Abstract
The global nutraceutical industry is experiencing a paradigm shift, driven by an increasing demand for functional foods and dietary supplements that address malnutrition and chronic diseases such as obesity, diabetes, cardiovascular conditions, and cancer. Traditional plant- and animal-derived nutraceuticals face limitations in scalability, cost, and environmental impact, paving the way for microbial biotechnology as a sustainable alternative. Microbial cells act as bio-factories, converting nutrients like glucose and amino acids into valuable nutraceutical products such as polyunsaturated fatty acids (PUFAs), peptides, and other bioactive compounds. By harnessing their natural metabolic capabilities, microorganisms efficiently synthesize these bioactive compounds, making microbial production a sustainable and effective approach for nutraceutical development. This review explores the transformative role of microbial platforms in the production of nutraceuticals, emphasizing advanced fermentation techniques, synthetic biology, and metabolic engineering. It addresses the challenges of optimizing microbial strains, ensuring product quality, and scaling production while navigating regulatory frameworks. Furthermore, the review highlights cutting-edge technologies such as CRISPR/Cas9 for genome editing, adaptive evolution for strain enhancement, and bioreactor innovations to enhance yield and efficiency. With a focus on sustainability and precision, microbial production is positioned as a game-changer in the nutraceutical industry, offering eco-friendly and scalable solutions to meet global health needs. The integration of omics technologies and the exploration of novel microbial sources hold the potential to revolutionize this field, aligning with the growing consumer demand for innovative and functional bioactive products.
Introduction
The evolving global health landscape is characterized by a significant rise in diseases associated with malnutrition, largely driven by the increasing consumption of junk food. Conditions such as obesity, diabetes, cardiovascular disorders, and cancer have spurred a growing consumer interest in functional foods and nutraceutical products designed not only to nourish but also to prevent or mitigate disease [1,2].
Nutraceuticals, derived from food sources, offer health benefits beyond basic nutrition and encompass therapeutic properties such as antioxidant, anti-inflammatory, and antimicrobial activities [3]. Coined by Stephen DeFelice in 1989, the term “nutraceutical” merges “nutrition” and “pharmaceutical”, reflecting its dual role in health and therapy. These products are often described as dietary supplements delivering concentrated bioactive agents in non-food matrices at dosages exceeding those obtainable from regular food [4]. Unlike whole foods, nutraceuticals focus on isolated bioactive components, blurring the line between nutrition and pharmacology.
A more refined definition by [5] emphasizes nutraceuticals as substances cultivated, extracted, or synthesized under controlled conditions that, when administered orally, restore altered body structures and functions, improving overall health and well-being. This potential to prevent and treat chronic diseases, including cardiovascular diseases, cancer, and neurodegenerative disorders, underscores the growing global interest in nutraceuticals.
Traditionally, plant and animal sources have been the primary means of obtaining nutraceuticals. However, these sources face challenges such as seasonal variability, high extraction costs, and limited scalability [6]. In contrast, microbial production offers year-round consistency, easier genetic manipulation, and lower production costs, making it an attractive alternative. Microorganisms have emerged as critical platforms for producing bioactive compounds such as probiotics, peptides, polyunsaturated fatty acids (PUFAs), and polyphenols [5].
Microbial-based nutraceuticals hold immense promises for addressing the limitations of plant- and animal-based production. These techniques rely on fermentation technologies, where microorganisms either directly synthesize bioactive compounds or convert substrates into value-added products. This approach simplifies production, reduces costs, and enhances scalability. Microbial cells also serve as ideal hosts for genetic engineering, enabling the optimization of fermentation processes (Figure 1) and the utilization of simple carbon sources for efficient metabolite production [7]. Microbial nutraceuticals encompass a wide range of bioactive compounds, including vitamins, oligosaccharides, peptides, and pigments, many of which are already being commercially produced. These compounds offer sustainable and efficient alternatives to traditional sources and align with the global push for eco-friendly production practices.
This review will explore the current technologies, challenges, and future trends in microbial nutraceutical production. By focusing on these bio-based solutions, it aims to highlight how fermentation-driven microbial platforms are transforming the nutraceutical industry, ensuring sustainability and scalability while addressing global health challenges.
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Where Biology Meets Engineering: Scaling Up Microbial Nutraceuticals to Bridge Nutrition, Therapeutics, and Global Impact
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Selected chapter, chapter 16:
Relevance and Challenges of Commercial Nutraceutical Production
The commercial production of nutraceuticals is expanding rapidly, driven by increasing consumer demand for functional foods and health-enhancing supplements [117]. However, scaling up microbial bioproduction presents several challenges, particularly in optimizing microbial strains and improving manufacturing efficiency. Microbial strains must be carefully selected and studied to ensure that they can withstand processing conditions, utilize cost-effective substrates such as molasses, whey, starch, and agro-industrial residues, and maintain consistent productivity [18,100]. A well-controlled fermentation process is essential to achieving high yields and product quality. Techniques such as batch, fed-batch, and perfusion fermentation are commonly used, with strict monitoring through Process Analytical Technology (PAT) tools to maintain precision and reproducibility [37,91]. Despite advances, limitations in microbial metabolism and process efficiency still hinder large-scale production.
To overcome these challenges, advanced biotechnological approaches are being explored to enhance strain performance and streamline production. Omics technologies, including genomics, transcriptomics, proteomics, and metabolomics, provide valuable insights into microbial metabolism, enabling the development of more robust strains with improved productivity [18,88,89].
Analyzing host genomics has provided insights into how the host genome shapes microbiome diversity and influences host phenotypes [121]. Likewise, metabolomics sheds light on the metabolic functions of the gut microbiome and its impact on the host gut metabolome [122]. Consequently, a multi-omics approach that integrates metagenomics and untargeted metabolomics at both intra- and inter-domain levels has uncovered interactions between host and microbial metabolites, offering valuable insights into the microbiota’s role in aging [122]. Additionally, genomics plays a key role in identifying probiotic potential within commensal microbes [123], while metagenomics helps decipher the interactions between probiotics and the microbiome [124]. The integration of transcriptomics, proteomics, and metabolomics provides a comprehensive profile of host–microbe interactions and the effects of probiotic supplementation on the host [125]. Transcriptomics, particularly through RNA sequencing or microarray gene expression analysis, is crucial for determining the immunomodulatory properties of probiotics. Meanwhile, proteomics utilizes protein chips containing antibodies, nucleic acids, or other protein-binding molecules to assess proteome changes [126]. The application of multi-omics techniques facilitates the purification of antimicrobial peptides from microbes through liquid chromatography–tandem mass spectrometry (LC–MS/MS), while integrated genomics and proteomics analyses help characterize the genes responsible for these compounds [127]. This approach also supports evolutionary studies through comparative genomics and the discovery of novel antimicrobial compounds. Mass spectrometry plays a critical role in identifying antimicrobial peptide sequences and their molecular masses within complex mixtures, providing insights into their structural and functional properties [128,129,130]. The next section will discuss these innovative strategies and their role in optimizing nutraceutical production.
16.1. Strain Selection for Nutraceutical Production
Several studies have focused on improving fermentation techniques, emphasizing enhanced functionality, better hygiene procedures, increased yields, and the standardization of fermentation processes using specific starter strains [5]. Generally Recognized as Safe (GRAS) microorganisms are widely used in industry due to their safety and versatility in pharmaceutical and food applications. Microbial cell factories such as S. cerevisiae, Y. lipolytica, A. pullulans, Corynebacterium glutamicum, E. coli, Lactiplantibacillus plantarum, and Lactiplantibacillus rhamnosus offer significant advantages for producing high-value nutraceuticals, including polyphenols, omega-3 fatty acids, vitamins, and probiotics [118,119]. The regulation of genetically modified microorganisms (GMMs) for nutraceutical production presents ethical, legal, and social challenges. Lawmakers, regulatory bodies, and industry stakeholders must address these concerns as biotechnology laws continue to evolve. The challenge lies in balancing technological advancements with safety, fairness, and public confidence [131,132,133]. The World Health Organization (WHO) plays a key role in setting international biotechnology standards. In the United States, multiple federal agencies regulate biotechnology, each with specific responsibilities. The Food and Drug Administration (FDA) oversees the approval of biotechnology products related to human health, such as genetically modified drugs, vaccines, and gene therapies. This process includes extensive pre-market testing, clinical trials, and post-market monitoring to assess potential risks, including allergic reactions, unintended genetic modifications, and long-term health effects [131,132,133].
In Europe, biotechnology regulation is managed at both national and European Union (EU) levels. The European Medicines Agency (EMA) regulates gene therapies, cell-based treatments, and biologics. The EU has some of the world’s strictest regulations for GMMs, and public resistance remains high in several member states. Regulatory approaches to GMMs vary across countries [132,134]. Canada, Australia, and Japan also have their own regulatory frameworks. However, the absence of global agreement on key issues such as GMM oversight, biotechnology patent rights, and ethical concerns related to gene editing makes establishing uniform international standards challenging [132,134].
16.2. Synthetic Biology and Heterologous Expression
Synthetic biology serves as a cornerstone of modern nutraceutical production, offering unprecedented control over microbial biosynthetic potential. It has revolutionized the production of complex nutraceuticals by enabling the creation of novel metabolic pathways in microorganisms. One of the most significant advantages of synthetic biology is its ability to combine genes from different organisms, creating biosynthetic pathways that do not naturally exist in a single organism. For example, the heterologous expression of plant genes in yeast as a microbial host has facilitated the yeast synthesis of plant natural products (PNPs) such as artemisinic acid and farnesene [120,135]. Traditionally extracted from plants, these compounds can now be produced at scale using engineered yeasts, eliminating issues like seasonal availability and low yields from natural sources. Furthermore, E. coli has been engineered with genes from various plant species to produce PNPs such as resveratrol, a compound known for its anti-inflammatory and antioxidant properties [136,137].
Another notable achievement in synthetic biology is the ability to rewire metabolic fluxes to boost the production of key intermediates. For instance, S. cerevisiae has been optimized to increase the production of artemisinic acid, a precursor to the antimalarial drug artemisinin, by redirecting carbon flux toward the mevalonate pathway [138]. Such advances make microbial platforms viable alternatives to plant-based extraction, reducing environmental impact and enhancing sustainability.
Synthetic biology has also enabled the production of a wide range of nutraceuticals. Flavonoids such as naringenin and bioactive peptides have been successfully synthesized in microbial hosts by introducing heterologous biosynthetic pathways [79]. Compared to traditional extraction methods, microbial production offers a more efficient, cost-effective, and scalable process. Furthermore, synthetic biology allows for improvements in the bioavailability and stability of nutraceuticals. Advanced techniques, such as enzyme engineering and pathway optimization, have led to higher yields, improved purity, and enhanced functional properties of the final products, addressing the growing global demand for nutraceuticals [107,139].
Through advanced synthetic biology techniques, researchers can redesign or construct entirely new metabolic networks in microbial hosts like E. coli and S. cerevisiae, enabling the efficient production of high-value nutraceuticals such as polyphenols, carotenoids, and bioactive peptides. These compounds, often difficult or inefficient to extract from natural sources, can be sustainably and scalably produced through microbial fermentation [140]. An additional advantage of this approach is the use of non-food lignocellulosic feedstocks, such as agricultural residues and municipal waste. These alternative substrates provide an environmentally friendly and cost-effective solution for generating valuable bioactive compounds, reducing reliance on traditional food crops and enhancing scalability [89].
16.3. Advances in CRISPR/Cas9 Technology
One of the most transformative technologies in microbial genetic engineering is CRISPR/Cas9, a genome editing tool that allows for precise modifications of microbial DNA. This tool enables scientists to tailor microbial metabolism for enhanced nutraceutical production by knocking out undesirable genes or introducing new biosynthetic pathways, thereby improving the efficiency of bioactive compound production. CRISPR/Cas9 has been developed to enable precise genetic editing, optimizing production pathways and stoichiometry while addressing challenges such as expanding carbon utilization, reducing metabolic burden, and enhancing strain stability. For instance, S. cerevisiae has been successfully engineered using CRISPR/Cas9 to boost the production of resveratrol [141].
In addition, CRISPR/Cas9 was successfully employed in a comprehensive metabolic engineering study to expand the carbon fermentation capabilities of S. cerevisiae to include glycerol [142]. This strategy demonstrated promising applications for utilizing biomass resources in bioethanol production [143,144]. Moreover, a similar CRISPR/Cas9 approach enabled S. cerevisiae to convert glycerol into 2,3-butanediol [145]. These engineered S. cerevisiae strains hold significant potential as platforms for redirecting metabolic fluxes toward nutraceutical production, particularly when targeting production from agricultural residues and acid-catalyzed glycerolysis [144].
Notably, Y. lipolytica efficiently utilizes glycerol natively for citric acid production [146]. Furthermore, the application of CRISPR/Cas9 genome editing and UV mutagenesis enhanced the conversion of glycerol to erythritol, a nutraceutical compound, achieving production titers as high as 150 g/L, with yields of 0.62 g/g and productivities of 1.25 g/L/h [147]. Ongoing research and development in such recombinant strains, as well as further synthetic pathways or heterologous expression of effective genes, are crucial for overcoming current limitations and achieving commercial viability.
Therefore, the capability of synthetic biology and heterologous expression pathways not only enhances production efficiency but also contributes to the sustainability of nutraceutical production by reducing reliance on traditional food-based feedstocks. However, despite the efficiency gains provided by CRISPR/Cas9, significant challenges remain, particularly in the regulatory landscape governing genetically modified organisms (GMOs). In markets with stringent labeling and approval requirements, such as the European Union, regulatory hurdles can delay or complicate the commercial use of CRISPR-modified organisms. Addressing these regulatory barriers will be critical for expanding the commercial use of CRISPR/Cas9 in nutraceutical production. Effective collaboration with regulatory agencies, transparent safety testing, and consumer education will be essential to overcome these challenges and foster broader acceptance of GMO-derived nutraceuticals.
16.4. Adaptive Evolution: Enhancing Microbial Efficiency
Adaptive evolution is an effective strategy in microbial strain development that leverages selective pressure to enhance microorganisms’ nutrient utilization and metabolite production capabilities. Unlike targeted genetic engineering, this method simulates natural selection by subjecting microbial populations to specific environmental stressors, leading to the development of strains with improved traits.
In adaptive evolution, microorganisms are cultivated under controlled conditions where selective pressures, such as nutrient scarcity or exposure to toxic byproducts, promote the survival of mutants with advantageous adaptations. These mutations can result in increased metabolic efficiency, enhanced tolerance to stressful conditions, or elevated production of target compounds like polyphenols, probiotics, and fatty acids. Research has demonstrated the successful application of adaptive evolution to enhance microbial strains for nutraceutical production. For example, S. cerevisiae has evolved to tolerate high ethanol concentrations—a toxic byproduct of fermentation—significantly boosting bioethanol production [148]. E. coli has been adapted to enhance GABA production using glycerol as the carbon source [69]. A heavy-ion mutagenesis was used in adaptive evolution of Aurantiochytrium sp. and increased the production of DHA from 0.18 to 0.27 g/L/h and the titer yield from 21 to 27 g/L [90].
One of the key advantages of adaptive evolution is its potential to enhance microbial performance without extensive genetic modification. This method promotes the natural development of beneficial traits, which can then be integrated with genetic engineering techniques for further optimization. Moreover, adaptive evolution is applicable to a broad range of microorganisms, including those that are challenging to modify using conventional genetic tools. Another benefit is the long-term stability of the evolved traits. Once a microbial strain adapts to a particular environment, its enhanced characteristics are generally preserved over successive generations, making adaptive evolution a robust and dependable method for strain improvement in industrial applications [149].
16.5. Fermentation Technologies for Nutraceutical Production
Fermentation remains the backbone of microbial production for nutraceuticals, providing a controlled environment where microorganisms can efficiently produce bioactive compounds such as polyphenols, probiotics, and fatty acids. Bioreactors enable precise control over factors such as temperature, pH, and oxygen levels, optimizing growth conditions and enhancing production efficiency. Advances in fermentation methods have improved yields, reduced costs, and facilitated scalability to meet global demand for nutraceuticals [139,141,150].
Batch fermentation is one of the most traditional and widely used methods for producing nutraceuticals. In this closed system, all ingredients are added at the beginning, and fermentation proceeds without any further input until completion. This method is particularly suitable for the production of probiotics and vitamins, where strict control of microbial growth and nutrient depletion is essential. Lactic acid bacteria (LAB), commonly used in batch fermentation, play a key role in probiotic production [150].
To overcome the limitations of batch fermentation, fed-batch fermentation allows for the continuous addition of nutrients during the fermentation process [150]. This ensures that microorganisms continue to grow and produce metabolites at optimal rates without being inhibited by nutrient depletion or toxic byproducts [151]. Fed-batch fermentation is commonly used to produce carotenoids such as β-carotene from Y. lipolytica [152], as well as from S. cerevisiae [153]. The production rate and total yield of β-carotene are significantly different when using bioreactors compared to flasks. The S. cerevisiae SM14 strain, developed through adaptive evolution, produced up to 21 mg/g dry cell weight (DCW) of β-carotene in shake flask cultures, while the βcar1.2 strain, generated by overexpressing carotenogenic genes, produced only 5.8 mg/g DCW. In fed-batch bioreactors, however, βcar1.2 outperformed SM14, achieving higher biomass and β-carotene productivity rates of 1.57 g/L/h and 10.9 mg/L/h, respectively, compared to SM14’s 0.48 g/L/h and 3.1 mg/L/h [154]. By continuous glucose feeding, the yeast cells maintain optimal metabolic activity, leading to 10.5% higher β-carotene production [153]. Continuous fermentation systems allow for the steady addition of nutrients and the removal of products and byproducts in real time, creating an uninterrupted production cycle. Microorganisms are kept in their exponential growth phase, where they are most productive, and products are harvested continuously. Continuous fermentation has been successfully employed in the production of Bifidobacterium species, a probiotic used in supplements to improve gut health. The system allows for the constant production of probiotic cultures, ensuring consistent quality and quantity of the product [155].
Solid-state fermentation involves growing microorganisms on solid substrates rather than in liquid media. This method is gaining attention for producing nutraceuticals with unique properties. Aspergillus oryzae is used in solid-state fermentation to produce antioxidants from agricultural byproducts, such as rice bran, for use in nutraceuticals [156]. Co-cultivation involves growing multiple microbial strains in the same bioreactor to exploit their synergistic interactions. Co-cultivation of Lactobacillus and Bifidobacterium species in the same bioreactor has been used to enhance the production of multi-strain probiotic supplements. These probiotics work together to improve the balance of gut microflora [157]. Membrane bioreactors (MBRs) use semi-permeable membranes to separate biomass from the fermentation broth, allowing for continuous filtration and product extraction. Membrane bioreactors are employed in the production of amino acids such as L-lysine by C. glutamicum. The MBR system allows for continuous removal of L-lysine, enhancing overall yield and simplifying downstream processing [141].
16.6. Process Optimization
The efficiency and yield of microbial fermentation for nutraceutical production can be significantly improved through process optimization. Critical parameters such as pH, temperature, oxygen levels, and nutrient supply must be precisely controlled to ensure the optimal growth of microorganisms and the production of desired metabolites [158]. Modern fermentation processes now integrate advanced monitoring and control systems, such as Process Analytical Technology (PAT), which provides real-time monitoring of key fermentation parameters, allowing for dynamic adjustments to optimize the fermentation process [159]. In the production of resveratrol by engineered E. coli, PAT tools are used to monitor the concentration of glucose, oxygen levels, and biomass during fermentation. These real-time data allow for fine-tuning of the process to maximize carotenoid yield [159]. In the production of omega-3 fatty acids from Schizochytrium species, a marine microalga, maintaining precise control over oxygen levels and pH, is critical to optimizing lipid accumulation, which is essential for high yields of omega-3 fatty acids. Automated bioreactors are employed to continuously monitor and adjust these parameters, ensuring that the microalga remains in its optimal growth phase throughout the fermentation process [160].
Higher yields and quality, along with lower costs and waste generation, will result from being able to depend on automated smart systems that need minimal human/manual intervention, which is crucial for bio-based biopharma products, particularly nutraceuticals. Gathering extensive sets of pertinent data is the greatest barrier to the adoption of smart manufacturing in bio-based industries [161,162]. According to studies, creating mathematical models has many advantages, particularly for the control and optimization of bioprocesses, to ensure operational reproducibility, quality control, and consistency [162]. As a result, sensors are needed to monitor physical factors like temperature and pressure, chemical quantities like pH and dissolved oxygen, and biological characteristics like cell density or metabolite concentrations. As shown in Figure 6, monitoring techniques and the sensors and analyzers that go along with them can be further categorized based on where they are in relation to the process unit. An in-line sensor constantly generates data (no sampling), and it is either in direct contact with the process medium (invasive) or is separated from it by a glass window (also known as an on-line sensor, non-invasive). These sensors enable continuous process control by providing continuous information. Samples close to the bioreactor are analyzed by at-line sensors. Even though the samples are collected regularly (manually or automatically), the analysis-related time delays (depending on the equipment) make such data ideal for monitoring but not for control. Finally, off-line measurement samples are manually or automatically collected before being sent to the lab for analysis. Due to the lengthy delays that result, these measurements are unable to control the dynamic process behavior [163].
Elazzazy, A.M.; Baeshen, M.N.; Alasmi, K.M.; Alqurashi, S.I.; Desouky, S.E.; Khattab, S.M.R. Where Biology Meets Engineering: Scaling Up Microbial Nutraceuticals to Bridge Nutrition, Therapeutics, and Global Impact. Microorganisms 2025, 13, 566.
https://doi.org/10.3390/microorganisms13030566
