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Robust systematic approaches for the metabolic engineering of cell factories remain elusive. The available models for predicting phenotypical responses and mechanisms are incomplete, particularly within the context of compound toxicity that can be a significant impediment to achieving high yields of a target product. This study describes a Multi-Omic Based Production Strain Improvement (MOBpsi) strategy that is distinguished by integrated time-resolved systems analyses of fed-batch fermentations. As a case study, MOBpsi was applied to improve the performance of an Escherichia coli cell factory producing the commodity chemical styrene. Styrene can be bio-manufactured from phenylalanine via an engineered pathway comprised of the enzymes phenylalanine ammonia lyase and ferulic acid decarboxylase. The toxicity, hydrophobicity, and volatility of styrene combine to make bio-production challenging. Previous attempts to create styrene tolerant E. coli strains by targeted genetic interventions have met with modest success. Application of MOBpsi identified new potential targets for improving performance, resulting in two host strains (E. coli NST74ΔaaeA and NST74ΔaaeA cpxPo) with increased styrene production. The best performing re-engineered chassis, NST74ΔaaeA cpxPo, produced ∼3 × more styrene and exhibited increased viability in fed-batch fermentations. Thus, this case study demonstrates the utility of MOBpsi as a systematic tool for improving the bio-manufacturing of toxic chemicals.

1,3-butadiene (butadiene) is a by-product produced during naphtha steam cracking, predominantly used in tyre manufacturing. Recently, steam crackers have converted to using more cost effective, lighter feedstocks such as shale gas, yielding less butadiene. The potential shortfall, coupled with concerns around increasing greenhouse gas emissions, provides a unique opportunity for renewable production. This study investigated the techno-economics and greenhouse gas emissions associated with renewable butadiene production routes within the context of a China located pulp mill. A hybrid bio-catalytic route, utilising black liquor, was compared against two chemo-catalytic routes using forestry residues and pulpwood. The hybrid bio-catalytic route uses a novel aerobic gas fermentation platform, employing heat integrated supercritical water gasification and aerobic gas fermentation to produce acetaldehyde, followed by chemo-catalytic upgrading (Acet-BD). The two chemo-catalytic routes catalytically upgrade biomass derived syngas; where one route (Eth-BD) passes through an ethanol intermediate, and the other (Syn-BD) utilises a series of commercialised catalytic technologies with propene as an intermediate. The hybrid bio/chemo-catalytic route, Acet-BD, was the only route profitable using the nominal techno-economic inputs, producing a Net Present Value of $2.8 million and Minimum Selling Price of $1367 tn−1. In contrast, the two chemo-catalytic routes produced Minimum Selling Prices of $1954 tn−1 (Eth-BD) and $2196 tn−1 (Syn-BD), demonstrating the competitiveness of this novel platform. Sensitivity analyses highlighted the equipment capital as the main contributor to increased Minimum Selling Price for all cases, and the Acet-BD route presented a 19% probability of achieving a positive net present value. Moreover, owed to the low process emissions and sequestration of biogenic carbon, all routes produced net negative emissions within a cradle-to-gate framework. As such, renewable butadiene production has potential as a net carbon sink for pulp mill residues conventionally destined for energy recovery.

Electrochemical oscillatory baffled reactors fabricated with additive manufacturing for efficient continuous-flow oxidations

Ethylene is a small hydrocarbon gas widely used in the chemical industry. Annual worldwide production currently exceeds 150 million tons, producing considerable amounts of CO2 contributing to climate change. The need for a sustainable alternative is therefore imperative. Ethylene is natively produced by several different microorganisms, including Pseudomonas syringae pv. phaseolicola via a process catalyzed by the ethylene forming enzyme (EFE), subsequent heterologous expression of EFE has led to ethylene production in non-native bacterial hosts including E. coli and cyanobacteria. However, solubility of EFE and substrate availability remain rate limiting steps in biological ethylene production. We employed a combination of genome scale metabolic modelling, continuous fermentation, and protein evolution to enable the accelerated development of a high efficiency ethylene producing E. coli strain, yielding a 49-fold increase in production, the most significant improvement reported to date. Furthermore, we have clearly demonstrated that this increased yield resulted from metabolic adaptations that were uniquely linked to the EFE enzyme (WT vs mutant). Our findings provide a novel solution to deregulate metabolic bottlenecks in key pathways, which can be readily applied to address other engineering challenges.

Supercritical water gasification (SCWG) is a combined thermal decomposition and hydrolysis process for converting wet biomass feedstock with high water content potentially (80 wt%) to syngas. The process bypasses the need for an energy intensive pre-drying step and also needs relatively shorter residence times (of the order of seconds to minutes) when compared to conventional gasification. The main target of SCWG is to obtain syngas rich in hydrogen whilst minimising char formation. In recent years, SCWG studies have advanced from using model compounds (e.g. glucose and cellulose) towards the use of real biomass and its waste (e.g. sugarcane trash). The use of biomass as a feedstock creates real opportunities for the technology since it is available in some form, regardless of location. This review discusses the findings from SCWG studies that have used real biomass as a feedstock. The effects of reaction temperature, pressure, residence time and feedstock concentration to the hydrogen yields are presented. The relationship between the main components in biomass (cellulose, hemicellulose and lignin) and hydrogen yields are also discussed. Homogeneous and heterogeneous catalysts have been used to enhance SCWG with real biomass feedstock and the benefits of these approaches are also considered. The economic benefits of running the catalytic SCWG at 400 °C compared to non-catalytic operation at 600 °C is evaluated. Reactor configuration and process conditions vary across the literature, and various authors describe the associated challenges (char formation and plugging, corrosion) as well as promising solutions to tackle these key challenges.

The manufacturing industry must diverge from a ‘take, make and waste’ linear production paradigm towards more circular economies. Truly sustainable, circular economies are intrinsically tied to renewable resource flows, where vast quantities need to be available at a central point of consumption. Abundant, renewable carbon feedstocks are often structurally complex and recalcitrant, requiring costly pretreatment to harness their potential fully. As such, the heat integration of supercritical water gasification (SCWG) and aerobic gas fermentation unlocks the promise of renewable feedstocks such as lignin. This study models the technoeconomics and life cycle assessment (LCA) for the sustainable production of the commodity chemicals, isopropanol and acetone, from gasified Kraft black liquor. The investment case is underpinned by rigorous process modelling informed by published continuous gas fermentation experimental data. Time series analyses support the price forecasts for the solvent products. Furthermore, a Monte Carlo simulation frames an uncertain boundary for the technoeconomic model. The technoeconomic assessment (TEA) demonstrates that production of commodity chemicals priced at ~US$1000 per tonne is within reach of aerobic gas fermentation. In addition, owing to the sequestration of biogenic carbon into the solvent products, negative greenhouse gas (GHG) emissions are achieved within a cradle-to-gate LCA framework. As such, the heat integrated aerobic gas fermentation platform has promise as a best-in-class technology for the production of a broad spectrum of renewable commodity chemicals.

Summary: The chemical industry must decarbonize to align with UN Sustainable Development Goals. A shift toward circular economies makes CO2 an attractive feedstock for producing chemicals, provided renewable H2 is available through technologies such as supercritical water (scH2O) gasification. Furthermore, high carbon and energy efficiency is paramount to favorable techno-economics, which poses a challenge to chemo-catalysis. This study demonstrates continuous gas fermentation of CO2 and H2 by the cell factory, Cupriavidus necator, to (R,R)-2,3-butanediol and isopropanol as case studies. Although a high carbon efficiency of 0.75 [(C-mol product)/(C-mol CO2)] is exemplified, the poor energy efficiency of biological CO2 fixation requires ∼8 [(mol H2)/(mol CO2)], which is techno-economically infeasible for producing commodity chemicals. Heat integration between exothermic gas fermentation and endothermic scH2O gasification overcomes this energy inefficiency. This study unlocks the promise of sustainable manufacturing using renewable feedstocks by combining the carbon efficiency of bio-catalysis with energy efficiency enforced through process engineering.

Globally, 2.01 billion tonnes of municipal solid waste (MSW) were generated in 2016, about 37% of which was disposed of into landfills. This study evaluates the environmental and financial viability of producing ethanol from autoclaved MSW via fermentation. Experimental screening of four different microorganisms (i.e., S. cerevisiae, Z. mobilis, E. coli, and S. pombe) and process modelling indicate that MSW-derived ethanol can significantly reduce greenhouse gas emissions relative to gasoline (84% reduction following EU Renewable Energy Directive accounting methodology, and by 156–231% reduction following the US Energy Independence and Security Act methodology). Utilisation of wastes for biofuel production in the UK benefits from policy support and financial support for renewable fuels (Renewable Transport Fuel Certificates). Financial analysis highlights that microorganisms achieving higher ethanol yield and productivity (S. cerevisiae and Z. mobilis) can achieve financial viability with higher cumulative net present value than E. coli, S. pombe. However, the positive net present value can be achieved primarily due to the benefit of gate fees received by diverting wastes to autoclave and ethanol production (64% of total revenues), rather than from revenues from ethanol sales (7% of total revenues). Key process improvements must be achieved to improve the financial viability of ethanol production from MSW and deliver a clear advantage over waste incineration, specifically improving hydrolysis yield, reducing enzyme loading rate and, to a lesser extent, increasing solid loading rate. The results provide significant insights into the role of policy and technology development to achieve viable waste-to-biofuel systems.

Living cells do not interface naturally with nanoscale materials, although such artificial organisms can have unprecedented multifunctional properties, like wireless activation of enzyme function using electromagnetic stimuli. Realizing such interfacing in a nanobiohybrid organism (or nanorg) requires (1) chemical coupling via affinity binding and self-assembly, (2) the energetic coupling between optoelectronic states of artificial materials with the cellular process, and (3) the design of appropriate interfaces ensuring biocompatibility. Here we show that seven different core-shell quantum dots (QDs), with excitations ranging from ultraviolet to near-infrared energies, couple with targeted enzyme sites in bacteria. When illuminated by light, these QDs drive the renewable production of different biofuels and chemicals using carbon-dioxide (CO2), water, and nitrogen (from air) as substrates. These QDs use their zinc-rich shell facets for affinity attachment to the proteins. Cysteine zwitterion ligands enable uptake through the cell, facilitating cell survival. Together, these nanorgs catalyze light-induced air-water-CO2 reduction with a high turnover number (TON) of ∼106-108 (mols of product per mol of cells) to biofuels like isopropanol (IPA), 2,3-butanediol (BDO), C11-C15 methyl ketones (MKs), and hydrogen (H2); and chemicals such as formic acid (FA), ammonia (NH3), ethylene (C2H4), and degradable bioplastics polyhydroxybutyrate (PHB). Therefore, these resting cells function as nanomicrobial factories powered by light.