Search Results (203)

Background: Modern viral vector production needs to consider process intensification for higher yields from smaller production volumes. However, innate antiviral immunity triggered in the producer cell may limit virus replication. While commonly used cell lines (e.g., Vero or E1A-immortalised cells) are already compromised in antiviral pathways, the redundancy of innate signaling complicates host cell optimization by genetic engineering. Small molecules that are hypothesized to target antiviral pathways (Viral Sensitizers, VSEs) added to the culture media offer a versatile alternative to genetic modifications to increase permissiveness and, thus, viral yields across multiple cell lines. Methods: To explore how the yield for a chimeric Zika vaccine candidate (YF-ZIK) could be further be increased in an intensified bioprocess, we used spin tubes or an Ambr15 high-throughput microbioreactor system as scale-down models to optimize the dosing for eight VSEs in three host cell lines (AGE1.CR.pIX, BHK-21, and HEK293-F) based on their tolerability. Results: Addition of VSEs to an already optimized infection process significantly increased infectious titers by up to sevenfold for all three cell lines tested. The development of multi-component VSE formulations using a design of experiments approach allowed further synergistic titer increases in AGE1.CR.pIX cells. Scale-up to 1 L stirred-tank bioreactors and 3D-printed mimics of 200 or 2000 L reactors resulted in up to threefold and eightfold increases, respectively. Conclusions: Addition of single VSEs or combinations thereof allowed a further increase in YF-ZIK titers beyond the yield of an already optimized, highly intensified process. The described approach validates the use of VSEs and can be instructive for optimizing other virus production processes. Full article

Bioprocess simulation is a powerful tool for leveraging circular economy principles in the analysis of large-scale bioprocesses, enhancing decision-making for efficient and sustainable production. By simulating different process scenarios, researchers and engineers can evaluate the techno-economic feasibility of different approaches. This approach enables the identification of cost-effective and sustainable solutions, optimizing resource use and minimizing waste, thereby enhancing the overall efficiency and viability of bioprocesses within a circular economy framework. In this review, we provide an overview of circular economy concepts and trends before discussing design methodologies and challenges in large-scale bioprocesses. The analysis highlights the application and advantages of using process simulators like SuperPro Designer v.14 in bioprocess development. Process design methodologies have evolved to use specialized software that integrates chemical and biochemical processes, physical properties, and economic and environmental considerations. By embracing circular economy principles, these methodologies evaluate projects that transform waste into valuable products, aiming to reduce pollution and resources use, thereby shifting from a linear to a circular economy. In process engineering, exciting perspectives are emerging, particularly in large-scale bioprocess simulations, which are expected to contribute to the improvement of bioprocess technology and computer applications. Full article

Large-scale bioprocesses often suffer from spatial heterogeneities, which impact microbial performance and often lead to phenotypic population heterogeneity. To better understand these effects at the single-cell level, this study applied, for the first time, automated real-time flow cytometry (ART-FCM) to monitor L-phenylalanine production with an Escherichia coli triple reporter strain in a fed-batch process with glycerol as the carbon source. The strain was cultivated in both a well-mixed stirred-tank bioreactor (STR) and a scale-down two-compartment bioreactor (TCB), consisting of an STR and a coiled flow inverter (CFI) in bypass, to simulate spatial heterogeneities. ART-FCM enabled autonomous, high-frequency sampling every 20 min, allowing for real-time tracking of fluorescence signals linked to growth (rrnB-mEmerald), oxygen availability (narGHIJ-CyOFP1), and product formation (aroFBL-mCardinal2). The STR exhibited uniform reporter expression and higher biomass accumulation, while the TCB showed delayed product formation and pronounced phenotypic diversification depending on the set mean residence time in the CFI. Single-cell fluorescence distributions revealed that the shorter mean residence time in the CFI resulted in pronounced subpopulation formation, whereas longer exposure attenuated heterogeneity, indicating transcriptional adaptation. This finding highlights a critical aspect of scale-down studies: increased exposure duration to perturbations can enhance population robustness. Overall, this study demonstrates the relevance of ART-FCM, in combination with a multi-reporter strain, as a pioneering tool for capturing dynamic cellular behavior and correlating it to process performance, providing deeper insights into microbial heterogeneity under fluctuating bioprocess conditions. Full article

Clostridium novyi is a common pathogen in domestic animals and humans, and alpha-toxin is the main cause of its pathogenesis. Because it is a fastidious organism, obtaining alpha-toxin is expensive. Therefore, we proposed an in silico study to synthesize epitopes in cultures of Escherichia coli BL21 pLysS (DE3). First, we used a stirred-tank bioreactor, developing a dry mass yield (DMY) of 0.77 g/L in batch cultures and 1.03 g/L in fed-batch cultures, without acetic acid production. With scale-up using a system without mechanical agitation, there was a higher DMY (1.20 g/L) with 0.56 mmol/mL of alpha-toxin epitope 1 (DE3/Ep1) and 0.61 mmol/mL of alpha-toxin epitope 2 (DE3/Ep2), with a similar profile for O2 consumption, glucose, and no acetic acid production. The kinetic parameters µ(h⁻¹), YX/S, YP/S, QP, and QX did not differ significantly; however, the kinetic data were superior. Our results suggest that in silico tools allow epitope selection and bioprocess standardization. This system provides cost savings and technological advances for the veterinary vaccine industry. Full article

Fed-batch cultivation is a widely used strategy for microbial lipid production, offering flexibility in nutrient control and the potential for high lipid productivity. However, optimizing feeding strategies remains a complex challenge, as it depends on multiple factors, including strain-specific metabolism and process limitations. In this study, we developed a computational framework based on dynamic flux balance analysis and small-scale metabolic models to evaluate and optimize lipid production in Rhodosporidium toruloides strains. We proposed equations to estimate both the carbon and energy source mass feed rate ($F_{in}·s_r$) and its concentration in the feed ($s_r$) based on lipid accumulation targets, and defined minimum feeding flow rate ($F_{in}$) according to process duration. We then assessed the impact of these parameters on commonly used bioprocess metrics—lipid yield, titer, productivity, and intracellular accumulation—across wild-type and engineered strains. Our results showed that the selection of $F_{in}·s_r$ was strongly strain-dependent and significantly influenced strain performance. Moreover, for a given $F_{in}·s_r$, the specific values of $s_r$, and the resulting $F_{in}$, had distinct and non-equivalent effects on performance metrics. This methodology enables the rational pre-selection of feeding strategies and strains, improving resource efficiency and reducing the probability of failed experiments. Full article

Marine microbial-derived docosahexaenoic acid (DHA) has garnered significant attention as a sustainable and health-promoting alternative to fish oil-derived DHA. However, its industrial production from marine heterotrophic microorganisms faces challenges related to high costs and suboptimal oil quality, which hinder its broader application. This review focuses on recent strategies aimed at achieving low-cost and high-quality marine microbial DHA production, emphasizing heterotrophic systems that dominate commercial supply. Key aspects include: Fermentation optimization using waste-derived feedstocks and bioprocess engineering to enhance DHA yields; Critical refining techniques—including degumming, neutralization, decolorization, and deodorization—are analyzed for improving DHA oil purity and quality, with emphasis on process optimization to adapt to the unique biochemical properties of microbial-derived oils. Additionally, strategies for oxidative stabilization, such as antioxidant protection, are discussed to extend the shelf life and preserve the nutritional value of marine microbial DHA oil. By integrating techno-economic and biochemical perspectives, this work outlines a holistic framework to guide the industrial optimization of marine microbial-sourced DHA oil production, addressing cost and quality challenges to facilitate its large-scale application as functional foods and nutraceuticals, thereby reducing reliance on marine resources and advancing sustainable omega-3 production. Full article

Proteins are crucial for medicine, pharmaceuticals, food, and environmental applications since they are used in various fields such as synthesis of drugs, industrial enzyme production, biodegradable plastics, bioremediation processes, etc. Xylanase is an important and versatile enzyme with applications across various industries, including pulp and paper, biofuel production, food processing, textiles, laundry detergents, and animal feed. Key parameters in biotechnological protein production include temperature, pH, and working volumes and especially medium compositions where optimization is crucial for large-scale applications due to cost considerations. Machine learning methods have emerged as effective alternatives to traditional statistical approaches in optimization. This study focuses on optimizing xylanase production via bioprocesses by employing regression analysis on datasets from various studies. The extreme gradient boosting (XGBoost) regression model was applied to predict xylanase activity under different experimental conditions, accurately predicting xylanase activity and identifying the significance of each variable. This study utilized experimentally derived datasets from peer-reviewed publications, in which the corresponding laboratory experiments had already been conducted and validated. The results demonstrate that machine learning methods can effectively optimize protein production processes, offering a strong alternative to traditional statistical approaches. Full article

Large-scale cultivation of cyanobacteria is often limited by the high cost of synthetic culture medium and the environmental impact of nutrient consumption. Cheese whey, a major agro-industrial waste product, is rich in organic and inorganic nutrients, making it a promising low-cost alternative for microbial growth while addressing waste bioremediation. This study investigates the growth performance and the biochemical composition of four different cyanobacterial species (Phormidium sp., Synechocystis sp., Chlorogloeopsis fritschii, and Arthrospira platensis), cultivated in cheese whey (CW). Pretreated CW was used at 20% and 100% v/v concentrations. All species grew satisfactorily in both concentrations, reaching biomass above 4 g L⁻¹ (in 100% v/v CW) and 2 g L⁻¹ (in 20% v/v CW). The highest μ_max value (0.28 ± 0.02 d⁻¹) was presented by Synechocystis sp. grown in 20% CW. Waste bioremediation of both 20 and 100% v/v CW demonstrated effective nutrient removal, with COD removal exceeding 50% for most species, while total nitrogen (TN) and total phosphorus (TP) removals reached up to 33% and 32%, respectively. Biochemical composition analysis revealed high carbohydrate and protein content, while lipid content remained below 15% in all cases. Interestingly, C. fritschii accumulated 11% w/w polyhydroxyalkanoates (PHAs) during the last day of cultivation in 20% v/v CW. These findings highlight the potential of C. fritschii as a valuable candidate for integration into bioprocesses aimed at sustainable bioplastic production. Its ability to synthesize PHAs from agro-industrial waste not only enhances the economic viability of the process but also aligns with circular economy principles. This study is a primary step towards establishing a biorefinery concept for the cultivation of cyanobacterial species in cheese whey-based wastewater streams. Full article

In this review, we trace the evolution of liquid chromatography from the pioneering work of Tennikova and Svec to the current monolithic polymethacrylate supports for performing liquid chromatography with biological macromolecules and nanoparticles, which offer rapid, high-throughput separations. By using interconnected channels with a tailored channel diameter, monoliths minimize the diffusion limitations typical of particle-based systems. Radial flow designs and optimized channel architectures enable the direct loading of complex biological fluids, reducing the need for sample preparation and optimizing the purification of large biomolecules and nanoparticles such as proteins, nucleic acids, extracellular vesicles, and viruses. Recent work has integrated monoliths into immunoaffinity and enzyme reactor platforms, streamlining analytical workflows and preparative applications in vaccine production and gene therapy. The ongoing advances in monolithic materials, channel geometry, and continuous processing hold promise for even greater efficiency and scalability in future applications. Full article

Bone tissue engineering (BTE) emerged as a practical approach to tackle prosthetic industry limitations. We merge aspects from developmental biology, engineering and medicine with the aim to produce fully functional bone tissue. Mesenchymal stem cells have the capability of self-renewal and specific lineage differentiation. Herein lies their potential for BTE. Among MSCs, human dental pulp stem cells have a higher proliferation rate, shorter doubling times, lower cellular senescence, and enhanced osteogenesis than hBM-SCs under specific conditions. In addition, these cells are readily accessible and can be extracted through a subtle extraction procedure. Thus, they garner fewer moral concerns than most MSCs available and embody a promising cell source for BTE therapies able to replace hBM-MSCs. Interestingly, their study has been limited. Conversely, there is a need for their further study to harness their true value in BTE, with special emphasis in the design of bioprocesses able to produce viable, homogenous bone constructs in a clinical scale. Here, we study the osteogenic differentiation of hDPSCs encapsulated in alginate hydrogels under suspended culture in a novel perfusion bioreactor. The system is compared with traditional 3D static and fed-batch culture methodologies. The novel system performed better, producing higher alkaline phosphatase activity, and more homogeneous, dense and functional bone constructs. Additionally, cell constructs produced by the in-house-designed system were richer in mature osteoblast-like and mineralizing osteocyte-like cells. In conclusion, this study reports the development of a novel bioprocess able to produce hDPSC-based bone-like constructs, providing new insights into hDPSCs’ therapeutic potential and a system able to be transferred from the laboratory bench into medical facilities. Full article

Propionic acid (PA) is an important organic acid with applications in food preservation, feed additives, and bio-based chemical production. While industrial PA is mostly derived from petrochemical processes, sustainable microbial alternatives are gaining attention. In this study, we explored a co-fermentation strategy using lactic acid bacteria (LAB) with complementary metabolic capabilities to enhance PA biosynthesis via the 1,2-propanediol (PDO) pathway. Genome-based screening identified a metabolic division between strains capable of producing PDO (e.g., Carnobacterium maltaromaticum IBB3447) and those converting PDO to PA (e.g., Levilactobacillus brevis IBB3735). Notably, we discovered that C. maltaromaticum IBB3447 is capable of PDO 24 biosynthesis, a function previously undescribed in this species. Phenotypic assays confirmed glycerol metabolism and acid tolerance among strains. In co-culture fermentation trials, the highest PA concentration (6.87 mM) was achieved using simultaneous fermentation in a fructose–sorbitol–glucose (FRC-SOR-GLC) medium, accompanied by prior PDO accumulation (up to 13.13 mM). No single strain produced PA independently, confirming that metabolic cooperation is required. These findings reveal a novel LAB-based bioprocess for sustainable PA and PDO production, using cross-feeding interactions and the valorization of industrial waste streams. The study supports future optimization and scale-up for circular bioeconomy applications. Full article

Decades of technological advancements have led to major environmental concerns, particularly the bioaccumulation of heavy metals, which pose persistent risks to ecosystems and human health. Consequently, research has increasingly shifted from conventional remediation techniques toward more sustainable, environmentally friendly solutions. This review explores recent advancements, ongoing challenges, and future perspectives in the field of bioremediation, emphasizing its potential as a green technology for heavy metal decontamination. Despite significant progress, key challenges remain, including scalability limitations and the management of bioremediation by-products, along with the influence of regulatory policies and public perception on its large-scale implementation. Emerging approaches such as genetic engineering and nanotechnology show promise in overcoming these limitations. Gene editing allows the tailoring of specific metabolic traits for bioprocesses targeted towards increased tolerance to pollutants and higher biodegradation efficiency, higher enzymatic specificity and affinity, and improved yield and fitness in plants. Nanotechnologies, particularly biogenic nanostructures, open up the possibility of repurposing waste materials as well as harnessing the advantages of the biosynthesis of NPs with higher stability, biocompatibility, and biostimulant capacities. Furthermore, biopolymers and bio-based nanocomposites can improve the efficiency and costs of bioremediation protocols. Even so, further research is essential to evaluate their long-term risks and feasibility. Full article

The projected global population is expected to reach 9.7 billion by 2050, necessitating a significant increase in food production. Malnutrition remains a global health challenge that contributes to over 3.5 million deaths annually and accounts for 45% of all child mortalities. Microalgae, including cyanobacteria, are a promising solution because of their rich composition of bioactive compounds such as polyunsaturated fatty acids, carotenoids, proteins, vitamins, and minerals. These biomolecules provide various health benefits, including antioxidant, antidiabetic, anticancer, anti-inflammatory, and cardioprotective properties, making microalgal biomass a valuable ingredient in functional food formulations. However, the large-scale adoption of microalgae for food production faces several challenges, including species-specific variations in biochemical composition, inconsistencies in biomass yield, structural alterations during extraction and purification, sensory issues, and bioprocessing inefficiencies. Furthermore, regulatory challenges and concerns regarding bioavailability and safety continue to limit their widespread acceptance. Despite these limitations, microalgal bioactives have significant potential for the development of next-generation nutraceuticals and functional foods. This review examines the bioactive compounds found in microalgae, detailing their biological activities and functional applications in the food industry. Additionally, it explores the key challenges preventing their integration into food products and proposes strategies to overcome these challenges, ultimately facilitating the commercialization of microalgae as a sustainable and health-promoting food source. Full article

The projected global population of 9.22 billion by 2075 necessitates sustainable food sources that provide health benefits beyond essential nutrition, as the relationship between food biochemistry and human well-being is becoming increasingly significant. Microalgae are simple microscopic organisms rich in various bioactive compounds, such as pigments, vitamins, polyunsaturated fatty acids, polysaccharides, bioactive peptides, and polyphenols, which can be used to develop novel foods with potential health benefits. Bioactive substances offer numerous health benefits, including anti-inflammatory, anticancer, antioxidant, anti-obesity, and heart-protective effects. However, incorporating microalgal biomass into functional food products presents several challenges, including species diversity, fluctuations in biomass production, factors affecting cultivation, suboptimal bioprocessing methods, inconclusive evidence regarding bioavailability and safety, and undesirable flavors and aromas in food formulations. Despite these challenges, significant opportunities exist for the future development of microalgae-derived functional food products. Extensive investigations are essential to overcome these challenges and enable the large-scale commercialization of nutritious microalgae-based food products. This review aims to examine the potential of microalgae as natural ingredients in functional food production, explore the factors limiting their industrial acceptance and utilization, and assess the safety issues associated with human consumption. Full article

In this study, solid-state fermentation for growth and bacteriocin production by Lactiplantibacillus plantarum LD1 was carried out using wheat bran, a lignocellulosic substrate. This is the first report showing bacteriocin production using L. plantarum LD1 in solid-state fermentation. Wheat bran supported higher production of bacteriocin (391.69 ± 12.58 AU/mL) than other substrates. Appropriate conditions were achieved using statistical designs. Significant factors identified by Plackett–Burman Design and their interactions were studied using response surface methodology. Enhanced production of bacteriocin (582.86 ± 0.87 AU/mL) and optimal growth (log₁₀ CFU/mL 8.56 ± 0.42) were attained in wheat bran medium supplemented with peptone (1.13%), yeast extract (1.13%), glucose (1.56%), and tri-ammonium citrate (0.50%). Growth in non-optimized medium (MRS) was almost similar (log₁₀ CFU/mL 8.15 ± 0.20), but the bacteriocin production level was lower (391.69 ± 0.58 AU/mL). Bacteriocin production was sustainable using varied quantities of wheat bran, showing the suitability of the optimized bioprocess for large-scale production. The cost for bacteriocin production in the optimized medium was found to be 444,583.60 AU/USD, which is about 4 times more economical than the cost of the commercial MRS medium, 121,497.18 AU/USD). Thus, an almost 1.5-fold improvement in bacteriocin production was achieved using wheat bran as the substrate. The cost of the production medium was reduced by approximately 25%, making the bioprocess economical. Full article