Biocatalysis: applications and potentials for the chemical industry

The chemical industry is exploring the use of renewable feed stocks to improve sustainability, prompting the exploration of bioprocesses for the production of chemicals. Attractive features of biological systems include versatility, substrate selectivity, regioselectivity, chemoselectivity, enantioselectivity and catalysis at ambient temperatures and pressures. However, a challenge facing bioprocesses is cost competitiveness with chemical processes because capital assets associated with the existing commercial processes are high. The chemical industry will probably use biotechnology with existing feed stocks and processes to extract higher values from feed stocks, process by-products and waste streams. In this decade, bioprocesses that offer either a process or a product advantage over traditional chemical routes will become more widely used.     

Toward a Carbon Dioxide Neutral Industrial Park

The industrial park of Herdersbrug (Brugge, Flanders, Belgium) comprises 92 small and medium‐sized enterprises, a waste‐to‐energy incinerator, and a power plant (not included in the study) on its site. To study the carbon dioxide (CO₂) neutrality of the park, we made a park‐wide inventory for 2007 of the CO₂ emissions due to energy consumption (electricity and fossil fuel) and waste incineration, as well as an inventory of the existing renewable electricity and heat generation. The definition of CO₂ neutrality in Flanders only considers CO₂ released as a consequence of consumption or generation of electricity, not the CO₂ emitted when fossil fuel is consumed for heat generation. To further decrease or avoid CO₂ emissions, we project and evaluate measures to increase renewable energy generation. The 21 kilotons (kt) of CO₂ emitted due to electricity consumption are more than compensated by the 25 kt of CO₂ avoided by generation of renewable electricity. Herdersbrug Industrial Park is thus CO₂ neutral, according to the definition of the Flemish government. Only a small fraction (6.6%) of the CO₂ emitted as a consequence of fossil fuel consumption (heat generation) and waste incineration is compensated by existing and projected measures for renewable heat generation. Of the total CO₂ emission (149 kt) due to energy consumption (electricity + heat generation) and waste incineration on the Herdersbrug Industrial Park in 2007, 70.5% is compensated by existing and projected renewable energy generated in the park. Forty‐seven percent of the yearly avoided CO₂ corresponds to renewable energy generated from waste incineration and biomass fermentation.     

Microalgal process-monitoring based on high-selectivity spectroscopy tools: status and future perspectives

Microalgae are well known for their ability to accumulate lipids intracellularly, which can be used for biofuels and mitigate CO₂ emissions. However, due to economic challenges, microalgae bioprocesses have maneuvered towards the simultaneous production of food, feed, fuel, and various high-value chemicals in a biorefinery concept. On-line and in-line monitoring of macromolecules such as lipids, proteins, carbohydrates, and high-value pigments will be more critical to maintain product quality and consistency for downstream processing in a biorefinery to maintain and valorize these markets. The main contribution of this review is to present current and prospective advances of on-line and in-line process analytical technology (PAT), with high-selectivity – the capability of monitoring several analytes simultaneously – in the interest of improving product quality, productivity, and process automation of a microalgal biorefinery. The high-selectivity PAT under consideration are mid-infrared (MIR), near-infrared (NIR), and Raman vibrational spectroscopies. The current review contains a critical assessment of these technologies in the context of recent advances in software and hardware in order to move microalgae production towards process automation through multivariate process control (MVPC) and software sensors trained on “big data”. The paper will also include a comprehensive overview of off-line implementations of vibrational spectroscopy in microalgal research as it pertains to spectral interpretation and process automation to aid and motivate development.     

Biological upgrading of pyrolysis-derived wastewater: Engineering Pseudomonas putida for alkylphenol,furfural, and acetone catabolism and (methyl)muconic acid production

While biomass-derived carbohydrates have been predominant substrates for biological production of renewable fuels, chemicals, and materials, organic waste streams are growing in prominence as potential alternative feedstocks to improve the sustainability of manufacturing processes. Catalytic fast pyrolysis (CFP) is a promising approach to generate biofuels from lignocellulosic biomass, but it generates a complex, carbon-rich, and toxic wastewater stream that is challenging to process catalytically but could be biologically upgraded to valuable co-products. In this work, we implemented modular, heterologous catabolic pathways in the Pseudomonas putida KT2440-derived EM42 strain along with the overexpression of native toxicity tolerance machinery to enable utilization of 89% (w/w) of carbon in CFP wastewater. The dmp monooxygenase and meta-cleavage pathway from Pseudomonas putida CF600 were constitutively expressed to enable utilization of phenol, cresols, 2- and 3-ethyl phenol, and methyl catechols, and the native chaperones clpB, groES, and groEL were overexpressed to improve toxicity tolerance to diverse aromatic substrates. Next, heterologous furfural and acetone utilization pathways were incorporated, and a native alcohol dehydrogenase was overexpressed to improve methanol utilization, generating reducing equivalents. All pathways (encoded by genes totaling ~30 kilobases of DNA) were combined into a single strain that can catabolize a mock CFP wastewater stream as a sole carbon source. Further engineering enabled conversion of all aromatic compounds in the mock wastewater stream to (methyl)muconates with a ~90% (mol/mol) yield. Biological upgrading of CFP wastewater as outlined in this work provides a roadmap for future applications in valorizing other heterogeneous waste streams.     

基于自然的碳达峰、碳中和解决方案：关键议题

实现碳达峰、碳中和是我国生态文明建设的重点战略方向。按照“目标设计-要素布局-方案实施”的基本逻辑，可以对碳达峰、碳中和的工作要点进行系统梳理。基于自然的解决方案(Nature-based Solutions, NbS)是国际社会普遍认可的生态恢复前沿理念，具备可持续性、系统性、尺度性、适应性四方面核心特质，能够有效回应“双碳”目标的工作要求。碳达峰、碳中和与NbS在目标导向和实践思路上高度契合，二者均以可持续发展为主要目标，NbS八项实施准则亦能够为“双碳”工作的积极推进提供全面系统的行动框架。结合NbS实施准则，具体提出基于自然的碳达峰、碳中和八大关键议题：(1)统筹减污降碳协同增效的政策体系；(2)强化尺度级联的传导机制；(3)构建基于自然的固碳途径；(4)完善多元化的碳汇交易；(5)兼顾社会公平的参与机制；(6)促进基于权衡的路径选择；(7)创新动态连续的监测手段；(8)推动NbS在“双碳”领域主流化，以期支撑国家碳达峰、碳中和可持续解决方案的落实。     

Sustainable conversion of coffee and other crop wastes to biofuels and bioproducts using coupled biochemical and thermochemical processes in a multi-stage biorefinery concept

The environmental impact of agricultural waste from the processing of food and feed crops is an increasing concern worldwide. Concerted efforts are underway to develop sustainable practices for the disposal of residues from the processing of such crops as coffee, sugarcane, or corn. Coffee is crucial to the economies of many countries because its cultivation, processing, trading, and marketing provide employment for millions of people. In coffee-producing countries, improved technology for treatment of the significant amounts of coffee waste is critical to prevent ecological damage. This mini-review discusses a multi-stage biorefinery concept with the potential to convert waste produced at crop processing operations, such as coffee pulping stations, to valuable biofuels and bioproducts using biochemical and thermochemical conversion technologies. The initial bioconversion stage uses a mutant Kluyveromyces marxianus yeast strain to produce bioethanol from sugars. The resulting sugar-depleted solids (mostly protein) can be used in a second stage by the oleaginous yeast Yarrowia lipolytica to produce bio-based ammonia for fertilizer and are further degraded by Y. lipolytica proteases to peptides and free amino acids for animal feed. The lignocellulosic fraction can be ground and treated to release sugars for fermentation in a third stage by a recombinant cellulosic Saccharomyces cerevisiae, which can also be engineered to express valuable peptide products. The residual protein and lignin solids can be jet cooked and passed to a fourth-stage fermenter where Rhodotorula glutinis converts methane into isoprenoid intermediates. The residues can be combined and transferred into pyrocracking and hydroformylation reactions to convert ammonia, protein, isoprenes, lignins, and oils into renewable gas. Any remaining waste can be thermoconverted to biochar as a humus soil enhancer. The integration of multiple technologies for treatment of coffee waste has the potential to contribute to economic and environmental sustainability.     

Biodiesel from microalgae

Continued use of petroleum sourced fuels is now widely recognized as unsustainable because of depleting supplies and the contribution of these fuels to the accumulation of carbon dioxide in the environment. Renewable, carbon neutral, transport fuels are necessary for environmental and economic sustainability. Biodiesel derived from oil crops is a potential renewable and carbon neutral alternative to petroleum fuels. Unfortunately, biodiesel from oil crops, waste cooking oil and animal fat cannot realistically satisfy even a small fraction of the existing demand for transport fuels. As demonstrated here, microalgae appear to be the only source of renewable biodiesel that is capable of meeting the global demand for transport fuels. Like plants, microalgae use sunlight to produce oils but they do so more efficiently than crop plants. Oil productivity of many microalgae greatly exceeds the oil productivity of the best producing oil crops. Approaches for making microalgal biodiesel economically competitive with petrodiesel are discussed.     

中国省域旅游业碳中和时空分异与模拟

旅游业碳中和的实现对于旅游业的绿色高质量发展和可持续发展至关重要。基于全国尺度以30个省份为分析单元,在承接团队前期关于旅游业碳达峰研究成果的基础上,借助土地利用数据、碳吸收系数和灰色预测模型分别测算与模拟了近20年和未来40年各省旅游业碳汇,通过旅游业碳中和指数反映旅游业碳排放和旅游业碳汇之间的动态变化,并利用空间自相关探索旅游业碳中和指数的时空差异。结果表明：（1）未来40年,我国省域旅游业碳汇总体呈现出"南北高,中间低"的空间分布特征,大多数省份的旅游业碳汇不断增长。东北部地区和长江流域以南各地区的旅游业碳汇较为富余,华东地区的山东、江苏和上海等省份的旅游业碳汇则相对匮乏。（2）不同情景中,低碳情景下的旅游业碳中和实现情况最好,有云南、四川和青海等7个省份在2060年之前实现了旅游业碳中和,而中等情景和基准情景下均仅有黑龙江和云南2个省份能够如期或提前实现。其中,西部和北部沿边省份的旅游业碳中和实现情况都要优于其它地区。（3）各省旅游业碳中和指数在未来40年的等级分区大致呈现出从Ⅰ级区逐步向Ⅴ级区转变的趋势,我国总体旅游业碳赤字率由2030年的96.67%逐渐降至2060年的76.67%。（4）未来40年,我国省域旅游业碳中和指数在空间上总体处于集聚态势,热点和冷点的空间分布特征较为明显,且演化趋势较为稳定。其中,热点区和次热点区主要分布在西北、西南、华南和东北地区,冷点区和次冷点区集中分布在华北、华中和华东地区。研究有效探讨了旅游业碳中和研究的理论与范式,并为中国旅游业碳中和的实现提供了一定的现实参考。     

Biorefinery: Toward an industrial metabolism

Fossil fuel reserves are running out, global warming is becoming a reality, waste recycling is becoming ever more costly and problematic, and unrelenting population growth will require more and more energy and consumer products. There is now an alternative to the 100% oil economy; it is a renewable resource based on agroresources by using the whole plant. Production and development of these new products are based on biorefinery concept. Each constituent of the plant can be extracted and functionalized in order to produce non-food and food fractions, intermediate agro-industrial products and synthons. Three major industrial domains can be concerned: molecules, materials and energy. Molecules can be used as solvent surfactants or chemical intermediates in substitution of petrol derivatives. Fibers can be valorized in materials like composites. Sugars and oils are currently used to produce biofuels like bioethanol or biodiesel, but second-generation biofuels will use lignocellulosic biomass as raw material. Lipids can be used to produce a large diversity of products like solvent, lubricants, pastes or surfactants. Industrial biorefinery will be linked to the creation of new processes based on the twelve principles of green chemistry (clean processes, atom economy, renewable feedstocks…). Biotechnology, especially white biotechnology, will take a major part into these new processes with biotransformations (enzymology, micro-organisms…) and fermentation. The substitution of oil products by biobased products will develop a new bioeconomy and new industrial processes respecting the sustainable development concept. Industrial biorefinery can be developed on the principle that any residues of one can then be exploited as raw material by others in an industrial metabolism.     

Irrigating poplar energy crops with landfill leachate negatively affects soil micro- and meso-fauna

Increased municipal solid waste generated worldwide combined with substantial demand for renewable energy has prompted testing and deployment of woody feedstock production systems that reuse and recycle wastewaters as irrigation and fertilization. Populus selections are ideal for such systems given their fast growth, extensive root systems, and high water usage rates. Maintaining ecological sustainability (i.e., the capacity for an ecosystem to maintain its function and retain its biodiversity over time) during tree establishment and development is an important component of plantation success, especially for belowground faunal populations. To determine the impact of solid waste leachate on soil micro- and meso-fauna, we compared soilfrom eight different Populus clones receiving municipal solid waste landfill leachate irrigation with clones receiving fertilized (N, P K) well water irrigation. Microfauna (i.e., nematodes) communities were more diverse in control soils. Mesofauna (i.e., insects) were associated with all clones; however, they were four times more abundant around trees found within the control plot than those that received leachate treatments. Nematode and insect abundance varied among Populus clones yet insect diversity was greater in the leachate-treated soils. Phytotechnologies must allow for soil faunal sustainability, as upsetting this balance could lead to great reductions in phytotechnology efficacy.     

Microbial lignin valorization through depolymerization to aromatics conversion

Lignin is the most abundant source of renewable aromatic biopolymers and its valorization presents significant value for biorefinery sustainability, which promotes the utilization of renewable resources. However, it is challenging to fully convert the structurally complex, heterogeneous, and recalcitrant lignin into high-value products. The in-depth research on the lignin degradation mechanism, microbial metabolic pathways, and rational design of new systems using synthetic biology have significantly accelerated the development of lignin valorization. This review summarizes the key enzymes involved in lignin depolymerization, the mechanisms of microbial lignin conversion, and the lignin valorization application with integrated systems and synthetic biology. Current challenges and future strategies to further study lignin biodegradation and the trends of lignin valorization are also discussed.     

Water-based woody biorefinery

The conversion of biomass into chemicals and energy is essential in order to sustain our present way of life. Fossil fuels are currently the predominant energy source, but fossil deposits are limited and not renewable. Biomass is a reliable potential source of materials, chemicals and energy that can be replenished to keep pace with our needs. A biorefinery is a concept for the collection of processes used to convert biomass into materials, chemicals and energy. The biorefinery is a “catch and release” method for using carbon that is beneficial to both the environment and the economy. In this study, we discuss three elements of a wood-based biorefinery, as proposed by the SUNY College of Environmental Science and Forestry (ESF): hot-water extraction, hydrolysis, and membrane separation/concentration. Hemicelluloses are the most easily separable main component of woody biomass and thus form the bulk of the extracts obtained by hot-water extraction of woody biomass. Hot-water extraction is an important step in the processes of woody biomass and product generation, replacing alternative costly pre-treatment methods. The hydrolysis of hemicelluloses produces 5-carbon sugars (mainly xylose), 6-carbon sugars (mainly glucose and mannose), and acetic acid. The use of nano-filtration membranes is an efficient technology that can be employed to fractionate hot-water extracts and wood hydrolysate. The residual solid mass after hot-water extraction has a higher energy content and contains fewer easily degradable components. This allows for more efficient subsequent processing to convert cellulose and lignin into conventional products.     

A sustainable woody biomass biorefinery

Woody biomass is renewable only if sustainable production is imposed. An optimum and sustainable biomass stand production rate is found to be one with the incremental growth rate at harvest equal to the average overall growth rate. Utilization of woody biomass leads to a sustainable economy. Woody biomass is comprised of at least four components: extractives, hemicellulose, lignin and cellulose. While extractives and hemicellulose are least resistant to chemical and thermal degradation, cellulose is most resistant to chemical, thermal, and biological attack. The difference or heterogeneity in reactivity leads to the recalcitrance of woody biomass at conversion. A selection of processes is presented together as a biorefinery based on incremental sequential deconstruction, fractionation/conversion of woody biomass to achieve efficient separation of major components. A preference is given to a biorefinery absent of pretreatment and detoxification process that produce waste byproducts. While numerous biorefinery approaches are known, a focused review on the integrated studies of water-based biorefinery processes is presented. Hot-water extraction is the first process step to extract value from woody biomass while improving the quality of the remaining solid material. This first step removes extractives and hemicellulose fractions from woody biomass. While extractives and hemicellulose are largely removed in the extraction liquor, cellulose and lignin largely remain in the residual woody structure. Xylo-oligomers, aromatics and acetic acid in the hardwood extract are the major components having the greatest potential value for development. Higher temperature and longer residence time lead to higher mass removal. While high temperature (>200°C) can lead to nearly total dissolution, the amount of sugars present in the extraction liquor decreases rapidly with temperature. Dilute acid hydrolysis of concentrated wood extracts renders the wood extract with monomeric sugars. At higher acid concentration and higher temperature the hydrolysis produced more xylose monomers in a comparatively shorter period of reaction time. Xylose is the most abundant monomeric sugar in the hydrolysate. The other comparatively small amounts of monomeric sugars include arabinose, glucose, rhamnose, mannose and galactose. Acetic acid, formic acid, furfural, HMF and other byproducts are inevitably generated during the acid hydrolysis process. Short reaction time is preferred for the hydrolysis of hot-water wood extracts. Acid hydrolysis presents a perfect opportunity for the removal or separation of aromatic materials from the wood extract/hydrolysate. The hot-water wood extract hydrolysate, after solid-removal, can be purified by Nano-membrane filtration to yield a fermentable sugar stream. Fermentation products such as ethanol can be produced from the sugar stream without a detoxification step.     

Unlocking the biosynthesis of sesquiterpenoids from methane via the methylerythritol phosphate pathway in methanotrophic bacteria,using α-humulene as a model compound

Isoprenoids are an abundant and diverse class of natural products with various applications in the pharmaceutical, cosmetics and biofuel industries. A methanotroph-based biorefinery is an attractive scenario for the production of a variety of value-added compounds from methane, because methane is a promising alternative feedstock for industrial biomanufacturing. In this study, we metabolically engineered Methylotuvimicrobium alcaliphilum 20Z for de novo synthesis of a sesquiterpenoid from methane, using α-humulene as a model compound, via optimization of the native methylerythritol phosphate (MEP) pathway. Expression of codon-optimized α-humulene synthase from Zingiber zerumbet in M. alcaliphilum 20Z resulted in an initial yield of 0.04 mg/g dry cell weight. Overexpressing key enzymes (IspA, IspG, and Dxs) for debottlenecking of the MEP pathway increased α-humulene production 5.2-fold compared with the initial strain. Subsequently, redirecting the carbon flux through the Embden–Meyerhof–Parnas pathway resulted in an additional 3-fold increase in α-humulene production. Additionally, a genome-scale model using flux scanning based on enforced objective flux method was used to identify potential overexpression targets to increase flux towards isoprenoid production. Several target reactions from cofactor synthesis pathways were probed and evaluated for their effects on α-humulene synthesis, resulting in α-humulene yield up to 0.75 mg/g DCW with 18.8-fold enhancement from initial yield. This study first demonstrates production of a sesquiterpenoid from methane using methanotrophs as the biocatalyst and proposes potential strategies to enhance production of sesquiterpenoid and related isoprenoid products in engineered methanotrophic bacteria.     

生物炼制技术的发展

随着石油资源的日益枯竭,化石经济面临严峻的挑战,而以町再生生物质资源为基础的生物炼制正展现出其广阔的前景。通过石油炼制与生物炼制的对比,分析了生物炼制任各个方面所具有的优势。并以生物乙烯为例,阐述了其发展现状、存在问题及解决途径、未来发展趋势等。     

生物炼制技术的发展

随着石油资源的日益枯竭,化石经济面临严峻的挑战,而以町再生生物质资源为基础的生物炼制正展现出其广阔的前景。通过石油炼制与生物炼制的对比,分析了生物炼制任各个方面所具有的优势。并以生物乙烯为例,阐述了其发展现状、存在问题及解决途径、未来发展趋势等。[编者按]     

Homogeneous,heterogeneous and enzymatic catalysis for transesterification of high free fatty acid oil (waste cooking oil) to biodiesel: A review

In the last few years, biodiesel has emerged as one of the most potential renewable energy to replace current petrol-derived diesel. It is a renewable, biodegradable and non-toxic fuel which can be easily produced through transesterification reaction. However, current commercial usage of refined vegetable oils for biodiesel production is impractical and uneconomical due to high feedstock cost and priority as food resources. Low-grade oil, typically waste cooking oil can be a better alternative; however, the high free fatty acids (FFA) content in waste cooking oil has become the main drawback for this potential feedstock. Therefore, this review paper is aimed to give an overview on the current status of biodiesel production and the potential of waste cooking oil as an alternative feedstock. Advantages and limitations of using homogeneous, heterogeneous and enzymatic transesterification on oil with high FFA (mostly waste cooking oil) are discussed in detail. It was found that using heterogeneous acid catalyst and enzyme are the best option to produce biodiesel from oil with high FFA as compared to the current commercial homogeneous base-catalyzed process. However, these heterogeneous acid and enzyme catalyze system still suffers from serious mass transfer limitation problems and therefore are not favorable for industrial application. Nevertheless, towards the end of this review paper, a few latest technological developments that have the potential to overcome the mass transfer limitation problem such as oscillatory flow reactor (OFR), ultrasonication, microwave reactor and co-solvent are reviewed. With proper research focus and development, waste cooking oil can indeed become the next ideal feedstock for biodiesel.     

Estimating environmental impacts of early-stage bioprocesses

The use of bioprocesses in industrial production promises resource- and energy-efficient processes starting from renewable, nonfossil feedstocks. Thus, the environmental benefits must be demonstrated, ideally in the early development phase with standardized methods such as life cycle assessment (LCA). Herein we discuss selected LCA studies of early-stage bioprocesses, highlighting their potential and contribution to estimating environmental impacts and decision support in bioprocess development. However, LCAs are rarely performed among bioprocess engineers due to challenges such as data availability and process uncertainties. To address this issue, recommendations are provided for conducting LCAs of early-stage bioprocesses. Opportunities are identified to facilitate future applicability, for example, by establishing dedicated bioprocess databases that could enable the use of LCAs as standard tools for bioprocess engineers.     

Phenotype analysis of cultivation processes via unsupervised machine learning: Demonstration for Clostridium pasteurianum

A novel approach of phenotype analysis of fermentation‐based bioprocesses based on unsupervised learning (clustering) is presented. As a prior identification of phenotypes and conditional interrelations is desired to control fermentation performance, an automated learning method to output reference phenotypes (defined as vector of biomass‐specific rates) was developed and the necessary computing process and parameters were assessed. For its demonstration, time series data of 90 Clostridium pasteurianum cultivations were used which feature a broad spectrum of solventogenic and acidogenic phenotypes, while 14 clusters of phenotypic manifestations were identified. The analysis of reference phenotypes showed distinct differences, where potential conditionalities were exemplary isolated. Further, cluster‐based balancing of carbon and ATP or the use of reference phenotypes as indicator for bioprocess monitoring were demonstrated to highlight the perks of this approach. Overall, such analysis depends strongly on the quality of the data and experimental validations will be required before conclusions. However, the automated, streamlined and abstracted approach diminishes the need of individual evaluation of all noisy dataset and showed promising results, which could be transferred to strains with comparably wide‐ranging phenotypic manifestations or as indicators for repeated bioprocesses with clearly defined target.     

Biochemistry,genetics and biotechnology of glycerol utilization in Pseudomonas species

The use of renewable waste feedstocks is an environment-friendly choice contributing to the reduction of waste treatment costs and increasing the economic value of industrial by-products. Glycerol (1,2,3-propanetriol), a simple polyol compound widely distributed in biological systems, constitutes a prime example of a relatively cheap and readily available substrate to be used in bioprocesses. Extensively exploited as an ingredient in the food and pharmaceutical industries, glycerol is also the main by-product of biodiesel production, which has resulted in a progressive drop in substrate price over the years. Consequently, glycerol has become an attractive substrate in biotechnology, and several chemical commodities currently produced from petroleum have been shown to be obtained from this polyol using whole-cell biocatalysts with both wild-type and engineered bacterial strains. Pseudomonas species, endowed with a versatile and rich metabolism, have been adopted for the conversion of glycerol into value-added products (ranging from simple molecules to structurally complex biopolymers, e.g. polyhydroxyalkanoates), and a number of metabolic engineering strategies have been deployed to increase the number of applications of glycerol as a cost-effective substrate. The unique genetic and metabolic features of glycerol-grown Pseudomonas are presented in this review, along with relevant examples of bioprocesses based on this substrate – and the synthetic biology and metabolic engineering strategies implemented in bacteria of this genus aimed at glycerol valorization.