Biotechnology for Biofuels最新文献

The shift toward sustainable biomanufacturing necessitates microbial platforms that efficiently convert low-cost, non-food feedstocks into high-value chemicals. Sucrose, a widely available and economical carbon source, remains underutilized in industrial Escherichia coli fermentation due to its low metabolic efficiency. This study investigates the production of L-lactic acid monomer in E. coli using sucrose, a cost-effective carbon source. Initially, we found that the recombinant strain 090S with the cscR gene knocked out exhibited an enhanced aerobic growth rate; however, during anaerobic fermentation for acid production, synthesis of the lactic acid monomer ceased after 3–4 h, indicating an impediment in sucrose metabolism under anaerobic conditions. Furthermore, we analyzed its transcriptional characteristics under aerobic-anaerobic phases through dynamic transcriptomic profiling and found significant differences. Specifically, for the csc operon, all three genes (cscB, cscK, and cscA) saw a significant decrease in expression when transferred into anaerobic conditions, retaining less than 10% of their aerobic expression levels. Here, we address this critical challenge by engineering optimized anaerobically active promoters to decouple sucrose utilization from native transcriptional constraints. Ultimately, the recombinant strain 091S, in which overexpresses the cscA and cscB genes by using the gapA promoter, produced 129.7 g/L of L-lactic acid in a 5-L bioreactor within 30 h of fermentation, with an average volumetric productivity of 4.32 g/(L·h), marking a 3.04-fold increase over the control. Additionally, an industrial fermentation process was simulated in a 30-L bioreactor under scaled-up conditions, resulting in a higher L-lactic acid yield of 145.7 g/L and a productivity of 4.96 g/(L·h), which was similar to that of glucose as a carbon source. This study elucidates the impact of oxygen content changes on gene transcription levels during the fermentation of E. coli using sucrose as a carbon source, offering a scalable and economically viable strategy for the efficient production of bio-products from sucrose or sucrose-rich feedstocks by E. coli.

Renowned for their distinctive aromas, terpenoid flavor compounds and their precursors are widely used in medicine, food, and the flavor and fragrance industries. Rapid advances in synthetic biology, including the modification of microbial chassis cells, the design of synthetic pathways for novel target products, and the integration of large-scale microbial fermentation, have enabled the development of microbial cell factories for the green and efficient production of terpenoid flavor compounds and their precursors, offering broader market potential. This review examines common biosynthetic mechanisms, recent progress in the field, and strategies for enhancing the biosynthetic efficiency of terpenoid flavor compounds and their precursors. This study aims to support the advancements of sustainable production technologies and promote industrial application within the flavor and fragrance sector.

The rising demand for natural products is accelerating research into sustainable methods for producing bio-based flavourings like ethyl butyrate. In this study, ethyl butyrate was successfully produced through the enzymatic esterification of butyric acid and ethanol, which were derived from the co-cultivation of Clostridium tyrobutyricum and Saccharomyces cerevisiae. Initial monoculture experiments with both strains were performed to investigate compromised fermentation conditions for co-cultivation. Based on these findings, anaerobic co-cultivation conditions were established at 37 °C and 150 rpm, with the pH controlled at 6. The effects of varying inoculation times in co-culture were examined, considering the solvent and acid tolerance of both strains. Due to the limited acid tolerance of S. cerevisiae, with significant inhibition at butyric acid concentrations above 10 g L¯¹, a time-delayed inoculation with C. tyrobutyricum was implemented. In batch experiments, the final concentrations of butyric acid and ethanol were 13.98 ± 3.06 g L¯¹ and 21.43 ± 1.66 g L¯¹, respectively. Further enhancement of product concentrations was explored through a fed-batch cultivation strategy yielding up to 45.62 ± 3.82 g L¯¹ of butyric acid and 18.61 ± 4.11 g L¯¹ of ethanol. Ethyl butyrate was formed from the fermentation products by lipase-catalysed enzymatic esterification in a two-phase system through the addition of an organic phase. The ester concentration in the organic phase reached a maximum of 23.93 ± 0.68 g L¯¹ (esterification yield 25%). This study presents a viable approach to the production of bio-based ethyl butyrate offering a sustainable alternative to traditional chemical synthesis methods.

Graphical Abstract

Waste activated sludge (WAS) represents a significant operational and environmental challenge for wastewater treatment plants (WWTPs) due to its low biodegradability, attributed to extracellular polymeric substances (EPS) that hinder enzymatic hydrolysis. Electrochemical (EC) pretreatment has shown promise in improving organic matter solubilization. However, conventional systems often face limitations related to high energy demand, mineralization of organic matter and electrode degradation. This study evaluates EC pretreatment with two dimensionally stable anodes, Ti/RuO₂ and Ti/RuO₂–ZrO₂–Sb₂O₅, as scalable alternatives for improving WAS biodegradability and energy recovery. The EC with both electrodes using WAS as the sole electrolyte, with an applied current of 10 mA/cm² for 30 min, achieved significantly enhanced solubilization with minimal mineralization. This effectively enhances the anaerobic biodegradability of WAS and increases methane recovery while maintaining low energy consumption and avoiding chemical additives. Methane yields increased to 168 and 342 N-L_CH4/kg_VS for WAS pretreated with Ti/RuO₂ and Ti/RuO₂–ZrO₂–Sb₂O₅, respectively, compared to 85 N-L_CH4/kg_VS for untreated sludge. Energy analysis revealed a net gain of 1.64 kW-h/kg_VS, outperforming other EC systems reported in the literature. In this sense, the implementation of this process could be integrated at an industrial scale in WWTPs as a cost-effective strategy for sludge valorization and resource recovery, in line with circular economy principles.

Gangliosides are essential glycosphingolipids critical in neurodevelopment and cell signaling. Traditionally sourced from animal tissues, their production raises ethical concerns and faces challenges in scalability and cost. Chemoenzymatic methods have emerged as alternatives but lack flexibility and broad industrial applicability of microbial systems. However, complete microbial biosynthesis remains challenging due to the complexity of reconstructing the biosynthetic pathway in non-native hosts. We report the first successful complete microbial synthesis of gangliosides by engineering the industrial filamentous fungus Ashbya gossypii. Using modular metabolic engineering, we heterologously expressed human and yeast enzymes to reconstruct a functional ganglioside biosynthetic pathway. Pathways for producing activated N-acetylneuraminic acid, lactosylceramide, and sialylated intermediates were integrated, yielding GM3 and GD3 at milligram-per-liter levels. These titers were further enhanced by introducing a heterologous Leloir pathway for galactose metabolism. This work represents a foundational advance in microbial glycoengineering, offering a scalable, animal-free microbial platform for ganglioside production with broad applications.

Background

Biological deconstruction of lignocellulose for sustainable chemical production offers an opportunity to harness evolutionarily specialized enzymes and organisms for industrial bioprocessing. While hydrolysis of cellulose and hemicellulose by CAZymes yields fermentable sugars, ligninolysis releases a heterogeneous mix of aromatic compounds that likely play a crucial role in shaping microbial communities and microbial metabolism. Here, we interrogated the metabolomic and transcriptomic response of a lignocellulolytic anaerobic fungus, Neocallimastix californiae, to a heterogeneous mixture of aromatic compounds derived from lignin.

Results

Through exposing the fungus to both a concentration it might experience in its native environment and an elevated concentration of alkaline lignin, we observe that N. californiae transforms vanillin and that supplying alkaline lignin at 0.125 g/L, alongside cellulose, enhances the growth and polysaccharide-degrading activity of N. californiae. Altogether, our results further suggest that vanillin consumption, increased polymer-degrading activity, increased metabolic activity, and transcriptomic remodeling of amino acid synthesis genes all coincide with increased melanin production by fungal cells. These observations challenge previous notions that aromatics from lignocellulose only inhibit the growth and polymer deconstruction capabilities of the biomass-degrading anaerobic fungi (Neocallimastigomycetes).

Conclusions

This study demonstrates that anaerobic fungi have a complex relationship with aromatic chemicals derived from lignin and hemicellulose and shift their metabolism in response to the addition of lignocellulose-derived aromatics to their growth medium. Further, as no known pathways for the biochemical transformation of aromatics were detected in these organisms despite observed transcriptome remodeling in the presence of aromatics, we suggest they might encode novel biochemical routes for scavenging amino acid building blocks from aromatic monomers derived from hemicellulose side chains and lignin.

Background

Oleaginous microorganisms are promising lipid producers that accumulate an abundance of lipids from different carbon sources. However, the cost of the carbon source in the culture medium is a significant component of the total substrate cost. In this study, lignocellulose from corncob hydrolysate (CBH) was used instead of glucose as a low-cost medium for Schizochytrium fermentation.

Results

Eicosapentaenoic acid (EPA) content was 7.31%, after 110 h of fermentation, when the total sugar concentration of CBH was 80 g/L, which was greater than that of pure glucose medium. Replacing 40% of freshwater with fermentation wastewater (FW) resulted in biomass, lipid titer, and EPA titer of 42.16 g/L, 23.05 g/L, and 1.72 g/L, respectively. Compared with the initial CBH medium, the lipid and EPA titers in the 7.5-L bioreactor employing the FW recycling strategy using CBH as a carbon source increased by 12.10% and 9.26%, respectively.

Conclusions

Corncob hydrolysate can be used as a potential low-cost and effective carbon source for EPA production by Schizochytrium sp. The recycling of FW provides a reference for reducing freshwater consumption and environmental pollution and realizing green and economic recycling fermentation.

This review critically examines the entire value chain of microalgal biorefineries, with the central aim of elucidating the key technological, economic, and environmental enablers and barriers that govern their transition from pilot-scale demonstrations to commercially viable, circular-economy applications. A systematic literature search was conducted across five major scientific databases using predefined Boolean strings: “algal biorefineries,” “microalgae biofuel,” “techno-economic analysis,” “life-cycle assessment,” and “bioproduct recovery.” Inclusion criteria encompassed peer-reviewed studies and authoritative policy documents published between January 2007 and March 2025 that provided empirical data on upstream cultivation, midstream processing, and downstream conversion, as well as techno-economic assessments (TEA) and life-cycle analyses (LCA). Exclusion criteria included non-English commentaries, purely theoretical models without experimental validation, and studies that focused exclusively on single-product streams. Unlike previous reviews that address isolated segments of the algal biorefinery pipeline, this work delivers a novel, integrative framework that synthesizes recent advances across cultivation modes, genetic and metabolic engineering, AI‐enabled optimization, and IoT‐driven monitoring. This review critically evaluates the trade-offs between CAPEX and OPEX, energy penalties associated with harvesting and drying, and inconsistencies in LCA to identify, where performance improvements yield the most significant economic and environmental returns. Finally, this review proposes a targeted research roadmap, spanning multivariate strain engineering, hybrid cultivation architectures, low‐energy harvesting technologies, cascade‐compatible fractionation platforms, standardized LCA/TEA protocols, and supportive policy mechanisms, that charts a clear path toward overcoming current bottlenecks. This comprehensive, evidence‐based synthesis aims to inform both academic research and industrial strategy, thereby advancing the field of sustainable algal biorefineries.

Population growth throughout the world has led to increased pollution and overconsumption of fossil resources. Microalgae are increasingly recognized as sustainable biofactories for producing lipids and astaxanthin, two commercially significant metabolites with wide-ranging applications in biofuel, pharmaceutical, cosmetic, and nutraceutical industries. Enhancing the yields of these compounds remains a major challenge due to growth–productivity trade-offs and limited understanding of regulatory mechanisms. This review aims to bridge that gap by providing a comprehensive and comparative analysis of traditional and modern strategies employed to enhance lipid and astaxanthin production in microalgae. We critically evaluate stress-based methods (e.g., salinity, light, nutrient limitation), phytohormone treatments, cultivation system optimization, and genome editing technologies, including CRISPR/Cas9. Special emphasis is given to gene-level responses and pathway-level regulation involved in these enhancements. This review article highlights the novel synchronization between astaxanthin and fatty acid biosynthesis under various stress conditions which emphasizes the role of diacylglycerol acyltransferase (DGAT) enzymes to enhance astaxanthin accumulation. Editing technologies with base suggest a novel strategy to reduce off-target effects and enhance metabolic efficiency related to lipid and astaxanthin biosynthesis.

Graphical Abstract

Enzymes capable of catalysing the production of hydrocarbons hold promise for sustainable fuel synthesis. However, the native activities of these enzymes are often insufficient for their exploitation in industrial bioprocesses. Enzyme engineering approaches including directed evolution (DE) can be used to improve the properties of enzymes to meet desirable standards for their industrial application. In this review, we summarise DE methods for engineering hydrocarbon-producing enzymes, including both screening- and selection procedures. The efficacy of DE depends on several factors, including sensitive and accurate detection of enzyme activity, the throughput of screening or selection steps, and the scale of diversity generation. Although DE is a well-established approach, its application in engineering hydrocarbon-producing enzymes has not been widely demonstrated. This can be attributed to the physiochemical properties of the target molecules, such as aliphatic hydrocarbons, which can be insoluble, gaseous, and chemically inert. Detection of these molecules in vivo presents several unique challenges, as does dynamically coupling their abundance to cell fitness. We conclude with a discussion on future directions and potential advancements in this field.