodium alginate-only counterparts (Ni et al., 2023).
The inherent antioxidant property of gelatin also endows the delivery system with special capabilities. The amino acids it contains, such as arginine (Arg), tyrosine (Tyr), and phenylalanine (Phe), are key sources of antioxidant activity (Shiao et al., 2021). In the DPPH experiment, pure gelatin demonstrated high antioxidant activity, and the antioxidant activity of the probiotics double-layer encapsulated with gelatin and sodium phytate (Gel/Phy/Bs) prepared by Mehdi Dadmehr et al. was even more remarkable, with a free radical inhibition rate of 85.12 % (Dadmehr et al., 2024).
5.2.6. Maillard reaction products
The Maillard reaction, a non-enzymatic interaction between amino groups (proteins, peptides, or amino acids) and carbonyl groups (reducing sugars), plays a critical role in modifying protein functionality to enhance food quality and stability (Naik et al., 2022). For instance, Maillard conjugation of casein with pectin or arabinogalactan reduces casein's surface hydrophobicity, increases its absolute zeta potential, and significantly improves solubility, foaming capacity, emulsification, antioxidant activity, and thermal stability (Haro-González et al., 2024). Maillard reaction products (MRPs), formed via covalent conjugation of proteins and carbohydrates, serve as novel biopolymer wall materials with enhanced thermal stability and antioxidant activity, increasingly utilized for encapsulating and delivering bioactive compounds and probiotics. They exhibit marked advantages in minimizing losses of sensitive ingredients during food processing (Li et al., 2023). For example, SPI conjugated with glucose or α-lactose under alkaline conditions (pH 9.0) at 90 °C for 3 h generated MRPs. These conjugates formed covalent bonds between proteins and carbohydrates, altering protein structure and creating an ideal matrix for probiotics encapsulation. The resulting Lactobacillus plantarum-MRP (Lp-MRP) microcapsules achieved an encapsulation efficiency of 87.41 % ± 0.67 %. MRPs also demonstrated intrinsic antioxidant activity, mitigating lipid oxidation-induced membrane damage during storage. After 4 weeks at 4 °C, viability of encapsulated Lp90 decreased by only 0.40 log CFU/mL, with no significant weekly fluctuations, highlighting superior storage stability (Jike et al., 2024).
MRPs enhance the Tg to improve the protective efficacy of encapsulation systems for probiotics. For instance, conjugates prepared via the Maillard reaction between WPI and soybean hull polysaccharide (SHP) were used to encapsulate Lactobacillus casei. During freeze-drying, the high Tg of WPI-SHP conjugates effectively inhibited ice crystal growth, preventing mechanical damage to probiotics cells. Experiments demonstrated that encapsulated L. casei achieved a freeze-drying survival rate of 87.35 % ± 1.73 %, significantly higher than that of unencapsulated cells (24.95 % ± 0.55 %). In simulated gastrointestinal digestion, the conjugate structure acted as a barrier, slowing the penetration of gastric acid and bile salts, thereby reducing probiotics inactivation. While unencapsulated L. casei suffered substantial viability loss post-digestion, WPI-SHP-encapsulated cells maintained high viability. Furthermore, under 4 °C and 25 °C storage conditions, L. casei encapsulated with WPI-SHP4d (Tg = 140.51 °C) retained viability above 10 log CFU/mL and 7.98 log CFU/mL, respectively, after 4 weeks, whereas unencapsulated cells showed significant decline (Dadmehr et al., 2024).
5.3. Impact of prebiotics on probiotics encapsulation systems
Beyond the ongoing exploration of encapsulation material properties, a highly valuable research direction has emerged: the co-encapsulation of probiotics with bioactive substances. This approach has gained increasing attention from both academia and industry due to its core advantage in effectively maintaining probiotics viability while fostering synergistic effects between probiotics and bioactive compounds (Ma et al., 2023). Co-encapsulation is not merely a physical mixture but involves rational integration of these components into a unified system via tailored techniques, leveraging their intrinsic properties and interaction mechanisms. Prebiotics are primarily defined as carbohydrates (Neri-Numa and Pastore, 2020), predominantly oligosaccharides, though they may also include non-carbohydrate substances (e.g., polyphenols, peptide polymers) and specific polysaccharides (e.g., inulin). These substances exhibit significant therapeutic potential, such as modulating lipid metabolism, regulating energy homeostasis, supporting neurocognitive development, and enhancing mineral bioavailability (Lordan et al., 2020). This section will focus on the roles of prebiotics in polysaccharide-based encapsulation systems, providing insights for refining co-encapsulation delivery models that integrate probiotics and bioactive agents.
5.3.1. Polysaccharide-based delivery systems incorporating prebiotics
In practical applications, prebiotics are rarely used alone for probiotics delivery but are often combined with polysaccharides, proteins, or other materials. For instance, β-glucan exhibits low oral bioavailability and solubility due to the absence of degrading enzymes in the upper gastrointestinal tract, limiting its functionality. To address this, researchers have integrated β-glucan with lipids, proteins, or polysaccharides to form delivery complexes that could improve solubility, enhance physical barriers, and synergistically regulate the intestinal environment, effectively compensating for the low bioavailability of β-glucan due to the lack of degrading enzymes in the front part of the intestine. At the same time, it significantly enhanced the protective effect on probiotics. (Chen et al., 2024).
Similarly, inulin has been employed as a supplement to materials like alginate and chitosan for probiotics encapsulation via electrostatic interactions. A study demonstrated that co-encapsulating Lactobacillus acidophilus with inulin and sodium alginate achieved an encapsulation efficiency of 96.75 %. Remarkably, under −18°Cstorage, the inulin-supplemented group (AIN) maintained viable counts ≥6 log CFU/g for 120 days, outperforming other formulations (Poletto et al., 2019). This highlights inulin's role in enhancing probiotics resilience during frozen storage.
In another approach, researchers have employed low concentrations of quercetin (Que, 0.05 %) boosted Lactobacillus casei viability to 1.03 × 1013 CFU/mL. Que-incorporated microcapsules achieved an encapsulation rate of 68.44 %, significantly higher than the polyphenol-free control (51.55 %). Furthermore, rutin (Rt)-loaded zein-chitosan complex coacervates (ZCSC) demonstrated enhanced acid resistance through cross-linking reactions between zein's hydroxyl side chains and Rt's phenolic groups under low gastric pH. This structural stabilization ensured a post-gastric viability of 7.30 ± 0.03 log CFU/mL, meeting the threshold for effective colonic delivery (Ma et al., 2024).
The co-encapsulation of probiotics and prebiotics can further enhance the vitality of probiotics in harsh environments by promoting their growth and proliferation. The encapsulation efficiency was significantly enhanced from 78.9 % to 99.2 % when embedding the resistant starch-probiotic mixture in sodium alginate-chitosan microcapsules, with nearly complete retention within the microcapsule matrix. This substantial improvement was attributed to the optimal compatibility between the larger particle size and the calcium alginate network architecture, which effectively hindered their leakage. The structural configuration ensured sustained stability of the probiotic-prebiotic synergistic effect through physical entrapment and molecular interaction mechanisms. In simulated gastrointestinal fluids, the free-state Lactobacillus plantarum without resistant starch addition experienced a drastic reduction to 1.74 ± 0.23 log CFU/g, while encapsulated cells still showed a 3.4-log cycle decrease. In contrast, co-encapsulation with resistant starch reduced the viability loss to approximately 3-log cycles. After 90 days of storage at 25 °C, co-encapsulation with resistant starch enhanced the survival rate of Lactobacillus plantarum by nearly 1-log cycle, reaching 93.61 %, thereby effectively maintaining probiotics functionality and extending shelf life (Li et al., 2024).
In addition to enhancing probiotics viability, prebiotics can further improve probiotics survival rates by suppressing the growth of pathogenic bacteria (Chauhan and Sharma, 2023). Inulin is currently the most intensively studied prebiotic category. It exerts prebiotic effects in Lactobacillus plantarum ATCC 14917 encapsulated within CMC and cellulose nanofibers (CNFs). During storage, the survival rate of probiotics in the CMC-inulin nanobiocomposite increased by 36 %. Furthermore, the composite film demonstrated antimicrobial properties against pathogenic bacteria (Zabihollahi et al., 2020). By optimizing the composition of inulin, crystalline nanocellulose (CNC), and WPI, encapsulation efficiency was enhanced, and probiotics survival in simulated digestive fluids improved (Maleki and Milani, 2020). Encapsulating probiotics with inulin as a microencapsulation material not only enhances gastrointestinal stability, survival rates, and storage activity of probiotics but also exerts prebiotic activity by modulating gut microbiota. Additionally, inulin regulates intestinal microbial communities to stimulate immunity against various disease conditions (Chen et al., 2024). Inulin promotes the growth of Lactobacillus and Bifidobacterium species. By stimulating short-chain fatty acid (SCFA) production, it reduces mucosal lesion scores and alleviates mucosal inflammation. The combination of inulin with Lactobacillus acidophilus CHO-220 significantly lowered low-density lipoprotein (LDL) cholesterol levels. Inulin improves and preserves gut microbial diversity while helping prevent pathogen invasion and translocation into host cells (Correa et al., 2024).
In summary, as indigestible bioactive compounds, prebiotics not only provide essential nutrients for probiotics growth but also enhance their resistance to acid, heat, and dehydration. Additionally, they suppress the proliferation of pathogenic microorganisms in the gastrointestinal tract, thereby reducing competitive pressures on probiotics survival. These multifaceted functionalities position prebiotics as a continually explorable class of bioactive agents, warranting their expanded application in the development of advanced probiotics delivery systems.
6. Conclusions and future insights
Probiotics play an indispensable role in maintaining human health, with their efficacy in disease prevention, metabolic regulation, and gut health management having been extensively validated across multiple fields. However, practical applications of probiotics face significant challenges, as their viability is prone to degradation during processing, storage, and gastrointestinal transit, leading to reduced bioavailability. Polysaccharide-based encapsulation techniques provide an effective strategy to address the viability loss of probiotics during processing, storage, and gastrointestinal transit. Conventional approaches including embedding, coating, and their hybrid combinations establish preliminary protective barriers. Nevertheless, single-polysaccharide systems exhibit significant limitations: sodium alginate shows weak protective capacity in gastric low-pH environments (pH < 3.0), while chitosan cannot function alone due to its inherent antimicrobial activity against probiotics, cellulose struggles to form independent nanofibrillar structures, starch inherently lacks stability, while pectin undergoes swelling in acidic environments. Comparatively, co-encapsulation with diverse polymers demonstrates superior performance. Specifically, polysaccharide-polysaccharide composites enhance encapsulation efficiency, protective efficacy, and targeted release capabilities through intermolecular interactions. Polysaccharide-protein hybrids synergistically optimize microcapsule functionality. Furthermore, co-encapsulation of prebiotics with probiotics promotes bacterial proliferation, enhances viability, and suppresses pathogens, thereby advancing delivery system design optimization.
Therefore, suitable encapsulation materials can be selected according to different requirements and purposes. For targeted release, options include pectin, resistant starch, and sodium alginate-chitosan complexes. Materials such as gum arabic, pullulan, and sodium alginate-gelatin composites are preferable for achieving high encapsulation efficiency and stability. Functional synergy can be achieved through the combination of prebiotics and functional polysaccharides. Embedding and coating technologies can be adapted to different materials: embedding techniques like spray drying are suitable for hydrophilic polysaccharides such as gum arabic, while ionotropic gelation is effective for anionic polysaccharides including sodium alginate. Coating methods include LbL self-assembly, which relies on oppositely charged polymers, and covalent crosslinking applicable to modifiable polysaccharides. However, composite materials face challenges such as complex preparation processes, insufficient release controllability, variability in biocompatibility and degradability, and suboptimal functional synergy efficiency.
Future studies should prioritize the exploration of novel encapsulation materials, specifically targeting the inherent limitations of single polysaccharides such as sodium alginate with poor low-pH protective capacity and chitosan exhibiting biphasic release characteristics, through systematic design of pH-sensitive modified materials. Meanwhile, the polysaccharide-polysaccharide and polysaccharide-protein composite systems face phase separation risks due to component ratio imbalances, particularly manifested through thermodynamic incompatibility at high protein concentrations. This necessitates systematic interfacial interaction analysis to optimize compositional ratios. Moreover, laboratory-scale techniques like layer-by-layer (LbL) self-assembly and electrospinning demonstrate limited scalability, with current research lacking industrially viable alternative processes and comprehensive analyses of performance deterioration mechanisms during production. Future development must prioritize scalable manufacturing technologies to bridge the lab-to-industry gap. Additionally, current probiotic delivery systems show inadequate adaptation to individual intestinal microenvironments, including variations in microbiota composition, pH gradients, and enzyme activity profiles. Precision engineering strategies based on gut microbiome signatures are required to customize carrier systems and regulate release kinetics, transitioning from universal protection to targeted delivery. Ultimately, these advancements will drive the translation of probiotic encapsulation technologies into practical applications such as functional food additives and clinical-grade microecological formulations.
Glossary
N/A.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
This work was financially supported by the Project for International Science and Technology Cooperation/Science and Technology Cooperation with Hong Kong, Macao and Taiwan of Hebei Province, China (25297102D), Subsidy funds for the Innovation Center of Pig Processing Technology (247790259H), and the National Nature Science Foundation of China (32101878).
Handling Editor: Dr. Xing Chen
Contributor Information
Dandan Zhao, Email: zdd6364@126.com, zdd2016@hebust.edu.cn.
Xiaoguang Gao, Email: gaoxiaoguang999@126.com.
Jianxiong Hao, Email: cauhjx@163.com.
Data availability
Data will be made available on request.
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Associated Data
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Data Availability Statement
Data will be made available on request.