Microbial Fermentation: A Sustainable Strategy for Producing High-Value Bioactive Compounds for Agriculture, Animal Feed, and Human Health

Abstract

Microbial fermentation is a key biotechnological tool for producing bioactive metabolites such as alkaloids, carotenoids, essential oils, and phenolic compounds, among others, with applications in human health, agriculture, and food industries. This review comprehensively reviews recent information on the synthesis of valuable compounds and enzymes through fermentation processes. Here, we discuss the advantages of the different types of fermentation, such as submerged and solid-state fermentation, in optimizing metabolite production by bacteria, fungi, and yeast. The role of microbial metabolism, enzymatic activity, and fermentation conditions in enhancing the bioavailability and functionality of these compounds is discussed. Integrating fermentation with emerging biotechnologies, including metabolic engineering, further enhances yields and specificity. The potential of microbial-derived bioactive compounds in developing functional foods, pharmaceuticals, and eco-friendly agricultural solutions positions fermentation as a pivotal strategy for future biotechnological advancements. Therefore, microbial fermentation is a sustainable tool to obtain high-quality metabolites from different sources that can be used in agriculture, animal, and human health.

1. Introduction

Bioactive compounds are found in plants, fruits, vegetables, and cereals. These compounds, such as alkaloids, antibiotics, mycotoxins, phenolics, pigments, polysaccharides, and vitamins, are among the compounds that microalgae can synthesize or microorganisms can excrete during the fermentation process. These metabolites are linked with promoting benefits for human health, such as antioxidants, anti-tumor, anti-diabetes, cardiovascular protection, and hypoglycemic activities. Therefore, metabolites are a target due to the promising ingredients of different nutraceutical, pharmaceutical, and cosmetic products [1,2,3,4]. Additionally, plant growth factors, biostimulation, elicitation, and the inhibition of phytopathogenic diseases on plants have been studied, as well as the potential activities of metabolites such as polysaccharides, enzymes, and microorganisms in submerged and solid-state fermentations for agricultural uses [5,6,7]. Recently, the effect of fermentation of food waste to enhance the diet to improve the health and growth of different animals has been studied; in this sense, the use of microorganisms as probiotics on animal models, as well as the metabolites obtained by fermentation of food waste as ingredients in animal feed, have been shown to enhance the health and growth of goats, pigs, and other animals; therefore, the use of fermentation of food waste could be a promising alternative to commercial animal products, contributing to a circular economy by enabling waste materials to be reintegrated into the production cycle and continue generating value [8,9,10,11]. This study highlights the importance of microbial fermentation as an innovative and sustainable alternative to obtain metabolites related to human health, agriculture, and animal uses.

2. Literature Research Methodology

To collect relevant literature on the subject, we performed literature research using the databases Scopus, Web of Science, and Google Scholar using the following keywords: fermentation, “bioactive compounds”, “one-health”, “metabolite production”, “solid-state fermentation”, enzymes, polyphenols, carotenoids, essential oils, polysaccharides, applications, lactic acid bacteria, Lactobacillus, Saccharomyces, Aspergillus, “functional foods”, pharmaceuticals. We mainly included literature from 2015 to 2025 and only reports published in English. At the end, we included 296 articles in this review paper (Figure 1).

3. Fermentation as a Strategy to Produce Metabolites of Interest

Fermentation is a preservation technique used since ancient times to extend the shelf life of food and improve the bioaccessibility and bioavailability of nutrients from different sources [12]. This process occurs through microorganisms such as bacteria, fungi, and yeast that transform substrates into new ingredients such as textures, fragrances, flavors, enzymes, and bioactive compounds [13,14]. The fermented products are affected by several factors, such as temperature, pH, aeration, agitation, and microorganism-to-substrate ratio; these parameters are monitored and studied to control the fermentation process [14,15]. The fermentation process is divided into submerged fermentation (SF) and solid-state fermentation (SFF), which depend on the substrate and nutrient medium based on the cultivation of microorganisms [14]. These fermentation types have some advantages, which are related to the growth of microorganisms, specificity, and adaptation to fermentation conditions (Figure 2) [16,17]. In this sense, Candida, Galactomyces, Kluyveromyces, Monascus, Pediococcus, Streptococcus, Torulaspora, and Weisella microorganisms have been studied on submerged fermentation, while Lactococcus, Lentimus, Mucor, Thamnidium, Trichoderma, and Yarrowia microorganisms have been used on solid-state fermentation [17,18,19,20,21,22].

3.1. Submerged Fermentation

Submerged fermentation is performed in a liquid substrate where microorganisms can adapt to high humidity. This fermentation method is one of the most studied because it is considered technologically easy to scale up to large-scale processes. Less extreme and more economical conditions are required to grow, diffuse, and separate microorganisms and compounds [17,19]. Submerged fermentation can be carried out in various ways, including batch, fed-batch, and continuous fermentation. The fermentation method used needs to be oriented to cover the characteristics of the microorganism and nutrients studied, as well as the infrastructure and separation steps required to obtain product compounds [16,17,23]. This type of fermentation is used in other processes. Also, it has been reported that a wide range of raw materials has been studied to extract different compounds such as flavonoids, antioxidants and antifungal peptides, dietary fibers, and volatile compounds, among others of industrial importance [17]. However, submerged fermentation requires a longer fermentation time due to its lower performance. Also, high effluent volumes are produced, and waste needs to be stabilized before being dropped into the environment [24]. The most studied microorganisms in submerged fermentation are bacteria (Bacillus, Bifidobacterium, Lactobacillus, Pediococcus, and Streptococcus), yeast (saccharomyces), and some fungi (Cordyceps, Galactomyces, kluyveromyces, Monascus, Picchia, Rhizopus, Torulaspora, and Weisella) that can grow in high humidity and transform the carbohydrates into organic acids, alcohols, other compounds, and new ingredients with health benefits, as well as enzymes that are secreted into the liquid medium [17,25].

3.2. Solid-State Fermentation

Solid-state fermentation is a technique in which microorganisms grow in the absence, partially or completely, of water and metabolize the nutrients of the solid substrate to transform carbohydrates into other compounds [1]. In recent years, various substrates have been studied for solid-state fermentation. Among these, agricultural biomass is considered an alternative for obtaining metabolites of agricultural, cosmetic, and pharmacological importance using fungi, yeast, and certain bacteria. Also, this solid-state fermentation has been considered cheaper in contrast to submerged fermentation due to processing conditions, simpler equipment, lower sterilization cost, and reduced downstream processing, among others [1,6,20,22]. Likewise, a higher yield of metabolites in shorter times has been obtained under solid-state fermentation compared to submerged fermentation, which could be due to specific growth conditions of microorganisms under stressed conditions, as well as uncompetitive inhibition with other microorganisms to consume the nutrients of substrates [6,12,26].

4. Microbial Metabolism and Fermentation Mechanism

4.1. Metabolic Changes in Microorganisms

In general, to be considered a “fermentation”, microorganisms must produce alcohol, commonly via pyruvate, converted into ethanol and carbon dioxide; this process is frequently related to yeasts, including Saccharomyces and non-Saccharomyces yeasts [27,28]. Nonetheless, the fermentation process is not exclusive to yeasts; there are several fermentative bacteria, such as lactic acid bacteria, acetic acid bacteria, and native microbiota present in raw materials, which participate in the fermentation of several food and beverage industries [3,29,30,31,32]. In this regard, bacterial fermentation holds special interest due to the increasing demand for fermented produce, which serves as a potential source of rich biomass and bioactive compounds, offering consumer benefits [33,34,35].

During fermentation, the process requires that fermentative bacteria start their metabolic activity, which commonly represents entering a new environment, and bacteria must adapt their genetic content in response to the surrounding conditions [36]. In this regard, there are some factors involved in the process, including temperature, pH, as well as the natural composition of the raw material, which induces the metabolic response of microorganisms [37], dealing with changes in the metabolic pathways, such as amino acids production; Liu, Zhang, Liu, Xu, Shang, Xia, Song and Liu [30] studied the changes in bacterial communities in fermented foods such as Chinese watermelon-soybean paste, revealing that microbial abundance may vary due to fermentation conditions, dealing with the diversity and richness of metabolites in the fermented food, and showing a positive correlation of Bacillus species with the production of all free amino acids and dipeptides, and the presence of 11 major metabolic pathways for the metabolism of tyrosine, arginine and proline, glycerophospholipid, sphingolipid, cofactor biosynthesis, alanine, aspartate, and glutamate; amino sugar and nucleotide sugar metabolism; as well as phenylalanine, tyrosine, and tryptophan biosynthesis. Temperature and time are important factors during fermentation, which may affect the results and composition of the fermented matrix. Zhao et al. [38] reported that lactic acid bacteria cells increased their levels at low salt fermentation and low temperature, contributing to acid production during the first 45 days of the process, as well as the increase in l-glutamine, l-ornithine, β-D-fructose, d-arabinose, d-gluconic acid, glycine, l-proline, and guanine as a product of the enrichment of 28 metabolic pathways, mainly for glycine, serine, and threonine metabolism, and aminoacyl-tRNA biosynthesis. As mentioned before, raw material composition plays an important role in the results of fermentation because it represents the “food” for the bacteria; in this regard, it is crucial to understand that the presence of essential nutrients, such as carbon sources, which are not necessarily highly diverse, contributes to a better response because there are some microorganisms that show a better growth and productive response in the presence of a sole carbon source and specific amino acids [39].

An example of fermentative bacteria is Lactiplantibacillus plantarum, which is considered a promising probiotic bacterium, given its potential for fermentation and high adaptability, improving the quality and properties of fermented foods [2]. The versatility of this microorganism resides in its genetic content; Lactiplantibacillus plantarum has a genome size of around 3.1 Mbp, which presents diverse genes associated with diverse metabolic functions, such as carbohydrate transport and metabolism, amino acid transport and metabolism, energy production and conversion, and other genes that contribute to the potential for vitamin production, biotin, and alpha- and beta-glucosidase, related to probiotic activity [40,41,42]. The versatility of fermentative bacteria such as Lactobacillus plantarum has been evaluated, reporting its ability to redirect around 44% of its gene expression according to each specific growth habitat, particularly by regulation of metabolic activity, including the metabolism of carbohydrates, pyruvate, energy production, and conversion, transport, and metabolism of amino acids, nucleotides, lipids, and inorganic ions. Pyruvate metabolism is essential for fermentative bacteria; in this regard, growth conditions may deal with marked differences in the gene regulation for its utilization, upregulating the expression of the pyruvate dehydrogenase complex, as well as the stress response against antimicrobial compounds produced during fermentation, demonstrating the metabolic versatility of these kinds of microorganisms and their potential for fermentation processes [43,44].

Although the metabolic versatility of single strains such as Lactobacillus or Bacillus is well-documented, current strategies are shifting towards the use of synthetic microbial communities that leverage synergistic interactions to overcome monoculture limitations, thereby addressing challenges in reproducibility and process efficiency [45,46]. Unlike monocultures, these multi-species consortia leverage synergistic interactions that enhance system stability against environmental fluctuations such as pH, temperature, and substrate concentration [46]. Furthermore, such interactions can trigger the activation of silent gene clusters, facilitating the discovery and overproduction of novel specialized metabolites unattainable by single strains [47]. By mimicking natural ecological niches, these strategies optimize metabolic cross-feeding, offering a superior alternative for sustainable bioactive compound production.

4.2. Enzymes Involved in Fermentation

Enzymes are considered environmentally friendly biological catalysts, revolutionizing food preparation methods. Their use is widespread across various global food industries, including dairy, brewing, meat, baking, beverages, cereals, legumes, oils, and fats (

Table 1. Microbial enzymes application in the food industry [21,22,51,52,53,54].

Microorganisms	Enzymes	Substrate	Application	Role
A. Niger, P. notatum, B. amyloliquefaciencs, B. stearothermophilus, B. licheniformis, S. cerevisiae, A. awamori, Rhizopus oryzae, Gluconacetobacter, Acetobacter xylinus, Komagataeibacter xylinus, Fusarium sp., B. brevis	α-Amylases	Grapes, rice, cereals	Beverages	Juice treatment, low-calorie beer, clarification of fruit juice.
	Cellulases			Degrade plant cell walls. In wine production is used to increase yield and quality.
	Esterase			Enhancement of flavor and fragrance in fruit juice.
	Glucoamylases			Convert the starch into maltose and fermentable sugar. Use in sake and light beer production.
	Laccases			Modification of color appearance and wine stabilization.
	Pectinases			Clarification of fruit juice
	Proteases			Improves fermentation of beer.
	Xylanases			Release arabinoxylans and lower oligosaccharides, reducing the muddy appearance and viscosity of the beer.
Lactobacillus acidophilus, B. mesentericus, S. boulardii, S. ellipsoideus, P. ostreatus, S. diastaticus, L. brevis, L. fermentum, R. oryzae, R. oligosporus, L. plantarum, A. oryzae, A. niger	α-Amylases	Wheat, maize, sorghum, millet, rice, soybean	Cereals and legumes	Increase the total starch in barley and peas. Decrease amylose content in rice. Decrease carbohydrates in sorghum.
	Arabinoxylanases			Decrease insoluble fiber.
	Cellulases			Low crude fiber in pearl millet Fiber decreases in sorghum and yellow maize.
	Lipases			Decreased fat in mung beans, pigeon peas, red beans, soy and wild vigna species, maize, and rice. Increase in fat in pearl millet.
	Polyphenol oxidases			Decrease tannins
	Proteases			Increase in some essential amino acids and IVPD in maize and sorghum. Increase in protein accumulation in pearl millet.
	Tannases			Decreases tannins in beans, oats, and sorghum.
	Xylanases			Decrease fiber in sorghum
Lactobacillus bulgaricus, Lactococcus lactis, L. acidophilus, L. cremoris, L. casei, L. paracasei, L. thermophilus, L. kefiri, L. caucasicus Penicillium camemberti, P. roqueforti, Acetobacter lovaniensis, Kluyveromyces lactis, S. cerevisiae, A. Niger, A orzyae, B. subtilis, S. boydii, B. subtilis	Catalases	Milk and casein	Dairy Products	Removes H2O2 and eliminate the effect of volatile sulfhydryl that is responsible for the cooked/off-flavor in ultra-pasteurized milk.
	Lactases			Lactose hydrolysis, whey hydrolysis.
	Lipases			Cheese flavor.
	Proteases			Protein hydrolysis, milk clotting, low-allergenic infant food formulation, enhanced digestibility and utilization, flavor improvement in milk and cheese.
Bacillus megaterium, Bacillus subtilis	Amine oxidases	Fish proteins	Aquatic products	Inhibit biogenic amine accumulation, which is responsible for decreasing the quality and safety of fish-fermented products
	Decarboxylases			Degrade saturated fatty acids, which influence flavor.
	Esterases			Enhance favorable texture (hardness, gumminess, springiness, and chewiness), flavor, and aroma properties.
	Glucosidases			Release aromatic compounds from flavorless precursors.
	Lipases			Contribute to the development of flavor in the products due to the degradation of lipids to free fatty acids.
	Lyases			Produce flavor substances
	Proteases			Can develop different fermentation outcomes, some of which improve the product, while others may not help and might be detrimental. Generating peptides with antioxidant and antibacterial activities.
	Transferases			Produce flavor substances
L. sakei, L. curvatus, L. plantarum, Leuconostoc carnosum, Leuconostoc gelidium, B. licheniformis, E. faecalis, E. hirae, E. durans, Bacillus subtilis, L. divergens, L. carnis, E. cecorum, B. lentus, T. longibrachiatum, A. niger, A. oryzae, S. aureus	Papain	Meat proteins	Meat	Myofibrillar degradation of as well as collagen proteins helps to tenderize meat
	Polyphenol oxidases			Improve textural characteristics of meat products.
	Proteases			Tendering tough buffalo and sheep meat.
	Transglutaminases			Modify the texture of meat and meat products

). Incorporating enzymes and microorganisms in food processing is a well-established traditional practice [18,22]. Microbial enzymes have been utilized in various industries, including fuel [48], human health [49], and soil [50], and are crucial to the food industry due to their greater stability than plant and animal enzymes. They can be produced cost-efficiently through fermentation processes, requiring less time and space [51].

Their high consistency also allows for easy process modifications and optimizations that improve vitamins, essential amino acids, proteins, food appearance, flavors, enhanced aroma, and reduced anti-nutrients [22].

4.3. Enzymes Used in the Food Industry

4.3.1. Amylase and Glucoamylase

Amylases are generally categorized into two main types based on their enzymatic class: hydrolases (Enzyme Commission, EC 3) and transferases (EC 2). Within the hydrolase class, they are further divided into two primary groups: endoamylases and exoamylases [55].

Glucoamylases, also called saccharifying enzymes, are within the group of exoamylases due to their ability to hydrolyze α-1,4 glycosidic bonds from the non-reducing starch, malto-oligosaccharides, and related substrates, releasing β-D-glucose [56]. They convert the starch to maltose and fermentable sugars. These enzymes are also produced from Saccharomyces cerevisiae during the fermentation with glucose to obtain ethanol. Also, glucoamylases are essential in brewing sake and soy sauce and creating light beer. These enzymes break down dextrins, transforming them into fermentable sugars, which result in beer with lower calorie content and reduced alcohol levels [51,56].

On the other hand, α-amylase (EC 3.2.1.1) is part of the family of starch-degrading endoamylase enzymes, which catalyzes the hydrolysis of α-1,4 glycosidic linkages in polysaccharides, producing short-chain dextrins. This enzyme is commonly synthesized by various organisms, including Archaea, fungi, bacteria, and animals [51]. Amylases have diverse applications in food processing, such as brewing, livestock feed, baking, fruit juice production, starch syrups, and starch liquefaction. Specifically, α-amylase breaks starches of flour into fermentable sugars, which are then utilized by yeast during bread production to enhance the bread’s taste and quality. This enzyme also helps slow staling when incorporated into bread-making [21].

4.3.2. Proteases

Proteases are crucial in commercial and industrial applications and catalyze the hydrolysis of peptide bonds of proteins to peptides [57]. They represent a large and diverse group of hydrolytic enzymes classified by their site of action, enzyme active site structure, and specific reaction mechanisms [58]. Proteases are categorized depending on their action sites and along polypeptide chains into exopeptidases and endopeptidases. Exopeptidases target chain ends, while endopeptidases cleave internal bonds. Endopeptidases are further grouped into six types according to the catalytic residue in their active site: serine endopeptidases, aspartic endopeptidases, cysteine endopeptidases, metalloendopeptidases, glutamic endopeptidases, and threonine endopeptidases [51,59].

Proteases are produced by diverse organisms, including plants, Archaea, fungi, bacteria, and animals, and are widely used in various food industry sectors, such as brewing, dairy, baking, food processing, and animal feed [21]. These microbial enzymes perform diverse biochemical, physiological, and regulatory roles and have been essential in producing traditional fermented foods [57]. The characteristic flavor of fermented products is highly related to proteolysis due to hydrolysates generating precursor compounds, which are related to some important flavor substances in fermented products [60]. Proteases are produced by diverse organisms, including plants, Archaea, fungi, bacteria, and animals. They are widely used in various food industry sectors, such as brewing, dairy, baking, food processing, and animal feed [21]. These microbial enzymes perform diverse biochemical, physiological, and regulatory roles and have been essential in producing traditional fermented foods [57]. The characteristic flavor of fermented products is highly related to proteolysis due to hydrolysates generating precursor compounds, which are related to some important flavor substances in fermented products [60].

4.3.3. Lipases

Lipases (EC 3.1.1.3) are a class of enzymes that catalyze the hydrolysis of long-chain triglycerides. They are widely distributed in animals, plants, and microorganisms. However, microbial lipases find an interesting role in biotechnology, as many of them are stable over a wide range of pH, at elevated temperatures, and in organic solvents. They signify the most important group of biocatalysts for industrial applications [61,62]. These enzymes can modify the properties of lipids by altering the location of fatty acid chains in the glyceride and replacing one or more fatty acids with new ones [61,63].

In the food industry, they are applied to enhance flavors in dairy products, particularly by hydrolyzing milk fats to produce desirable cheese flavors. In bread dough, they hydrolyze triglycerides into diglycerides, monoglycerides, and fatty acids, which improve softness, volume, over-fermentation tolerance, and shelf life [63].

4.3.4. Catalase

Catalase (EC 1.11.1.6) is an oxidoreductase enzyme that reduces reactive oxygen species, particularly hydrogen peroxide, produced during aerobic respiration, thereby serving as an antioxidant and protecting cells from oxidative stress [64]. It is widely used in the food industry, often with enzymes like glucose oxidase for food preservation. In milk processing, catalase removes peroxide; in egg whites, it eliminates glucose; and in baking and food packaging, it helps prevent oxidation and reduces perishability. Its application in cheese production is limited [51].

4.3.5. Cellulases and Xylanases

Cellulases are hydrolytic enzymes that catalyze the cleavage of β-1,4 glycosidic bonds in glucose chains, transforming cellulose into cello-oligosaccharides and glucose via chemical or enzymatic hydrolysis. This category includes endoglucanases (EC 3.2.1.4), exoglucanases (EC 3.2.1.91), and β-glucosidases (EC 3.2.1.21) [65]. Similarly, microorganisms produce xylanases to break down xylans, a key hemicellulose component. Three main enzymes—endoxylanases, exoxylanases, and β-xylosidases—work together to degrade the xylan structure. Specifically, endoxylanases (EC 3.2.1.8) cleave β-1,4 bonds within xylan, while exoxylanases (EC 3.2.1.37) act on the non-reducing ends, producing xylooligosaccharides [66].

In the baking industry, cellulase and xylanase, along with amylases, lipases, and oxidases, enhance dough softness and reduce stickiness by breaking down insoluble cellulose and arabinoxylans in wheat flour into simple sugars, thereby improving flavor and texture in products like bread and cookies [51,67].

These enzymes are also widely used in brewing and fruit juice production, often combined with pectinases and amylases. During brewing, grains such as barley, wheat, and corn undergo mashing and malting to activate enzymes that ferment starch into alcohol. In both brewing and juice processing, cellulases and xylanases, along with pectinases, aid in breaking down cell walls and releasing sugars, nutrients, pigments, and aromatic compounds. These macerating enzymes are essential for producing fruit juices and wine [18,65,66].

4.3.6. Lactases (β-Galactosidase)

Lactase, a hydrolase enzyme known as β-galactosidase, is sourced from plants, animals, and microorganisms. Microbial β-galactosidases are widely used in food technology to hydrolyze lactose in milk and its by-products, particularly beneficial for individuals with lactose intolerance. These enzymes break down β-galactopyranosides (like lactose) and produce galactooligosaccharides (GOS), which offer prebiotic health benefits. Additionally, β-galactosidase aids in producing lactose-based sweeteners from the high-lactose effluents of cheese production [51,68].

4.3.7. Tannases

Tannase, or tannin acylhydrolase (EC 3.1.1.20), is an extracellular inducible enzyme that catalyzes the hydrolysis of some tannins and gallic acid esters [69]. Tannases are widely applied in the food, brewing, and pharmaceutical industries. They are found across animal, plant, and microbial sources, with microbial tannases preferred for industrial applications. These enzymes hydrolyze tannins by breaking ester and depside bonds, releasing glucose and gallic acid [70]. In the food industry, tannases improve instant tea by enhancing compound extractability and cold-water solubility. Additionally, tannases prevent haze and undesirable phenolic compounds in beer and wine, improving quality. The enzyme also reduces haze and bitterness in fruit juices. In animal feed, tannases degrade anti-nutritional tannins in agro-industrial by-products, enhancing feed quality [69,71].

4.3.8. Esterases

Esterases are hydrolase enzymes that catalyze the formation and breakdown of ester bonds. Their regioselectivity, stereospecificity, and stability in organic solvents make them highly valuable for various industrial applications [72]. They are essential in food and alcoholic beverages, mainly for modifying oils and fats in fruit juices and creating flavors and fragrances. A key group, feruloyl esterases, breaks the ester bond between ferulic acid and polysaccharides in plant cell walls. By hydrolyzing lignocellulosic biomass, these enzymes are also crucial for waste management [51].

5. Bioactive Compounds Obtained from Microbial Fermentation

5.1. Carotenoids

Carotenoids are a group of compounds related to the pigments yellow, orange, red, and purple, mainly found in fruits, vegetables, and plants [73]. Carotenoids have a 40-carbon structure with double conjugated bonds and a polyene chain end that varies depending on whether or not an oxygen atom is present; in this sense, carotenoids are classified into two groups, carotenes and xanthophylls [73]. Carotene comprises hydrocarbons and carotenoids such as α-carotene, β-carotene, and lycopene [73,74]. Xanthophylls are carotenoids with an oxygen atom on the end polyene chain that constitutes a hydroxyl, carbonyl, or aldehyde molecule. Some examples of xanthophylls are lutein, astaxanthin, and fucoxanthin [73]. These compounds have been reported to have a high antioxidant activity that can protect the organisms from oxidative stress caused by biotic and abiotic factors; this effect could be associated with their structure of double conjugated bonds. In this sense, it has been reported that carotenoids are related to potential health effects such as UV-protective, antiproliferative, antidiabetic, anti-inflammatory, and antiatherosclerosis responses.

Carotenoids are a group of compounds related to the pigments yellow, orange, red, and purple, mainly found in fruits, vegetables, and plants [73]. Carotenoids have a 40-carbon structure with double conjugated bonds and a polyene chain end that varies depending on whether or not there is an oxygen atom; in this sense, carotenoids are classified into two groups, carotenes and xanthophylls [73]. Carotene comprises hydrocarbons and carotenoids such as α-carotene, β-carotene, and lycopene [73,74]. Xanthophylls are carotenoids characterized by an oxygen atom on the end polyene chain that constitutes a hydroxyl, carbonyl, or aldehyde molecule. Some examples of xanthophylls are lutein, astaxanthin, and fucoxanthin [73].

These compounds have been reported to have a high antioxidant activity that can protect the organisms from oxidative stress caused by biotic and abiotic factors; this effect could be associated with their structure of double conjugated bonds. In this sense, it has been reported that carotenoids are related to potential health effects such as UV-protective, antiproliferative, antidiabetic, anti-inflammatory, and antiatherosclerosis responses [75,76]; therefore, carotenoids are compounds widely used in different industries such as medicine, food, and in pharmacology [77].

Microorganisms have gained popularity as an alternative source to plants to obtain multiple bioactive compounds simultaneously, using cheaper raw materials as carbon sources to reduce the cost of biorefinery [78]. Recently, various microorganisms have been studied for their carotenoid production; among these, photosynthetic bacteria, fungi, marine archaea, and yeast have been utilized to obtain lycopene, ketolases, α-carotene, β-carotene, and other carotenoids through biotechnology strategies [73,77,79,80]. The use of microorganisms as carotenoid producers could be due to the controlled cultivation, high efficiency, target compound synthesis, and reduced production period by metabolism modifications [81]. Biotechnology strategies such as metabolic engineering tools for the manipulation of biosynthetic pathways for carotenoid production coupled with the fermentation process followed by separation and purification methods have been widely studied to enhance the production and extraction of carotenoids from microorganisms, where the principal factors studied are pH of culture medium, temperature, process time, moisture content, aeration rate, carbon and nitrogen sources, light, carbon/nitrogen ratio, sonication, and chemical supplements, among other parameters [77,82,83]. The use of microorganisms as factories for producing carotenoids has been widely studied due to their diverse color tones, the ability to manipulate them, and low equipment requirements [84]. However, some disadvantages, such as the high production costs, have limited their industrial application. As a result, other biotechnological approaches are being explored to enhance carotenoid production and reduce costs using various microorganisms [78,83,85,86,87].

In this sense, it has been reported that fermentation technology has been used in conjunction with biotechnological manipulations such as metabolic engineering to produce carotenoids with high production yields, efficiency, moderate cost of production, and environmental friendliness in contrast to chemical synthesis and plant or animal source extraction due to the fast-growing strains and downstream techniques to extract the target products [77,83]. In this sense, microorganisms such as Blakeslea trispora, Chryseobacterium artocarpi CECT 8497, Flavobacterium sp. cells, Serratia marcescens, Phaffia rhodozyma, Rhodosporidium toruloides, Sporobolomyces, Sporidioblous, and Xanthophyllomyces Dendrorhous has been used to obtain anthraquinones, astaxanthin, β-carotene, canthaxanthin, flexirubin, lutein, and zeaxanthin by an optimized fermentation process [77,80,84,88,89], while other microorganisms, such as Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris, and Yarrowia lipolytica, have been modified through biotechnology strategies as well as different extraction processes to obtain carotenoids with high yields (Figure 3) [15,83,87].

On the other hand, the effect of fermentation on carotenoids from different food sources and by-products has been studied [85]. In this sense, orange, sweet potato, carrot, tomato, marigold red pepper, goji berries, and marine by-products, among other sources, have been studied to obtain astaxanthin, carotenes (α-carotene and β-carotene), lutein, lycopene, and zeaxanthin [78,85,86,90]. Likewise, fermentation factors such as matrix sources and fermentation conditions, as well as the microorganism studied, have been reported to enhance or maintain the carotenoid content; also, the use of some microorganisms could favor the carotenoid extraction and bioavailability by the production of enzymes that separate the carotenoids from the food matrix and facilitate the extraction with downstream strategies [85,86].

5.2. Essential Oils

Essential oils are bioactive compounds derived from plants composed mainly of low molecular weight volatile compounds such as terpenes and hydrocarbons [91,92]. These substances have attracted the attention of pharmaceutical and food industries due to their therapeutic [93], anticancer [94], antimicrobial [95], antioxidant [96], and food safety properties [97]. Also, essential oils contain compounds with strong odors, and the fragrance industry has also been interested in the obtention of these materials. The production of essential oils can be carried out by direct plant extraction, plant cell cultures, and microbial fermentation [98]. Specifically, the application of microbial fermentation can be classified into biotransformation (fungi or yeast transform compounds based on certain substrates), methods that use waste residues (food waste), and metabolic pathways followed by genetically modified microorganisms (Figure 4).

Besides these methods, there are a few recent reports regarding the de novo synthesis of essential oils by microorganisms. For example, fungal cultures (13 strains belonging to Aspergillus sp., Ceratocystis sp., and Neurospora sp.) produced interesting volatile compounds that can be found in essential oils from plants, such as citral and β-citronellol [99], which have been studied for their antifungal [100] and vasorelaxant properties in rats [101]. On the other hand, Sen et al. [102] reported that the interaction of agarwood (Aquilaria malaccensis) callus with the associated fungus Fusarium produced by fermentation led to a significant accumulation of terpenes such as geranyl isovalerate and tetrapentacontane, 1,54, dibromo-. Also, the infection of juvenile A. malaccensis plants coupled with associated Fusarium registered the presence of sesquiterpenes such as agarospirol, γ-eudesmol, and (−)-aristolene, which were exclusive of the fermentation with this fungus. In this sense, the main issue with the de novo synthesis of essential oil compounds by microorganisms is the accumulation of the obtained metabolites that can result in toxic environments for the organisms. This is the case of geraniol and citronellol production by the fungal genus Ceratocystis, whose removal procedures consisted of integrated bioprocesses that had to be applied to eliminate contaminated substances [103].

Regarding the use of waste residues for obtaining essential oils (or their compounds) through microbial fermentation, Mahmoud et al. [104] proposed a solid-state fermentation process that employs fungi, bacteria, and yeast strains (Aspergillus, Trichoderma, Bacillus, and Saccharomyces) to produce volatile compounds from basil leaf and stem waste. Several substances of importance for human health and the food industry were identified, such as ϒ-Bisabolene, isoprenyl cinnamate, diacetyl, and ethyl isovalerate. These compounds are present in natural essential oils from plants, and they have been studied for antioxidant [105], antimicrobial [106], antifungal [107], and food preservation [108] properties. Another therapeutic compound in essential oils is D-limonene, which can be obtained via the fermentation of olive mill waste by Rhizopus oryzae fungus and Candida tropicalis yeast [109]. The quantification of this substance revealed that R. oryzae produced a higher amount of D-limonene than C. tropicalis in controlled conditions: 87.73 µg/kg and 11.95 µg/kg, respectively. Therefore, agro-industrial waste residues are suitable for producing volatile compounds with health benefits. For example, rice bran oil waste is rich in ferulic acid, a vanillin precursor. Zheng et al. [110] reported a consecutive bioconversion process from ferulic acid of rice bran oil residue fermented by Aspergillus niger to vanillic acid, and then this broth was fermented by Pycnoporus cinnabarinus to obtain vanillin. A maximum concentration of this compound (2.8 g/L) with a molar yield of 61.9% at 72 h was registered.

In the case of biotransformation as a method for obtaining essential oils from microbial fermentation, this process is defined as the induction of the microbe to follow a specific metabolic pathway from an introduced precursor [111]. For that, geranic acid has been synthesized from geraniol by Rhodococcus sp., a group of bacteria isolated from soil. This synthesis was controlled by reaching a maximum conversion (54.6%) of geranic acid after 96 h of incubation at 30 °C as the optimal temperature for the reaction [112]. Besides these bacteria, yeasts have been reported to produce volatile compounds via bioconversion. Oda et al. [113] developed a metabolic pathway by Hansenula and Pichia yeasts using acetyl-CoA to synthesize citronellol, finding that the accumulation of citronellol in the incubating system could intoxicate yeasts; therefore, sodium acetate was added to control the production rate of citronellyl acetate for all strains. In this sense, yeasts can be affected by their metabolites in the production of the fragrance compound 2-phenylethanol (2-PE) from phenylalanine by Saccharomyces cerevisiae, in which the use of hydrophobic polymethylmethacrylate (PMMA) microspheres has been reported to physically remove the accumulation of phenylethanol [114]. This interesting volatile compound has been studied for treating psychiatric disorders such as depression [115].

Based on the above-mentioned, yeasts have been broadly studied to produce compounds of essential oils. However, metabolic engineering has been applied to modify these microorganisms at the genetic level. An increase in geraniol concentration (from 43.19 mg/L to 523.96 mg/L) via synthesis performed by manipulating enzymes GES and Erg20WW in S. cerevisiae was reported [116]. Also, this compound’s production was enhanced by modifying the site where the geraniol synthesis occurred in S. cerevisiae peroxisomes, which improved the geraniol titers by 80% [117]. Therefore, S. cerevisiae represents an appropriate “factory” for essential oil generation. Important substances such as limonene have been obtained by orthogonal engineering, in which a metabolic pathway was created by genetic modification in this yeast, producing significant amounts of limonene (917.7 mg/L) in fed-batch fermentation [118]. In addition to reported experimental trials, Werner et al. [119] reported in an in-silico study that modifying gene expression of central metabolic enzymes in S. cerevisiae can increase β-ionone yields up to 4-fold. Most microorganisms used for essential oil production belonged to the fungi kingdom; however, gene modification has been commonly applied in prokaryotic organisms. One of these microorganisms is E. coli, which has been altered to produce volatile compounds such as patchoulol [120], viridiflorol and amorphadiene [121], ionone [122], and others.

5.3. Phenolic Compounds

Phenolic compounds are bioactive compounds obtained from plants with a benzene ring with one or more hydroxyl groups. These products can be found in various natural foods, such as cereals, vegetables, fruits, and beverages such as wine, coffee, cocoa, and tea [123,124]. Phenolic compounds, including flavonoids, tannins, lignans, and stilbenes, are abundant plant compounds synthesized via the shikimate pathway using L-phenylalanine and L-tyrosine as precursors [125]. These compounds are present in various foods and beverages, contributing to their health benefits and bioactivity. Cocoa and its derivatives are among the most studied food matrices due to their high polyphenol content. Bacterial fermentation of cocoa beans significantly increases the release of conjugated phenolic compounds, such as caffeoyl aspartic acid and p-coumaroyl tyrosine, within 24 h of fermentation [126]. Using strains like Saccharomyces cerevisiae and Pichia kudriavzevii further enhances phenolic content, showcasing their potential for improving food bioactivity [127]. In microbial fermentation, phenolic acids undergo extensive biotransformation by the gut microbiota, producing smaller bioactive metabolites. Microbial fermentation processes significantly influence protocatechuic and vanillic acids derived from cocoa shells. During colonic fermentation, the gut microbiota transforms these compounds into metabolites such as benzoic, phenylpropanoid, and phenylacetic acids. Additionally, fermentation of cocoa by-products generates other phenolic compounds, such as caffeoyl aspartic acid, p-coumaroyl aspartic acid, clovamide, and p-coumaroyl tyrosine. These transformations are heavily influenced by the dietary fiber content in cocoa shells, which modulates the release and microbial accessibility of phenolic acids (Table 2) [128]. Similarly, coffee pulp fermentation enhances the production of phenolic metabolites such as phenylpropanoic acids and phenyl-γ-valerolactones, while reducing the overall concentration of polyphenols [129]. The fermentation process, driven by microbial activity, modulates these phenolic transformations and generates bioactive compounds like benzoic and phenylacetic acids. Additionally, dietary fiber and alkaloid content, such as caffeine and trigonelline, influence the bioavailability and release of phenolic compounds during fermentation [130].

Cereals naturally contain phenolic compounds predominantly in bound forms, which limits their bioavailability [131,132]. Specific processing techniques, such as microbial fermentation, are required to enhance the release of these antioxidant compounds and increase their free-form levels. Fermentation with ragi tape effectively enhances phenolic content in black glutinous rice. Over 72 h, free phenolics increased by 49%, and free-conjugated phenolics peaked at 48 h with an 8% rise. This process significantly improved antioxidant activity, as indicated by reduced IC50 values. Key phenolic compounds, including caffeic and ferulic acids, showed notable increases, highlighting the potential of ragi tape fermentation to boost the antioxidant properties of this rice variety [133]. Solid-state fermentation (SSF) of sorghum grain (SG) enhances sensory characteristics by modifying its polyphenol content. Various microbial strains (Lactiplantibacillus plantarum, Saccharomyces cerevisiae, Rhizopus oryzae, Aspergillus oryzae, and Neurospora sitophila) were used to assess their impact on polyphenols. After SSF, tannin and free phenolic contents were reduced by 56.36% and 23.48%, respectively. Cellulase played a crucial role in degrading tannins and phenolic compounds, while xylanase initially released flavonoids, although microbial consumption altered this effect over time. These findings highlight the potential of SSF to modify the polyphenolic profile and improve SG bioactivity and processing characteristics [134]. In a study on rye flour fermentation, the microbial community of germinated rye was enriched during lactic acid fermentation with increased terpenoid and phenolic compounds. Metabolomic analysis revealed notable changes in phenolic compound profiles, including the accumulation of bioactive polyphenols. Specifically, compounds such as ferulic acid, caffeic acid, and p-coumaric acid were identified post-fermentation, suggesting that sourdough fermentation can enhance the content of bioactive polyphenols, contributing to the health-promoting potential of fermented rye flour [135]. Studies consistently demonstrate that microbial fermentation significantly enhances the phenolic profile and antioxidant activity of cereals. While cereals naturally contain phenolics, a substantial portion exists in bound forms, limiting their bioavailability [136]. Fermentation releases these bound phenolics, as observed in black glutinous rice with ragi tape, sorghum, and sourdough-fermented rye, where free phenolics like caffeic, ferulic, and p-coumaric acids increased, boosting antioxidant activity. This process is attributed to microbial enzymes like feruloyl esterases, which hydrolyze bonds linking phenolic acids to cell walls [137].

Microbial fermentation is a crucial bioprocess for preserving fruits and vegetables. This biological method reduces the risk of contamination by producing antimicrobial compounds like organic acids, ethanol, and bacteriocins. Beyond preservation, fermentation enhances these foods’ nutritional value and creates new and desirable tastes and textures [138]. Citrus fruits, including their peels, are abundant sources of bioactive compounds such as phenolic acids, flavanones, and polymethoxylated flavones, alongside carotenoids and ascorbic acid [139,140]. Microbial fermentation of citrus peels, including orange, lemon, and grapefruit, was conducted using Lactobacillus plantarum and Lactobacillus acidophilus as fermentation agents. This process significantly enhanced the phenolic profile by transforming bound phenolics into more bioavailable forms. Key phenolic acids such as ferulic acid, caffeic acid, and p-coumaric acid were notably increased. Additionally, flavonoids like naringenin and hesperidin showed considerable elevation, improving antioxidant activity [141]. In the case of orange peel, SSF significantly increased the release of phenolic compounds. The fermentation process primarily resulted in the formation of flavonoid aglycones, such as naringenin, hesperetin, and nobiletin, replacing the glycoside hesperidin found in unfermented peels. Additionally, the fermented peel showed enhanced antioxidant activity [142].

In chili peppers, microbial fermentation increases both the quantity and diversity of phenolic compounds [143]. Lactic acid bacteria secrete enzymes like amylase, β-glucosidase, and phenolic acid decarboxylase, facilitating the release of bound polyphenols and enhancing antioxidant activity [144]. Caffeic acid, a key phenolic, rises during early fermentation stages but degrades into compounds like vinyl catechol and ethyl catechol as fermentation progresses [145]. Ferulic acid, another significant phenolic found in chili peppers and carrots, can be metabolized into vanillic acid and protocatechuic acid, highlighting the transformative potential of fermentation on polyphenolic profiles [146]. In chili peppers, microbial fermentation increases both the quantity and diversity of phenolic compounds [143]. Lactic acid bacteria secrete enzymes like amylase, β-glucosidase, and phenolic acid decarboxylase, facilitating the release of bound polyphenols and enhancing antioxidant activity [144]. Caffeic acid, a key phenolic, rises during early fermentation stages but degrades into compounds like vinyl catechol and ethyl catechol as fermentation progresses [145]. Ferulic acid, another significant phenolic found in chili peppers and carrots, can be metabolized into vanillic acid and protocatechuic acid, highlighting the transformative potential of fermentation on polyphenolic profiles [146].

Onion by-products are abundant in fiber and bioactive compounds, offering a valuable source for developing various bioproducts, including polyphenols [147]. A study assessed the potential of onion skins for polyphenol release through fermentation by different microorganisms, including bacterial and yeast strains. Results showed that fermentation with specific strains like Lactiplantibacillus plantarum and Saccharomyces cerevisiae increased the content of bioactive compounds, particularly quercetin aglycone, by up to 60% in yellow onion skins [148].

Wild herbs and plants are rich in phenolic compounds, particularly flavonoids [149]. Fermentation with microorganisms such as bacteria is known to increase the bioactive compounds in plants, particularly those with antioxidant and antibacterial properties [150]. For example, Achillea millefolium L. (yarrow) and Origanum majorana L. were studied as sources of phenolic-rich extracts, focusing on their impact on human gut microbiota and microbial metabolism. SSF enhanced the bioavailability of phenolic compounds like rosmarinic acid and caffeoylquinic acids. The microbial metabolism yielded significant metabolites, including phloroglucinol and 3,4-dimethoxyphenylacetic acid [151]. Beverages such as wine, beer, coffee, and tea are substantial sources of polyphenols, the most abundant antioxidants in the human diet [152]. Fermentation is an emerging method used to enhance the phenolic content in beverages, thereby improving their antioxidant properties. For instance, microbial fermentation of tea extracts using Trichoderma reesei, Aspergillus niger, and lactic acid bacteria (LAB) transformed pivotal bioactive compounds. Galloylated catechins were hydrolyzed, and organic acids were decarboxylated, increasing phenolic bioavailability. Additionally, alcohols and ketones accumulated, enhancing the aroma profile [153]. This highlights fermentation as a promising tool for improving both the phenolic composition and sensory qualities of tea beverages. Blueberry pomace, a by-product of juice processing, has shown significant potential as a matrix for phenolic enrichment through SSF. In a recent study, various fungal and LAB strains, including Aspergillus niger (AN), Lactobacillus acidophilus (LA), and Lactobacillus plantarum (LP), were utilized to enhance its polyphenol profile. Fermentation increased the content of key phenolic acids such as gallic acid, caffeic acid, and chlorogenic acid, as well as flavonoids like quercetin. Notably, anthocyanin levels decreased overall, except in pomace fermented by AN, where anthocyanidins showed an upward trend. Enhanced antioxidant activities were observed, with ABTS, DPPH, and FRAP radical scavenging capacities rising by 33.56%, 59.89%, and 87.82%, respectively. Additionally, simulated gastrointestinal digestion revealed improved bioaccessibility of polyphenols, underscoring SSF as an effective method for value-added utilization of blueberry pomace in functional food development [154]. Albino bilberry (Vaccinium myrtillus L.) juice [155] and Chinese rice wine [156] exemplify the impact of fermentation on enhancing phenolic composition and flavor complexity. In albino bilberry juice, fermentation with non-Saccharomyces yeasts resulted in increased phenolic acids (21.8–42.5%), flavonols (26.8–47.2%), and flavan-3-ols (4.9–74.5%), with novel flavonols synthesized during the process. Hanseniaspora uvarum yielded the highest phenolic enhancements, showcasing yeast metabolism’s role in antioxidant enrichment. Similarly, in Chinese rice wine brewed from five raw materials, liquid-state fermentation and grain liquefaction influenced flavor compounds and microbial diversity. Buckwheat-fermented wine stood out with the highest ester (27.39 mg/L), amino acid (1.47 mg/mL), and phenolic acid (904.29 mg/L) contents, contributing to its complex sensory profile characterized by honey, floral, and umami notes. Metagenomic sequencing revealed variations in microbial communities, with dominant genera including Saccharomyces, Aspergillus, and Bacillus. Together, these studies emphasize the importance of fermentation parameters and raw material selection in tailoring phenolic profiles, microbial ecology, and sensory attributes in functional beverages.

Fermentation can generate the synthesis of phenolic compounds, further increasing their content and antioxidant capacity [157]. However, the lack of standardized methodologies for analyzing phenolic compounds, the use of various microorganisms, and different extraction methods present opportunities to further improve the understanding and application of microbial fermentation in enhancing phenolic profiles [158,159,160]. Standardizing analytical techniques, selecting the appropriate microorganisms, and extraction methods are essential to maximizing the benefits of fermentation for food quality improvement.

Table 2. Summary of microbial fermentation treatments applied to various food matrices. The table high-lights the phenolic compounds that were enhanced as a result of fermentation, showcasing the po-tential of microbial processes in improving the bioavailability and antioxidant properties of phe-nolic compounds in food.

Food Matrix	Fermentation Treatment	Phenolic Compounds Increased	Reference
Cocoa shells	In vitro colonic fermentation	Caffeoyl aspartic acid, p-coumaroyl aspartic acid, clovamide, p-coumaroyl tyrosine	[128]
Coffee pulp	In vitro colonic fermentation	Phenylpropanoic acids, phenyl-γ-valerolactones	[129]
Black glutinous rice	Ragi tape fermentation	Caffeic acid, ferulic acid	[133]
Sorghum grain	SSF with various microbial strains	Tannins, free phenolics (caffeic, ferulic, p-coumaric acids)	[134]
Rye flour	Lactic acid fermentation	Ferulic acid, caffeic acid, p-coumaric acid	[135]
Citrus peels (orange, lemon, grapefruit)	Lactobacillus plantarum, Lactobacillus acidophilus	Ferulic acid, caffeic acid, p-coumaric acid, naringenin, hesperidin and nobiletin	[141,142]
Chili peppers	Lactic acid bacteria	Caffeic acid, ferulic acid	[143]
Onion skins	Lactiplantibacillus plantarum, Saccharomyces cerevisiae	Quercetin aglycone	[148]
Achillea millefolium L. and Origanum majorana L.	SSF with various microbial strains	Rosmarinic acid, caffeoylquinic acids, phloroglucinol and 3,4-dimethoxyphenylacetic acid	[151,161]
Tea extracts	Trichoderma reesei, Aspergillus niger, Lactic acid bacteria	Galloylated catechins, organic acids	[151,153]
Blueberry pomace	Various fungal and LAB strains (Aspergillus niger, Lactobacillus acidophilus, Lactobacillus plantarum)	Gallic acid, caffeic acid, chlorogenic acid, quercetin	[153,154]
Albino bilberry juice	Non-Saccharomyces yeasts	Phenolic acids, flavonols, flavan-3-ols	[154,155]
Chinese rice wine	Saccharomyces, Aspergillus, Bacillus	Phenolic acids	[155,156]

Table 2. Summary of microbial fermentation treatments applied to various food matrices. The table high-lights the phenolic compounds that were enhanced as a result of fermentation, showcasing the po-tential of microbial processes in improving the bioavailability and antioxidant properties of phe-nolic compounds in food.

Food Matrix	Fermentation Treatment	Phenolic Compounds Increased	Reference
Cocoa shells	In vitro colonic fermentation	Caffeoyl aspartic acid, p-coumaroyl aspartic acid, clovamide, p-coumaroyl tyrosine	[128]
Coffee pulp	In vitro colonic fermentation	Phenylpropanoic acids, phenyl-γ-valerolactones	[129]
Black glutinous rice	Ragi tape fermentation	Caffeic acid, ferulic acid	[133]
Sorghum grain	SSF with various microbial strains	Tannins, free phenolics (caffeic, ferulic, p-coumaric acids)	[134]
Rye flour	Lactic acid fermentation	Ferulic acid, caffeic acid, p-coumaric acid	[135]
Citrus peels (orange, lemon, grapefruit)	Lactobacillus plantarum, Lactobacillus acidophilus	Ferulic acid, caffeic acid, p-coumaric acid, naringenin, hesperidin and nobiletin	[141,142]
Chili peppers	Lactic acid bacteria	Caffeic acid, ferulic acid	[143]
Onion skins	Lactiplantibacillus plantarum, Saccharomyces cerevisiae	Quercetin aglycone	[148]
Achillea millefolium L. and Origanum majorana L.	SSF with various microbial strains	Rosmarinic acid, caffeoylquinic acids, phloroglucinol and 3,4-dimethoxyphenylacetic acid	[151,161]
Tea extracts	Trichoderma reesei, Aspergillus niger, Lactic acid bacteria	Galloylated catechins, organic acids	[151,153]
Blueberry pomace	Various fungal and LAB strains (Aspergillus niger, Lactobacillus acidophilus, Lactobacillus plantarum)	Gallic acid, caffeic acid, chlorogenic acid, quercetin	[153,154]
Albino bilberry juice	Non-Saccharomyces yeasts	Phenolic acids, flavonols, flavan-3-ols	[154,155]
Chinese rice wine	Saccharomyces, Aspergillus, Bacillus	Phenolic acids	[155,156]

5.4. Polysaccharides

The fermentation process induces the production of several types of polysaccharides by microorganisms. Some of these compounds possess properties that are beneficial for human health, such as antioxidant, antitumor, and anti-inflammatory activities [162,163]. Also, they are considered biocompatible and biodegradable materials with high yield and reproducible production [162].

In this sense, a high amount of research can be found in the literature reporting that lactic acid fermentation by bacterial strains (Lactobacillus, Leuconostoc, and others) produces exopolysaccharides. Exopolysaccharides are a specific type of extracellular biopolymer synthesized by bacteria and some fungi, having several health and physicochemical properties [164,165]. For that, Table 3 summarizes different exopolysaccharides with biological activities produced by microbial fermentation.

One of these substances is dextran, a long-chain glycopolymer reported as a functional molecule for medical purposes such as a plasma volume expander, a wall material for encapsulation, and an alternative to tomography markers [166]. Schmid et al. [167] stated that the properties of dextran are dependent on its molecular weight (i.e., length of the polymer chain). In that report, dextran polymers with high molecular weights (1.09 × 10⁸–1.86 × 10⁸ Da) produced by Liquorilactobacillus hordei exhibited different rheological properties, indicating that the dextran with the longest chain had the best capacity to form films and gels.

Similarly, Esmaeilnejad-Moghadam, Mokarram, Hejazi, Khiabani and Keivaninahr [166] found that dextran synthesized by Leuconostoc mesenteroides in milk permeate culture media possessed the lowest molecular weight and increased solubility as compared to dextran produced in broth medium. Besides its techno-functional properties, an appropriate antioxidant activity has been detected (by ABTS and DPPH analyses) in dextran obtained by the fermentation with Leuconostoc pseudomesenteroides isolated from the Juçara palm tree [168]. However, some bacteria cannot produce exopolysaccharides in fermentation conditions. As an example, Lactobacillus fermentum was investigated during the fermentation of longan pulp, finding that the polysaccharides of the fruit changed in composition and chemical structure because of the action of the bacteria. The polysaccharides from the fermented pulp were composed mainly of arabinose (49%), galactose (23%), glucose (14%), and other monosaccharides (~14%), in which immunomodulatory and prebiotic activities were detected [169].

Also, the prebiotic activity has been attributed to fructooligosaccharides that can be synthesized by the enzymes inulosucrase and endoinulinase present in bacteria such as Leuconostoc citreum and Aspergillus niger, respectively. In this study, sucrose was more effective than inulin as a substrate for the chain reaction to produce these prebiotic compounds [170]. On the other hand, the production of functional polysaccharides by fungi has been commonly studied in fermentation conditions involving Tremella spp. The basidiospore fermentation by Tremella aurantialba in tofu wastewater has been proposed as a method for obtaining adequate yields of polysaccharides (~15.02 g/L) whose composition was based on monosaccharides such as glucose and mannose [171]. Similarly, Ge et al. [172] examined Tremella fuciformis in fermentation conditions, finding that this fungus produced macromolecular polysaccharides (yield ≈ 9.0 g/L) that were composed of xylose, mannose, and galactose residues and had antioxidant properties determined by the scavenging capacity of superoxide anions and hydroxyl radicals analyses. Besides the antioxidant activity, neuroprotective [173], immunomodulatory [174], and antitumor [175] activities have been reported for polysaccharides produced by Tremella spp.

Regarding the fermentation by yeasts, Chen et al. [176] addressed the biological properties of polysaccharides obtained by the fermentation of a flower, Dendrobium officinale, carried out by Saccharomyces cerevisiae and Wickerhamomyces anomalous. Both yeasts produced four polysaccharides (comprised of mannose and glucose, having the following ratios: 3.31:1, 5.56:1, 2.40:1, and 3.29:1) that were isolated, possessing an enhanced antioxidant property; however, the anti-inflammatory activity was inadequate for the proposed experimental conditions.

Also, the production of endopolysaccharides by yeast has been reported for the fermentation of biodiesel-derived crude glycerol, in which yeast strains belonging to Debaryomyces sp., Naganishia uzbekistanensis, Rhodotorula sp., and Yarrowia lipolytica generated biopolymers with potential for pharmaceutical applications [177].

Furthermore, bacteria can produce polysaccharides of interest alongside other microorganisms. In this sense, the fermentation of okara (residue from soybean) by lactic acid bacteria Lactobacillus bulgaricus and fungi Neurospora crassa produced polysaccharides (composed mainly of galacturonic acid, galactose, and arabinose) with blood sugar regulation, glucose adsorption delaying, and prebiotic properties [178]. In another study, the yeast Saccharomyces cerevisiae and the bacteria Bacillus subtilis enhanced the polysaccharides obtained by the fermentation of wheat bran, indicating that the isolated products (composed mostly of galactose, xylose, and galacturonic acid) possessed suitable antioxidant properties (measured by the DPPH test) [179]. Therefore, the combination of bacteria with other microorganisms, such as fungi and yeasts, can result in the enhancement of polysaccharides produced by fermentation of different sources.

5.5. Polyunsaturated Fatty Acids (PUFAs)

Chemically, fatty acids are composed of hydrocarbon chains terminated by a carboxyl group at one end and a methyl group at the other [180]. They are categorized into groups depending on their degree of saturation: saturated fatty acids, which contain no double bonds between carbon atoms; monounsaturated fatty acids, characterized by a single double bond; and polyunsaturated fatty acids, which possess two or more double bonds [180,181,182]. Structurally distinguished by the position of their first double bond, the most biologically relevant classes are the omega-3 (n-3) and omega-6 (n-6) series [183]. These lipids can improve human health by maintaining cell membrane fluidity, supporting neurodevelopment, and regulating cardiometabolic and inflammatory responses [184]. As the human body lacks the enzymes to synthesize the parent compounds like α-linolenic acid (ALA) and linoleic acid (LA), these are essential nutrients that must be obtained from the diet [183]. However, relying only on traditional sources is becoming increasingly unsustainable. Deep-sea oils are the primary commercial source of PUFAs, and these marine resources are limited by seasonal variations and are contributing to the depletion of global fish stocks [185]. Furthermore, plant-based oil production faces challenges regarding competition for arable land and climatic dependency, necessitating alternative production routes [186].

In response to these ecological limitations, oleaginous microorganisms, defined as those accumulating over 20% of their dry cell weight as lipids, have emerged as a robust biotechnological alternative, often referred to as Single Cell Oils (SCOs) [186,187]. Unlike non-oleaginous species, the lipid accumulation in these microorganisms is driven by the enzyme ATP citrate lyase (ACL). And under nutrient-limited conditions (typically nitrogen), ACL plays the pivotal role of cleaving cytosolic citrate into acetyl-CoA, providing the essential precursors for fatty acid biosynthesis [188]. This metabolic capability is found across diverse taxa, including filamentous fungi and oleaginous yeasts. Yeasts such as Yarrowia lipolytica are particularly notable for their industrial versatility, capable of growing on recalcitrant agro-industrial residues like palm by-products, thereby supporting circular bioeconomy frameworks [189]. Regarding the oleic profiles, marine protists such as Schizochytrium sp. (Thraustochytrids) are primary industrial sources of docosahexaenoic acid (DHA), a crucial component for neural and retinal tissue development [190]. Moreover, filamentous fungi like Mortierella alpina are exploited to produce omega-6 fatty acids, like arachidonic acid (ARA) and gamma-linolenic acid (GLA). ARA acts as a vital precursor for eicosanoids involved in immune signaling [185], while GLA is recognized for its anti-inflammatory properties and potential to alleviate conditions such as atopic eczema [186,191]. This taxonomic diversity allows for the targeted production of high-value lipids that are metabolically expensive or impossible to source efficiently from terrestrial plants.

Furthermore, to establish microbial fermentation as a commercially viable strategy for commodity markets like animal feed, production costs must be reduced by enhancing not just total lipid yield but specifically the fraction of high-value PUFAs. Recent metabolic engineering efforts have focused on two distinct biosynthetic pathways. In Thraustochytrids like Schizochytrium sp., DHA synthesis is governed by a polyketide synthase (PKS) system, a multi-enzyme complex that synthesizes long-chain PUFAs directly without oxygen-dependent desaturation steps [186,192]. Recent studies demonstrate that overexpression of specific PKS subunits, such as the Open Reading Frame A (ORFA), significantly increases DHA content while reducing saturated fatty acids [193], a process dependent on activation by specific phosphopantetheinyl transferases (PPTase) [192]. In contrast, microalgae such as Phaeodactylum tricornutum utilize the aerobic desaturase/elongase pathway. In these systems, key desaturases like PtFAD2 and PtFAD6, whose targeted overexpression or regulation under stress conditions, such as cold shock, can drastically shift the lipid profiles toward eicosapentaenoic acid (EPA) accumulation [194,195]. These focused genetic interventions allow for the customization of microbial oils to meet specific nutritional requirements.

Despite these biotechnological strides, the industrial competitiveness of SCOs remains constrained by production costs, where feedstock procurement can account for up to 70% of operational costs [187]. Moreover, the oxidative instability of PUFAs poses a major challenge during downstream processing. While synthetic biology has improved product stability by engineering lipid droplets to encapsulate and shield these sensitive fatty acids from oxidation [196,197], efficient recovery poses a significant challenge, as traditional solvent extraction risks product oxidation and toxicity. Consequently, Supercritical Fluid Extraction (SFE) using CO₂ has emerged as a superior green alternative, offering high selectivity for high-value lipids without solvent residues [198]. Future success depends on integrating robust, genetically optimized strains with these sustainable processing technologies.

Table 3. Exopolysaccharides synthesized by microbial fermentation and their health properties.

Microorganisms	Specie	Exopolysaccharide	Health Properties	Ref.
Bacteria	Acetobacter xylinum	Levan	Antioxidant, anti-inflammatory	[199]
Bacteria	Lactobacillus plantarum	Glucose and galactose residues	Antioxidant	[200]
Bacteria	Paenibacillus polymyxa	Heteroglycan formed by (1→4) and (1→6) hexose residues	Antioxidant, immunomodulatory, mitogenic, allergenic, anti-inflammatory	[201]
Bacteria	Escherichia coli (modified with a Leuconostoc citreum gene)	Alternan	Encapsulation capability	[202]
Bacteria	Bacillus sp. isolated from fermented pickles	Glucose and galactose residues	Antioxidant	[203]
Fungi	Polyporus umbellatus	Three polysaccharides composed by mannose, galactose and glucose (molar ratios: 43.6:2.5:1.0; 17.7:3.1:1.0 and 4.6:2.6:1.0)	Antioxidant, immunological, cellular aging delaying, DNA damage protecting	[204]

Table 3. Exopolysaccharides synthesized by microbial fermentation and their health properties.

Microorganisms	Specie	Exopolysaccharide	Health Properties	Ref.
Bacteria	Acetobacter xylinum	Levan	Antioxidant, anti-inflammatory	[199]
Bacteria	Lactobacillus plantarum	Glucose and galactose residues	Antioxidant	[200]
Bacteria	Paenibacillus polymyxa	Heteroglycan formed by (1→4) and (1→6) hexose residues	Antioxidant, immunomodulatory, mitogenic, allergenic, anti-inflammatory	[201]
Bacteria	Escherichia coli (modified with a Leuconostoc citreum gene)	Alternan	Encapsulation capability	[202]
Bacteria	Bacillus sp. isolated from fermented pickles	Glucose and galactose residues	Antioxidant	[203]
Fungi	Polyporus umbellatus	Three polysaccharides composed by mannose, galactose and glucose (molar ratios: 43.6:2.5:1.0; 17.7:3.1:1.0 and 4.6:2.6:1.0)	Antioxidant, immunological, cellular aging delaying, DNA damage protecting	[204]

6. Applications of Bioactive Compounds Obtained from Microbial Fermentation

To provide a critical evaluation of fermentation-derived bioactives, this section prioritizes their analysis by specific fields of application: animal nutrition, agriculture, and human health. For each sector, the discussion links the bioactive compounds to their underlying molecular mechanisms, ranging from gut microbiota modulation to enzymatic inhibition, and evaluates their reported efficacy in recent in vitro and in vivo models.

6.1. Animal Feed

The use of fermented feed is gaining attention as a sustainable approach to improving livestock health while addressing environmental concerns. Fermented feed is a biologically modified product resulting from microbial fermentation that transforms feed components into microbial proteins, bioactive peptides, amino acids, and beneficial probiotics [205]. This process enhances the digestibility of complex carbohydrates, proteins, and fibers, improving nutrient availability and reducing the impact of anti-nutritional factors such as phytates and protease inhibitors [206,207]. Fermented feed is gaining attention as a sustainable approach to improving livestock health while addressing environmental concerns. Fermented feed is a biologically modified product resulting from microbial fermentation that transforms feed components into microbial proteins, bioactive peptides, amino acids, and beneficial probiotics [205]. This process enhances the digestibility of complex carbohydrates, proteins, and fibers, improving nutrient availability and reducing the impact of anti-nutritional factors such as phytates and protease inhibitors [206,207].

Gut microbiota is crucial in livestock health and productivity [208]. Fermented feed has been shown to positively modulate gut microbiota composition by increasing the abundance and diversity of beneficial microorganisms. This promotes optimal nutrient absorption, gastrointestinal health, and immune function while reducing disease incidence and oxidative stress [209,210]. This modulation of gut microbiota through fermented feed is increasingly recognized as a sustainable alternative to antibiotics, improving animal welfare and production efficiency [211]. It also mitigates oxidative stress by protecting the gastrointestinal tract, aiding recovery, and boosting stress resilience, supporting efficient and sustainable livestock production [209].

The growing interest in fermented feed stems from its demonstrated benefits across various livestock species, though outcomes are closely tied to inoculation protocols and substrate composition. For example, in Bamei piglets, the inclusion of a complete feed (enriched with 5% whey powder) fermented for 5 days with Lactobacillus plantarum and Bacillus subtilis (1.0 × 10⁹ CFU/mL) led to higher weight gain, better feed efficiency, and improved immunity. Notably, this high-dose mixed-strain fermentation was essential to achieve the reported elevated levels of immunoglobulins and reduced inflammatory markers. The fermented feed also positively impacted the gut microbiota, enhancing the diversity of beneficial bacteria such as Lactobacillus and Prevotellaceae, which are crucial for maintaining intestinal health and optimizing nutrient absorption [212]. Similarly, fermenting the plant-based fraction of their feed (corn and soybean meals) for 4 days (>1.0 × 10⁹ CFU/g) with Lactobacillus and Bacillus subtilis significantly improved growth performance, meat quality, and nutrient utilization in broiler chickens. However, these benefits demonstrated dose-dependency: additional benefits included reduced cholesterol content and better feed conversion ratios at 10% inclusion, whereas 5% inclusion maximized weight gain, showcasing its potential to enhance poultry production efficiency and product quality through optimized dietary formulation [205]. Similar specificity is required in aquaculture, where the utility of fermentation lies in its ability to biologically detoxify plant-based ingredients. Rather than simply enriching protein content, the process must target the hydrolysis of species-specific antinutritional factors, such as phytates, saponins, and non-starch polysaccharides, to unlock nutrient digestibility, modulate gut health, and promote growth [213]. Furthermore, replacing cornmeal with Psophocarpus tetragonolobus (winged bean tubers) has shown promising results in ruminant diets by modulating in vitro ruminal fermentation kinetics. Specifically, a 30% inclusion optimized volatile fatty acid profiles, with increased propionic acid levels and energy efficiency, proving that the tuber substrate itself can enhance degradability without prior fermentation. This highlights the potential of fermented tuber pellets as a sustainable alternative to traditional feed ingredients like cornmeal, supporting both animal health and productivity [214]. In poultry, a study on Xuefeng black-bone chickens demonstrated that the combination of 30-day anaerobically microbial fermented feed (using a consortium of Saccharomyces cerevisiae, Bacillus subtilis, Lactobacillus plantarum, and Enterococcus faecium) with ginseng polysaccharides (200 mg/kg) further enhanced growth performance. Crucially, this combination exhibited a synergistic effect superior to either treatment alone, significantly improving feed efficiency and immune function [215]. Notably, contrasting these findings with previous broiler studies reveals a key distinction: while improved feed conversion is consistent across species, lipid modulation is context-dependent; unlike in broilers, where fermentation alone reduced cholesterol [205], in Xuefeng chickens this effect required ginseng supplementation, highlighting that specific metabolic outcomes may rely on functional synergies.

Regarding potential adverse outcomes, the reviewed studies indicate that risks such as residual oxidative stress are largely mitigated when controlled inoculants are used. In fact, fermentation consistently reduced serum malondialdehyde (MDA) levels and upregulated antioxidant enzymes (SOD, GSH-Px) in both broilers and piglets, indicating a protective rather than deleterious cellular effect [205,212]. However, evidence suggests that adverse outcomes are primarily functional or process-dependent rather than intrinsic to the microbial metabolism. Uncontrolled fermentation may carry risks of accumulating undesirable secondary metabolites (e.g., mycotoxins or biogenic amines) and pathogenic contamination if quality assurance fails [213]. Furthermore, even with safe protocols, metabolic trade-offs exist: for instance, high-level replacement of corn with winged bean tubers in ruminant diets reduced total volatile fatty acid concentrations [214], while in broilers, exceeding a 5% dietary inclusion compromised maximal weight gain despite improved feed conversion, likely due to palatability thresholds [205]. Thus, while toxicity is rare in controlled settings, diminishing returns and metabolic shifts at high inclusion rates represent the primary limitations needing mitigation.

These studies demonstrate the broad applicability of microbial fermentation across species such as swine, poultry, ruminants, and fish, yet they also underscore that efficacy relies on tailoring protocols to specific substrates and physiological targets to maximize gains while mitigating metabolic trade-offs. By shifting from generic applications to precision strategies that align inoculation doses and processing times and functional synergies with biological needs, fermented feeds represent a vital tool in advancing eco-friendly and highly efficient livestock management.

6.2. Agricultural Use

Microbial products offer a sustainable alternative to traditional agricultural chemicals and fertilizers by enhancing crop yields and improving soil health. Companies increasingly use microorganisms as biocontrol agents and biofertilizers through carrier-based inoculants, which enrich the soil by producing essential nutrients through their metabolic activities. Building upon these benefits, the critical role of microbial metabolites has been associated with the promotion of plant growth and the enhancement of disease resistance. This supports plant growth by enhancing nutrient availability, such as nitrogen, phosphorus, and potassium, while improving soil properties and boosting beneficial bacteria. These applications not only decrease reliance on chemical inputs but also contribute to soil biodiversity and health, which is crucial for maintaining long-term agricultural sustainability [216,217,218].

Recent studies have highlighted the pivotal role of microbial metabolites in plant growth promotion and disease resistance. Plant growth-promoting rhizobacteria produce metabolites such as auxins, cytokinins, and gibberellins, which enhance nutrient uptake and stimulate plant growth. However, these hormonal effects are concentration-dependent: while optimal levels promote elongation, excessive microbial auxin production can alter the plant’s endogenous hormonal balance, potentially inhibiting root growth depending on the host’s sensitivity [219,220]. Beyond growth promotion, these metabolites also improve disease resistance by inhibiting pathogens and inducing systemic resistance. Plants further influence their microbiomes by secreting specific compounds that recruit beneficial microbes to suppress pathogens and enhance plant immunity [221]. These insights demonstrate the potential of microbial metabolites to support resilient cropping systems while reducing dependency on chemical inputs.

In addition to promoting disease resistance, microbial metabolites also play a key role in helping plants cope with environmental stresses such as drought, salinity, and temperature extremes by enhancing plant physiology, boosting antioxidants, and improving water retention, ultimately increasing productivity under stress [222,223]. Exopolysaccharides secreted by soil bacteria improve soil aggregation and water retention, promoting plant hydration during drought by enhancing water infiltration [224]. Furthermore, under broad abiotic constraints like salinity and thermal stress, osmoprotectants like trehalose and proline produced by microbes function beyond simple osmotic adjustment; they act as chemical chaperones that stabilize membrane integrity and scavenge reactive oxygen species, thereby mitigating oxidative damage. Crucially, the efficacy of this mechanism is governed by the plant genotype, as the capacity to integrate microbial osmolytes for osmotic adjustment varies significantly among species, determining the final degree of stress tolerance [225]. Microbial compounds also enhance plant antioxidant systems, scavenging reactive oxygen species to prevent oxidative damage and preserve cellular integrity [226]. In addition, some metabolites induce the expression of stress-responsive genes, enabling better adaptation and tolerance to abiotic stresses [227].

Given their benefits, the adoption of microbial metabolites in agriculture represents a sustainable shift from traditional chemical fertilizers and pesticides, promoting eco-friendly farming practices. However, a critical comparison highlights a key trade-off: while chemical inputs offer immediate and consistent yields, biological agents are living systems susceptible to environmental variability, often resulting in less predictable field performance. Despite these challenges, microbial inoculants derived from beneficial bacteria and fungi, combined with renewable feedstocks in microbial fermentation, enhance soil fertility, support plant health, and reduce reliance on chemical inputs, contributing to sustainability by minimizing dependence on non-renewable resources [228]. Furthermore, microbial biotechnology bolsters soil biodiversity and health, with managed soil microbiomes significantly improving crop productivity and resilience [229].

Recent advancements in biotechnology, including the development of engineered microbes, have enhanced the production of microbial metabolites, providing sustainable alternatives to traditional fertilizers and pesticides by efficiently converting renewable feedstocks into biofertilizers and biopesticides [230]. These biotechnological advancements not only enhance microbial metabolite production but also significantly contribute to improving agricultural efficiency. By optimizing fermentation conditions and selecting suitable microorganisms, the yield and effectiveness of these metabolites have been enhanced. For example, engineered microbes are capable of converting renewable feedstocks into high-value biofertilizers and biopesticides, which support eco-friendly agricultural practices [231]. Additionally, the use of genetic engineering techniques has led to the development of tailored microbial strains that produce specific metabolites at higher concentrations, improving agricultural efficiency [232]. Microorganisms significantly important in agriculture have long been recognized as an effective and eco-friendly alternative in modern farming, reducing the dependence on synthetic fertilizers and pesticides [233]. Furthermore, biotechnological advancements in microbial applications support sustainable agriculture by minimizing reliance on non-renewable resources, as they utilize renewable feedstocks in fermentation processes. As these technological advances continue to evolve, they further drive the adoption of microbial-based practices, ultimately ensuring a more sustainable and productive agricultural system [234].

6.3. Human Health

Fermentation stands out as a promising alternative for improving health due to its content of bioactive compounds, which are linked to the metabolic and biotransformation activities carried out by microorganisms [235]. The transformation of the natural components of the raw material during fermentation results in the production of enzymes that cause the transformation of complex compounds into simple biomolecules, giving rise to compounds with biological activity beneficial to human health (Figure 5) [138,236].

Dairy fermentations such as milk, yogurt, cheese, cream, and ice cream contain probiotic bacteria, and so do some meats, sausages, bread, and cereal products [237]. The necessary amount of these viable microorganisms in a food to exert health benefits varies between 10⁸ and 10⁹ colony-forming units (CFU) per day. Their effectiveness depends on the type of microorganism and the physiological conditions of the consumer [238]. The mechanisms of action associated with the health effects of these microorganisms include competitive inhibition of pathogenic bacteria proliferation (through alterations in pH levels and a reduction in oxygen availability) and non-competitive inhibition via the production of bacteriocins [239]; synthesis of essential micronutrients (vitamins, amino acids, and enzymes) enhances the bioavailability of compounds [240]; and stimulation of the host immune system by promoting the production of interleukin-10 (IL-10) and immunoglobulin A (IgA) antibodies [241]. They also play a crucial role in protecting intestinal epithelial cells against inflammation and associated disorders by regulating the production of antibodies, lymphocytes, interleukins, cytokines, and chemokines [242]. Additionally, probiotic microorganisms present in fermented foods have been reported to produce the enzymes β-galactosidase and lactase, which help combat lactose intolerance by converting glucose and galactose into short-chain fatty acids [243].

On the other hand, the fermentation of whey protein has gained significant attention in recent years, as it enables the enzymatic hydrolysis of peptides that can act as bioactive compounds. This process is made possible by the activity of proteases, which generate low molecular weight peptides (˂10 kDa) with specific compositions that allow them to enter the cell nucleus and interact directly with DNA-associated proteins regulating gene expression [244,245]. These bioactive peptides have been linked to the positive regulation of cell proliferation in human cell cultures and the inhibition of growth in various types of cancer cells. Additionally, they have demonstrated immune system-like functions, such as promoting lymphocyte proliferation, antibody production, and cytokine regulation [246]. Furthermore, they stimulate the phagocytic capacity of macrophages and inhibit the secretion of specific cytokines. Antidiabetic, antihypertensive, and xanthine oxidase inhibitor activities are also reported by these bioactive molecules [247].

Following this, kefir is a fermented lactic acid derivative with characteristics similar to yogurt. Its main components, such as lactic acid bacteria, organic acids, polysaccharides, and bioactive peptides, have significant health benefits [248]. Several studies have demonstrated its antioxidant, anti-inflammatory, anti-hypertensive, anticancer, and antidiabetic properties. Additionally, kefir has been associated with therapeutic effects on bone health, the immune system, cognitive function, and the gut microbiota [249,250,251,252,253].

Continuing with lactic fermentations, but now in solid-state form, bioactive peptides from various cheese varieties have been shown to provide health benefits by acting as antioxidants, antihypertensives, and antidiabetics [254]. For example, in a study where individuals with hypertension consumed 30 g/day of Italian cheese (Grana Padano), a significant reduction in blood pressure (both systolic and diastolic) was observed after two months [255]. In another study, six different types of cheese were evaluated, including Gouda, which showed the best results in antioxidant activity, ECA inhibitory activity, and DPP-IV enzyme inhibition. The study concluded that consuming 10–20 g of Gouda cheese, as part of a balanced diet, could be sufficient to obtain health benefits. This finding underscores that bioefficacy is not only dose-dependent but also strain-specific; different starter cultures (e.g., Lactobacillus vs. Lactococcus) possess distinct proteolytic systems that cleave casein at specific sites, resulting in unique peptide profiles with varying degrees of ACE-inhibitory potency [256]. Another important compound in this type of lactic fermentation is conjugated linoleic acid. Reports have shown an increase in these bioactive levels during cheese maturation, which could enhance its potential health benefits, including antidiabetic, anticancer, anti-atherosclerotic, and antihypertensive effects [257].

On the other hand, soy fermentation is an important source of bioactive peptides with significant health benefits. For example, peptides obtained from tempeh fermentation have demonstrated antioxidant, anti-inflammatory, and antihypertensive effects [258,259]. Crucially, the bioavailability of these peptides is dictated by the fermentation type; SSF is particularly effective as it allows filamentous fungi like Rhizopus spp. To penetrate the soybean matrix with their hyphae. This physical penetration, combined with the secretion of specific proteases, facilitates a deeper hydrolysis of proteins into low-molecular-weight bioactive peptides compared to liquid fermentation systems [259,260]. Saponins and phytosterols, which are bioactive compounds naturally found in soy, have gained interest due to their medicinal properties [260]; regarding saponins, the presence of these compounds in some fermented foods has been associated with obesity prevention, positive regulation of the immune system, antiviral effects, and antitumor properties [261]. At the same time, sterols can inhibit cholesterol absorption in the intestine, potentially reducing the risk of cardiovascular diseases [262]. Another important compound in soy is gamma-aminobutyric acid (GABA), which acts as a neurotransmitter in the central nervous system; its health benefits include lowering blood pressure, promoting relaxation, and improving mood [262]. Fermented soy food has been reported to be a rich source of GABA due to the production of enzymes during fermentation, which facilitates its transformation through the action of glutamic acid decarboxylase [262]. Soy is also naturally rich in vitamins, minerals, and fiber, which are crucial for human growth and proper metabolism [259]. In addition to this, another important aspect of soy is its high content of allergenic proteins and other allergens, such as Gly m 1, P28, and P34 [263]. Current research confirms that fermentation with Rhizopus spp., Aspergillus oryzae, and Bacillus subtilis effectively degrades these allergenic epitopes via enzymatic hydrolysis. However, the efficiency of this reduction is not uniform; it is governed by strain-specific proteolytic activities and the fermentation method (e.g., solid-state vs. liquid). Furthermore, regarding consumption, the resulting immunomodulatory benefits, such as the shift in the Th1/Th2 balance and suppression of IgE, appear to be dose-dependent, highlighting the need to establish standardized intake levels to maximize safety and therapeutic efficacy [259,260,263]. Also, bioactive compounds such as phenolic acids, flavonoids, and isoflavones have been identified in soy-derived fermentations [264]. Some of these fermented products originate from Meju, a dry soybean block fermented with fungi and Bacillus sp., which has been attributed with anticancer potential due to the presence of trypsin inhibitors, isoflavones, vitamin E, and unsaturated fatty acids [265,266]. Additionally, Doenjang extracts have been linked to the activation of the enzyme glutathione S-transferase and the increased activity of natural killer cells [267].

It is important to mention that isoflavones are the main compounds responsible for the anticancer potential of fermented soy products, as they have demonstrated beneficial therapeutic effects in cell lines of various types of cancer, including stomach, colon, lymphoma, pancreas, prostate, breast, and neuroblastoma [266]. Furthermore, these compounds have been associated with cardiovascular health benefits, increasing HDL cholesterol levels while reducing LDL cholesterol and triglyceride levels [268]. On the other hand, the anti-inflammatory effects of soy fermentation have been widely studied. It has been found that consuming foods such as miso and soy sauce reduces serum levels of IL-6 and IL-18 and inflammatory markers, including high-sensitivity C-reactive protein (hs-CRP) [269]. Some studies have also linked isoflavones to neuroprotection, promoting neuronal regeneration and enhancing existing neuronal functions. Therefore, soy fermentations represent an important source of bioactive compounds, particularly genistein, with great potential in preventing various diseases [270].

Fermentation has also been used to develop natural cosmetics with potential benefits for skin health. In this regard, studies on Camellia sinensis var. Assamica have shown that the activity of enzymes produced during the fermentation process can promote the release of bioactive compounds with moisturizing properties (such as amino acids and sugars); these compounds may help improve skin hydration and support its barrier function [271]. Additionally, an increase in the content of compounds like gallic acid and epigallocatechin gallate (EGCG) has been observed in the extract of Camellia sinensis tea, which could enhance its antioxidant and anti-inflammatory properties, which may help protect the skin from damage caused by free radicals and UV radiation, delaying skin aging and contributing to cancer prevention [272].

On the other hand, solid-state fermentation of phenols has been reported to improve the nutritional quality and antioxidant properties of various legumes and cereals (rice, wheat bran, corn, oats, rye, and millet). This process involves enzymes such as amylases, proteases, and lipases, which hydrolyze polysaccharides, proteins, and lipids into products with lower toxicity, improved texture, flavor, and aroma, as well as a reduction in antinutritional compounds such as phytic acid, tannins, and gas-producing compounds [258].

According to several studies, a wide variety of phenolic compounds have also been identified in liquid fermentation, such as wine, including catechins, p-coumaric acid, resveratrol, rutin, quercetin, myricetin, anthocyanins, tannins, and flavan-3-ols, as well as several phenolic acids such as caffeic, ellagic, syringic, vanillic, and ferulic acids [273]. Additionally, the fermentation process of red wine leads to the formation of compounds like melatonin and hydroxytyrosol, which have been strongly associated with the numerous health benefits of this beverage [274,275]. The bioactive compounds in red wine have been extensively studied in both in vitro and in vivo investigations, demonstrating a wide range of health benefits, including antioxidant, antibacterial, anti-inflammatory, anticancer, and antidiabetic properties [276,277,278,279,280]. Other notable effects include antithrombotic, antidepressant, and neuroprotective activities, microbiota regulation, and anti-obesity effects, as they influence adipose tissue metabolism, hypocholesterolemia, and endothelial function [281,282,283,284].

Vinegar is a highly valued fermented product obtained from fruits such as grapes, apples, pomegranates, and cranberries through a fermentation process that can occur via two different pathways: alcoholic and acetic [285]. The microorganisms commonly used in this process belong to the Acetobacter and Komagataeibacter species, which have been associated with increased bioactive compounds such as polyphenols and organic acids [3]. Among these, acetic acid is considered the most important component of vinegar. In this context, apple cider vinegar has been shown to provide various health benefits, including improvements in cognitive, liver, and reproductive function, and therapeutic effects in diabetes [286,287,288,289,290]. It also exhibits antioxidant, antimicrobial, anti-inflammatory, and anti-obesity properties. Another significant effect studied is its ability to combat hypercholesterolemia by improving total cholesterol levels, triglycerides, LDL cholesterol, total cholesterol/HDL-C ratio, and LDL-C/HDL-C ratio [291]. These effects have been linked to bioactive compounds such as catechins, p-hydroxybenzoic acid, gallic acid, caffeic acid, p-coumaric acid, and chlorogenic acid [292].

Kombucha is a fermented liquid that is notable for its potential health benefits, including antioxidant, antitumor, hepatoprotective, and antidiabetic properties. It is prepared by fermenting black or green tea (derived from the Camellia sinensis plant) using a symbiotic culture of yeasts, acetic acid bacteria, and lactic acid bacteria [293,294]. This product contains bioactive compounds such as catechins, theaflavins, thearubigins, and vitamins B1, B6, B12, and C, which are associated with its reported health benefits [295,296]. In this context, in vivo studies have demonstrated the antidiabetic potential of kombucha, showing effects such as reduced blood glucose levels, increased plasma insulin, and modulation of gluconeogenic and glycolytic enzyme activity in experimentally induced diabetic rats [297]. Additionally, this beverage has been shown to inhibit the activity of enzymes like α-amylase and α-glucosidase, leading to a slower rate of blood glucose absorption [298]. The antiproliferative activity of kombucha has also been evaluated in various cancer cell lines, including MCF-7 (breast cancer), A549 (lung cancer), and HCT8, HCT 116, and CACO-2 (colon cancer) [293,299]. Furthermore, studies suggest it may exhibit selectivity toward cancer cells while sparing normal lung cells [300]. On the other hand, this type of fermentation has also been shown to reduce the levels of pro-inflammatory markers, including nitric oxide (NO), tumor necrosis factor (TNF), and interleukin-6 (IL-6) in macrophages activated by LPS. Additionally, it has been found to inhibit the enzyme 15-lipoxygenase (15-LOX), a key mediator in inflammatory processes [299,301]. However, the interpretation of these findings requires caution due to experimental inconsistencies. Studies indicate that anticancer efficacy is strictly substrate-dependent (e.g., green vs. black tea) [300] and that variations in fermentation parameters can drastically alter metabolic profiles, limiting the reproducibility of cytokine modulation effects [299].

Another important liquid fermentation is black tea, made by a unique microbial fermentation process involving some basic functional microorganisms of the genera Aspergillus, Bacillus, Candida, Cyberlindnera, Klebsiella, Lactobacillus, Penicillium, and Rasamsonia, among others. Compounds such as alkaloids, polyphenols, polysaccharides, and volatile compounds have been identified in this beverage, where caffeine stands out as one of the most abundant alkaloids and catechin in the group of polyphenols [302,303]. Multiple in vivo studies have shown that black tea has health benefits, for example, by reducing blood glucose levels and lowering the risk of diabetes by up to 45% in people with regular consumption of black tea (2–3 g per day) [304]. It has also been associated with significant weight and fat loss, improving the lipid profile and reducing hyperlipidemia in humans [305]. It has also been shown to ameliorate chemically induced colitis in mice by modulating the gut microbiota, to decrease insulin resistance and chronic kidney disease in rats by modulating insulin signaling and increasing Nrf2 expression [306], and to exert a hepatoprotective effect by modulating hepatic oxidative stress, inflammatory response, and gut microbiota dysfunction in mice [307], and it possesses potent lipid-lowering activity in a high-fat zebrafish model [308]. Additionally, in some in vitro and in vivo studies, it showed potential to prevent the onset of cardiological and neurodegenerative diseases associated with its interaction with proteins of the signaling pathways of these diseases, so this fermentation has potential for the prevention and treatment of chronic diseases [309,310].

Despite promising in vitro data, a translation gap persists for fermented beverages. Clinical trials on apple cider vinegar show significant heterogeneity, where outcomes depend more on intervention duration and baseline health than on bioactive concentration [291]. Similarly, the in vivo efficacy of kombucha and teas is constrained by the low bioavailability of polyphenols (e.g., theaflavins) and potential toxicity risks, indicating that health benefits require standardized therapeutic windows rather than simple dose-dependent consumption [294,304].

7. Integrative Challenges: One Health, Regulations, and Industrial Scalability

The broad spectrum of applications discussed in this review operates within a unified biological continuum, aligning with the One Health strategy. Interventions in one sector inevitably impact the others; specifically, the use of fermentation-derived biostimulants serves as a critical tool to reduce the environmental load of xenobiotics in the soil-plant interface. Recent evidence by Porras et al. [311] highlights that SSF not only enhances the extraction of bioactive compounds but also provides a sustainable pathway to replace chemical fertilizers, preventing their accumulation in the food chain. Simultaneously replacing prophylactic antibiotics with functional fermented feeds mitigates the selection pressure for antimicrobial resistance in livestock, creating a biological barrier against the transmission of resistant pathogens to humans [312]. Thus, fermentation technologies act as a measurable mechanism for the reduction of veterinary antibiotic consumption and serve as a key indicator of success.

However, translating these biological benefits into global industry practice requires navigating a complex regulatory dichotomy. Currently, traditional fermentation using defined, non-genetically modified (non-GMO) consortia, particularly LAB, represents the most mature route for immediate scale-up. As noted by Bergsma et al. [313], these microorganisms benefit from “Qualified Presumption of Safety (QPS) status in the European Union, facilitating their rapid approval as bioprotective agents. Conversely, advanced strategies relying on precision fermentation of genetically modified microorganisms (GMMs) face stricter scrutiny under comprehensive safety frameworks. D’Amore et al. [314] emphasize that new ingredients and processes must undergo rigorous risk assessments to ensure they do not introduce hazards, a process that significantly slows market entry compared to established QPS cultures. Consequently, the immediate future of the bioeconomy lies in optimizing wild-type consortia to bridge the gap between high-yield innovation and regulatory compliance.

8. Perspectives

The use of microorganisms as potential biofactories of bioactive compounds using biotechnological strategies such as fermentation, coupled with other techniques such as metabolic engineering tools, is on the rise due to advantages such as reduced use of chemicals and water and ease of cultivation and separation of compounds. Also, bioactive compounds obtained from microorganisms are considered to have a natural origin, and the commercial market has become more popular in contrast to chemical synthesis [15,31,78,84]. Likewise, the bioactive compounds such as carotenoids, essential oils, phenolic compounds, and polysaccharides, among others, obtained through microbial fermentation have been used for nutraceutical enrichment of animal feed [315], as well as the production of protein-rich animal feed using different sources such as agro-industrial and food wastes [207]. In recent years, microbial fermentation has been explored as an alternative to using agrochemicals to protect crops and reduce plant diseases caused by bacteria, fungi, nematodes, and yeast. Biostimulants have been developed to enhance the growth, yield, and defense of various crops [311,316]. In human health, microbial fermentation has been studied for the extraction of bioactive compounds as an alternative to traditional chemical extraction for the enrichment of food or for use as a nutraceutical because these compounds have been linked to the proper functioning of the organism and the reduction in non-communicable diseases [313,317,318].

Despite the use of microorganisms to obtain bioactive compounds using biotechnology strategies, research needs to continue developing efficient processes to release and enhance the extraction of the bioactive compounds because fermentation parameters such as fermentation time, solid-to-liquid ratio, temperature, pH, microorganism used, carbon source, and downstream techniques to separate the molecules of interest need to be optimized to reduce the high cost of the technologies and to guarantee an appropriate bioprocess to achieve high yield and selectivity and avoid microbial contamination during the fermentation process [1,22,84,162]. Specifically, addressing economic limitations requires sustainable scale-up strategies. A promising approach is the valorization of low-cost agro-industrial by-products, such as molasses, whey, or fruit pomace, to replace expensive synthetic media. This substitution not only mitigates operational costs but also upcycles environmental waste into high-value bioactive ingredients, thereby establishing a profitable circular economy model [312].

Critical analysis suggests that maximizing consistency requires specific metabolic targeting. For instance, SSF is superior for enzymes and peptides as it mimics fungal niches [259,260], whereas standardized submerged fermentation favors liquid-soluble antioxidants [299]. Moreover, employing controlled stress strategies such as low temperature and salt reduction can strategically enhance bioactive profiles [38], and shifting from spontaneous to defined microbial consortia is essential to stabilize metabolic outputs against environmental fluctuations [294].

9. Conclusions

Bioactive compounds, including alkaloids, phenolic compounds, pigments, vitamins, and others, are of interest in many industries for their potential as chemopreventive agents against many chronic diseases, such as cancer, cardiovascular diseases, and diabetes. They can also prevent aging-related diseases. These metabolites are mainly found in plants, and for their use, the extraction techniques often produce high quantities of toxic waste from solvents such as hexane, methanol, and acetonitrile, among others. Microbial fermentation is a sustainable tool for obtaining high-quality metabolites from various plant and animal matrices and producing them through biotechnology to achieve a higher yield of these compounds. These metabolites can be used in agriculture, as well as in animal and human health. The potential use of these metabolites nowadays relies on the biotechnological techniques implemented to produce each group of metabolites.