Production Platform Research Articles

Microbial chassis as the platform for production of dihydroxy xanthophyll-based carotenoids: an overview of recent advances in biomanufacturing.

Physiological and environmental cues prompt microbes to synthesize diverse carotenoids, including dihydroxy xanthophylls, facilitating their adaptation and survival. Lutein and its isomeric counterpart, zeaxanthin, are notable dihydroxy xanthophylls with bioactive properties such as antioxidative, anti-inflammatory, anticancer, and neuroprotective effects, particularly beneficial for human ocular health. However, global natural resources for co-producing lutein and zeaxanthin are scarce, with zeaxanthin lacking commercial sources, unlike lutein sourced from marigold plants and microalgae. Traditionally, dihydroxy xanthophyll production primarily relies on petrochemical synthetic routes, with limited biological sourcing reported. Nonetheless, microbiological synthesis presents promising avenues as a commercial source, albeit challenged by low dihydroxy xanthophyll yield at high cell density. Strategies involving optimization of physical and chemical parameters are essential to achieve high-quality dihydroxy xanthophyll products. This overview briefly discusses dihydroxy xanthophyll biosynthesis and highlights recent advancements, discoveries, and industrial benefits of lutein and zeaxanthin production from microorganisms as alternative biofactories.

High‐Throughput Indirect Gravure Printing Applied to Low‐Temperature Metallization of Silicon‐Based High‐Efficiency Solar Cells

Herein, this work is dedicated to a both exotic and promising printing technique in silicon (Si) photovoltaics (PV). Indirect gravure printing is investigated as a metallization technique focusing on low‐temperature applications, such as Si heterojunction (SHJ) and perovskite Si tandem solar cells. One advantage of this rotary printing technique is that its components are very durable. From graphical industry, it is known that gravure cylinders show almost no wearing during multiple printing. Additionally, rotary printing techniques offer great throughput. Therefore, indirect gravure printing can reduce costs significantly in PV production. In this work, short process cycle times of 0.9 s cell−1 for industrial wafers (edge length of 156.75 mm) are demonstrated. Using an appropriate production platform even enables cycle times of down to 0.5 s cell−1. Busbarless SHJ half cells with non‐optimized gravure‐printed front grids reach a mean photoconversion efficiency that is (1.7 ± 0.5) %abs lower compared to screen‐printed reference samples, however, cycle time is reduced. The metal contacts exhibit a mean shading width of (65 ± 16) μm and an average height of (5 ± 1) μm. Applied to SHJ solar cells’ rear, such metallization can replace screen‐printed contacts with 0.1%abs efficiency loss only, as device simulations reveal.

Enhancing astaxanthin accumulation through the expression of the plant-derived astaxanthin biosynthetic pathway in Dunaliella salina

Dunaliella salina, a microalga that thrives under high-saline conditions, is notable for its high β-carotene content and the absence of a polysaccharide cell wall. These unique characteristics render it a prime candidate as a cellular platform for astaxanthin production. In this study, our initial tests in an E. coli revealed that β-ring-4-dehydrogenase (CBFD) and 4-hydroxy-β-ring-4-dehydrogenase (HBFD) genes from Adonis aestivalis outperformed β-carotene hydroxylase (BCH) and β-carotene ketolase (BKT) from Haematococcus pluvialis counterparts by two-fold in terms of astaxanthin biosynthesis efficiency. Subsequently, we utilized electroporation to integrate either the BKT gene or the CBFD and HBFD genes into the genome of D. salina. In comparison to wild-type D. salina, strains transformed with BKT or CBFD and HBFD exhibited inhibited growth, underwent color changes to shades of red and yellow, and saw a nearly 50% decline in cell density. HPLC analysis confirmed astaxanthin synthesis in engineered D. salina strains, with CBFD + HBFD-D. salina yielding 134.88 ± 9.12 μg/g of dry cell weight (DCW), significantly higher than BKT-D. salina (83.58 ± 2.40 μg/g). This represents the largest amount of astaxanthin extracted from transgenic D. salina, as reported to date. These findings have significant implications, opening up new avenues for the development of specialized D. salina-based microcell factories for efficient astaxanthin production.

Diasporic citizen journalism: Exploring the discussion on the 2022 blank paper protests in the Chinese twitter community

Departing from the conventional nation-state framework, this paper explores citizen journalism from a translocal and transnational perspective. Focusing on Chinese diasporic communities on Twitter, the study presents an explorative inquiry into the phenomenon of diasporic citizen journalism. Through an empirical case study of the 2022 Blank Paper Protests (BPP) in China, the research reveals the pivotal role the Chinese Twitter-sphere played as a vital platform for cross-border news production and sense-making during a critical event. The study also illuminates the diversity, networked dynamics, and internal conflicts within the Chinese digital diaspora concerning the BPP. Theoretically, this paper underscores the imperative need to expand the horizons of citizen journalism research, suggesting diasporic citizen journalism as a new avenue for future exploration. Methodologically, the study showcases the potential of state-of-the-art computational methods, including Large Language Models (LLMs), in advancing journalism research. The methodology employed in this study exemplifies the integration of LLMs to enhance automated topic and sentiment analysis.

Electromagnetic drop-on-demand (DoD) technology as an innovative platform for amorphous solid dispersion production

Production of amorphous solid dispersions (ASDs) is an effective strategy to promote the solubility and bioavailability of poorly water soluble medicinal substances. In general, ASD is manufactured using a variety of classic and modern techniques, most of which rely on either melting or solvent evaporation. This proof-of-concept study is the first ever to introduce electromagnetic drop-on-demand (DoD) technique as an alternative solvent evaporation-based method for producing ASDs. Herein 3D printing of ASDs for three drug-polymer combinations (efavirenz-Eudragit L100-55, lumefantrine-hydroxypropyl methylcellulose acetate succinate, and favipiravir-polyacrylic acid) was investigated to ascertain the reliability of this technique. Polarized light microscopy, differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD), and Fourier Transform Infrared (FTIR) spectroscopy results supported the formation of ASDs for the three drugs by means of DoD 3D printing, which significantly increases the equilibrium solubility of efavirenz from 0.03 ± 0.04 µg/ml to 21.18 ± 4.20 µg/ml, and the equilibrium solubility of lumefantrine from 1.26 ± 1.60 µg/ml to 20.21 ± 6.91 µg/ml. Overall, the reported findings show how this new electromagnetic DoD technology can have a potential to become a cutting-edge 3D printing solvent-evaporation technique for on-demand and continuous manufacturing of ASDs for a variety of drugs.

Engineering the microenvironment of P450s to enhance the production of diterpenoids in Saccharomyces cerevisiae

Cytochrome P450 enzymes play a crucial role as catalysts in the biosynthesis of numerous plant natural products (PNPs). Enhancing the catalytic activity of P450s in host microorganisms is essential for the efficient production of PNPs through synthetic biology. In this study, we engineered Saccharomyces cerevisiae to optimize the microenvironment for boosting the activities of P450s, including coexpression with the redox partner genes, enhancing NADPH supply, expanding the endoplasmic reticulum (ER), strengthening heme biosynthesis, and regulating iron uptake. This created a platform for the efficient production 11,20-dihydroxyferruginol, a key intermediate of the bioactive compound tanshinones. The yield was enhanced by 42.1-fold through 24 effective genetic edits. The optimized strain produced up to 67.69 ± 1.33 mg/L 11,20-dihydroxyferruginol in shake flasks. Our work represents a promising advancement toward constructing yeast cell factories containing P450s and paves the way for microbial biosynthesis of tanshinones in the future.

Recombinant human fibroblast growth factor 7 obtained from stable Chinese hamster ovary cells enhances wound healing.

Although fibroblast growth factor 7 (FGF7) is known to promote wound healing, its mass production poses several challenges and very few studies have assessed the feasibility of producing FGF7 in cell lines such as Chinese hamster ovary (CHO) cells. Therefore, this study sought to produce recombinant FGF7 in large quantities and evaluate its wound healing effect. To this end, the FGF7 gene was transfected into CHO cells and FGF7 production was optimized. The wound healing efficacy of N-glycosylated FGF7 was evaluated in animals on days 7 and 14 post-treatment using collagen patches (CPs), FGF7-only, and CP with FGF7 (CP+FGF7), whereas an untreated group was used as the control. Wound healing was most effective in the CP+FGF7 group. Particularly, on day 7 post-exposure, the CP+FGF7 and FGF7-only groups exhibited the highest expression of hydroxyproline, fibroblast growth factor, vascular endothelial growth factor, and transforming growth factor. Epidermalization in H&E staining showed the same order of healing as hydroxyproline content. Additionally, the CP+FGF7 and FGF7-only group exhibited more notable blood vessel formation on days 7 and 14. In conclusion, the prepared FGF7 was effective in promoting wound healing and CHO cells can be a reliable platform for the mass production of FGF7.

Evaluation of an experimental kelp farm’s structural behavior using regression modelling and response amplitude operators derived from in situ measurements

An experimental kelp farming system for exposed ocean conditions was designed, deployed, planted with kelp and instrumented for evaluation of its dynamic response to ocean waves, tides, and currents. The farm featured a novel “lattice” mooring design and anchor lines and cultivation lines (horizontal lines used as kelp growth substrate) made of fiberglass rods. The farm was deployed at a site in Saco Bay, Maine with 13 m (MLLW) water depth. There the farm was exposed to waves with heights up to 5.9 m. Anchor line tension, tide and wave height time series were gathered and processed into response amplitude operators (RAOs) and least squared error linear regression models enabling recognition of meaningful patterns between the forcing factors and the mooring response. Mean mooring line tensions were shown to increase nonlinearly with tide. Anchor line tension response amplitudes were shown to exhibit high sensitivity to both low and high frequency wave forcing. Numerical free-release test simulations suggested natural frequencies in heave of 0.91 Hz, indicating that tension response sensitivities at high frequency could be the result of resonance. Low frequency tension response disproportionate to the low frequency wave forcing could be explained by wave forcing on kelp cultivation arrays modulated by wave group envelopes. Instances of high magnitude, potentially damaging peak tensions, deemed shock loads, were prevalent in most load cases. Anchor line tension dynamics including RAO and shock load magnitudes were shown to be sensitive to mooring stiffness (ratio of tension to resulting elongation) and, in some cases, significant wave amplitude. Patterns of anchor line response indicated that additional mooring elasticity or geometric compliance and use of floatation with less sensitivity to high frequency waves could help avoid the cause of and costly consequences of amplified high frequency loading and high amplitude shock loading. RAOs and regression model results also indicated a subdued response in frequencies associated with ocean swell waves, suggesting desirable performance in waves most dominant in extreme storm events. With the proposed improvements, the farm system design suggests merit as a robust and durable macroalgae biomass production platform.

Tailoring regulatory components for metabolic engineering in cyanobacteria.

The looming climate crisis has prompted an ever-growing interest in cyanobacteria due to their potential as sustainable production platforms for the synthesis of energy carriers and value-added chemicals from CO2 and sunlight. Nonetheless, cyanobacteria are yet to compete with heterotrophic systems in terms of space-time yields and consequently production costs. One major drawback leading to the low production performance observed in cyanobacteria is the limited ability to utilize the full capacity of the photosynthetic apparatus and its associated systems, i.e. CO2 fixation and the directly connected metabolism. In this review, novel insights into various levels of metabolic regulation of cyanobacteria are discussed, including the potential of targeting these regulatory mechanisms to create a chassis with a phenotype favorable for photoautotrophic production. Compared to conventional metabolic engineering approaches, minor perturbations of regulatory mechanisms can have wide-ranging effects.

Carbon dioxide valorization into resveratrol via lithoautotrophic fermentation using engineered Cupriavidus necator H16

BackgroundIndustrial biomanufacturing of value-added products using CO2 as a carbon source is considered more sustainable, cost-effective and resource-efficient than using common carbohydrate feedstocks. Cupriavidus necator H16 is a representative H2-oxidizing lithoautotrophic bacterium that can be utilized to valorize CO2 into valuable chemicals and has recently gained much attention as a promising platform host for versatile C1-based biomanufacturing. Since this microbial platform is genetically tractable and has a high-flux carbon storage pathway, it has been engineered to produce a variety of valuable compounds from renewable carbon sources. In this study, the bacterium was engineered to produce resveratrol autotrophically using an artificial phenylpropanoid pathway.ResultsThe heterologous genes involved in the resveratrol biosynthetic pathway—tyrosine ammonia lyase (TAL), 4-coumaroyl CoA ligase (4CL), and stilbene synthase (STS) —were implemented in C. necator H16. The overexpression of acetyl-CoA carboxylase (ACC), disruption of the PHB synthetic pathway, and an increase in the copy number of STS genes enhanced resveratrol production. In particular, the increased copies of VvSTS derived from Vitis vinifera resulted a 2-fold improvement in resveratrol synthesis from fructose. The final engineered CR-5 strain produced 1.9 mg/L of resveratrol from CO2 and tyrosine via lithoautotrophic fermentation.ConclusionsTo the best of our knowledge, this study is the first to describe the valorization of CO2 into polyphenolic compounds by engineering a phenylpropanoid pathway using the lithoautotrophic bacterium C. necator H16, demonstrating the potential of this strain a platform for sustainable chemical production.

A novel chemically defined medium for the biotechnological and biomedical exploitation of the cell factory Leishmania tarentolae

The development of media for cell culture is a major issue in the biopharmaceutical industry, for the production of therapeutics, immune-modulating molecules and protein antigens. Chemically defined media offer several advantages, as they are free of animal-derived components and guarantee high purity and a consistency in their composition. Microorganisms of the genus Leishmania represent a promising cellular platform for production of recombinant proteins, but their maintenance requires supplements of animal origin, such as hemin and fetal bovine serum. In the present study, three chemically defined media were assayed for culturing Leishmania tarentolae, using both a wild-type strain and a strain engineered to produce a viral antigen. Among the three media, Schneider's Drosophila Medium supplemented with Horseradish Peroxidase proved to be effective for the maintenance of L. tarentolae promastigotes, also allowing the heterologous protein production by the engineered strain. Finally, the engineered strain was maintained in culture up to the 12th week without antibiotic, revealing its capability to produce the recombinant protein in the absence of selective pressure.

A ReLU-based linearization approach for maximizing oil production in subsea platforms: An application to flow splitting

Oil and gas extraction from subsea reservoirs is a complex process that is highly reliant on accurate mathematical modeling for optimization. A key challenge is identifying control settings to maximize oil production while accounting for the nonlinear behavior and constraints of the underlying physical systems. This paper introduces a new method using Rectified Linear Units (ReLU) neural networks to model these complexities. We focus on a subsea system connecting multiple oil wells to a manifold with multiple headers, explicitly modeling flow splitting to optimize oil yield. Our approach employs mixed-integer reformulations of ReLU neural networks to approximate pressure-drop functions in risers, allowing efficient optimization of production platforms considering routing possibility and flow-splitting operations. Computational analysis employing a branch-and-cut approach and p-split partitioning (for the ReLU model) shows that our ReLU-based methodology converges faster than standard piecewise-linear (PWL) approaches while delivering comparably effective results. Notably, ReLU found solutions in scenarios where PWL failed to find an incumbent solution within the time limit. These results underscore a significant advancement in the optimization of offshore production platforms by using ReLU-based models.

The production of Spanish fiction series for SVOD: evolution, trends and strategies

The consolidation of digital television via subscription video on demand (SVOD) services has created an unprecedented increase in the consumption of serialised TV shows, which has had direct consequences on the content production sector. Faced with an increasingly demanding, hyperconnected and discerning audience and a supply of content never previously seen, television production and distribution strategies are involved in a process of change and adaptation. This paper seeks to analyse the production of Spanish television series created to be distributed on SVOD platforms. The aim is to know the role that production companies play in the creation of television series for SVOD so as to identify strategies and cooperative alliances between production companies and platforms and to detect possible patterns and trends in the operations of the audiovisual market. Using a mixed methodology that combines quantitative study with qualitative data analysis, the 88 Spanish TV series and miniseries that premiered on SVOD platforms from 2016 to 2021 are analysed, taking as a starting point the year in which these platforms began operating in Spain. The results confirm a trend towards cooperation between independent production companies and the platforms themselves, while no presence of in-house production is observed. Likewise, the atomisation of the role of production companies in the context of SVOD is shown, a phenomenon that already existed in generalist television production.

Lactate-mediated medium-chain fatty acid production from expired dairy and beverage waste

Fruits, vegetables, and dairy products are typically the primary sources of household food waste. Currently, anaerobic digestion is the most used bioprocess for the treatment of food waste with concomitant generation of biogas. However, to achieve a circular carbon economy, the organics in food waste should be converted to new chemicals with higher value than energy. Here we demonstrate the feasibility of medium-chain carboxylic acid (MCCA) production from expired dairy and beverage waste via a chain elongation platform mediated by lactate. In a two-stage fermentation process, the first stage with optimized operational conditions, including varying temperatures and organic loading rates, transformed expired dairy and beverage waste into lactate at a concentration higher than 900 mM C at 43 °C. This lactate was then used to produce >500 mM C caproate and >300 mM C butyrate via microbial chain elongation. Predominantly, lactate-producing microbes such as Lactobacillus and Lacticaseibacillus were regulated by temperature and could be highly enriched under mesophilic conditions in the first-stage reactor. In the second-stage chain elongation reactor, the dominating microbes were primarily from the genera Megasphaera and Caproiciproducens, shaped by varying feed and inoculum sources. Co-occurrence network analysis revealed positive correlations among species from the genera Caproiciproducens, Ruminococcus, and CAG-352, as well as Megasphaera, Bacteroides, and Solobacterium, indicating strong microbial interactions that enhance caproate production. These findings suggest that producing MCCAs from expired dairy and beverage waste via lactate-mediated chain elongation is a viable method for sustainable waste management and could serve as a chemical production platform in the context of building a circular bioeconomy.

Historical charcoal production as a factor in soil cover heterogeneity in a fluvioglacial landscape − A case study from northern Poland

Although in situ wood thermal processing, as a common practice in European forests in historical times, has been confirmed in many studies, the environmental implications of that process remain poorly explored. In this study, we focused on evaluating the impact of that process on forest vegetation and soil cover heterogeneity in a fluvioglacial landscape dominated by Brunic Arenosols. This study covered three 70–95-year-old stands of Scots pine in northern Poland. Three soil profiles, including from the relict charcoal hearth (RCH), from the surrounding ditch and a control, were examined from each stand, as well as associated vegetation. The studies showed that 19th century charcoal production platforms were overgrown with 20–60% fewer plant species than control stands. The effect on soils varied according to their characteristics. The presence of anthropogenic horizons consisting of a mixture of burnt soil material, ashes, charcoal particles, and other products of wood pyrolysis was the most distinguishing feature of RCH soils classified as Spolic Technosols (Arenic, Dystric, Pyric, Protospodic, Transportic). Typically, these horizons were characterised by lower bulk density, and higher porosity, as well as higher contents of total organic carbon, nitrogen, and exchange cations than the control soils. The differences between the RCH and control soils were highlighted by a comparison between the pools of the components in a soil column (pedon). Clear tendencies were not recorded for pH, the phosphorus content and stocks, the forms of iron and aluminium, the saturation of the soil sorption complex with bases or the trophic index. This may be due to the long-term neutralising effect of forest vegetation, dominated by Scots pine. Moreover, the results of our study may indicate the problem of toxicity of the RCH soils, which should be undertaken, considering the utilitarian importance of the issue.

A Coupled, Global/Local Finite Element Methodology to Evaluate the Fatigue Life of Flexible Risers Attached to Floating Platforms for Deepwater Offshore Oil Production

A Coupled, Global/Local Finite Element Methodology to Evaluate the Fatigue Life of Flexible Risers Attached to Floating Platforms for Deepwater Offshore Oil Production

The SARS-CoV-2 Spike Protein Receptor-Binding Domain Expressed in Rice Callus Features a Homogeneous Mix of Complex-Type Glycans.

The spike protein receptor-binding domain (RBD) of SARS-CoV-2 is required for the infection of human cells. It is the main target that elicits neutralizing antibodies and also a major component of diagnostic kits. The large demand for this protein has led to the use of plants as a production platform. However, it is necessary to determine the N-glycan structures of an RBD to investigate its efficacy and functionality as a vaccine candidate or diagnostic reagent. Here, we analyzed the N-glycan profile of the RBD produced in rice callus. Of the two potential N-glycan acceptor sites, we found that one was not utilized and the other contained a mixture of complex-type N-glycans. This differs from the heterogeneous mixture of N-glycans found when an RBD is expressed in other hosts, including Nicotiana benthamiana. By comparing the glycosylation profiles of different hosts, we can select platforms that produce RBDs with the most beneficial N-glycan structures for different applications.

Yeast Platforms for Production and Screening of Bioactive Derivatives of Rauwolscine.

Monoterpene indole alkaloids (MIAs) make up a highly bioactive class of metabolites produced by a range of tropical and subtropical plants. The corynanthe-type MIAs are a stereochemically complex subclass with therapeutic potential against a large number of indications including cancer, psychotic disorders, and erectile dysfunction. Here, we report yeast-based cell factories capable of de novo production of corynanthe-type MIAs rauwolscine, yohimbine, tetrahydroalstonine, and corynanthine. From this, we demonstrate regioselective biosynthesis of 4 fluorinated derivatives of these compounds and de novo biosynthesis of 7-chlororauwolscine by coexpression of a halogenase with the biosynthetic pathway. Finally, we capitalize on the ability of these cell factories to produce derivatives of these bioactive scaffolds to establish a proof-of-principle drug discovery pipeline in which the corynanthe-type MIAs are screened for bioactivity on human drug targets, expressed in yeast. In doing so, we identify antagonistic and agonistic behavior against the human adrenergic G protein-coupled receptors ADRA2A and ADRA2B, and the serotonergic receptor 5HT4b, respectively. This study thus demonstrates a proto-drug discovery pipeline for bioactive plant-inspired small molecules based on one-pot biocatalysis of natural and new-to-nature corynanthe-type MIAs in yeast.

Simultaneous product platform design and scheduling on unrelated additive/subtractive hybrid machines

Simultaneous product platform design and scheduling on unrelated additive/subtractive hybrid machines

Efficient production of l-glutathione by whole-cell catalysis with ATP regeneration from adenosine.

l-glutathione (GSH) is an important tripeptide compound with extensive applications in medicine, food additives, and cosmetics industries. In this work, an innovative whole-cell catalytic strategy was developed to enhance GSH production by combining metabolic engineering of GSH biosynthetic pathways with an adenosine-based adenosine triphosphate(ATP) regeneration system inEscherichia coli. Concretely, to enhance GSH production in E. coli, several genes associated with GSH and l-cysteine degradation, as well as the branched metabolic flow, were deleted. Additionally, the GSH bifunctional synthase (GshFSA) and GSH ATP-binding cassetteexporter (CydDC) were overexpressed. Moreover, an adenosine-based ATP regeneration system was first introduced into E. coli to enhance GSH biosynthesis without exogenous ATP additions. Through the optimization of whole-cell catalytic conditions, the engineered strain GSH17-FDC achieved an impressive GSH titer of 24.19 g/L only after 2 h reaction, with a nearly 100% (98.39%) conversion rate from the added l-Cys. This work not only unveils a new platform for GSH production but also provides valuable insights for the production of other high-value metabolites that rely on ATP consumption.