Improving photosynthesis and metabolic networks for the competitive production of phototroph-derived biofuels

To improve bioenergy production from photosynthetic microorganisms it is necessary to optimize an extensive network of highly integrated biological processes. Systematic advances in pathway engineering and culture modification have resulted in strains with increased yields of biohydrogen, lipids, and carbohydrates, three bioenergy foci. However, additional improvements in photosynthetic efficiency are necessary to establish a viable system for biofuel production. Advances in optimizing light capture, energy transfer, and carbon fixation are essential, as the efficiencies of these processes are the principal determinants of productivity. However, owing to their regulatory, catalytic, and structural complexities, manipulating these pathways poses considerable challenges. This review covers novel developments in the optimization of photosynthesis, carbon fixation, and metabolic pathways for the synthesis of targeted bioenergy carriers.     

Online control of cell culture redox potential prevents antibody interchain disulfide bond reduction

The phenomenon of monoclonal antibody (mAb) interchain disulfide bond reduction during manufacturing processes continues to be a focus of the biotechnology industry due to the potential for loss of product, increased complexity of purification processes, and reduced stability of the drug product. We hypothesized that antibody reduction can be mitigated by controlling the cell culture redox potential and subsequently established a threshold redox potential above which the mAb remained intact and below which there were significant and highly variable amounts of reduced mAb. Using this knowledge, we developed three control schemes to prevent mAb reduction in the bioreactor by controlling the cell culture redox potential via an online redox probe. These control methodologies functioned by increasing the concentration of dissolved oxygen (DO), copper (II) (Cu), or both DO and Cu to maintain the redox potential above the threshold value. Using these methods, we were able to demonstrate successful control of antibody reduction. Importantly, the redox control strategies did not significantly impact the cell growth, viability, mAb production, or product quality attributes including aggregates, C-terminal lysine, high mannose, deamidation, and glycation. Our results demonstrate that controlling the cell culture redox potential is a simple and effective method to prevent mAb reduction.     

Recent advances in membrane technologies for biorefining and bioenergy production

The bioeconomy, and in particular, biorefining and bioenergy production, have received considerable attention in recent years as a shift to renewable bioresources to produce similar energy and chemicals derived from fossil energy sources, represents a more sustainable path. Membrane technologies have been shown to play a key role in process intensification and products recovery and purification in biorefining and bioenergy production processes. Among the various separation technologies used, membrane technologies provide excellent fractionation and separation capabilities, low chemical consumption, and reduced energy requirements. This article presents a state-of-the-art review on membrane technologies related to various processes of biorefining and bioenergy production, including: (i) separation and purification of individual molecules from biomass, (ii) removal of fermentation inhibitors, (iii) enzyme recovery from hydrolysis processes, (iv) membrane bioreactors for bioenergy and chemical production, such as bioethanol, biogas and acetic acid, (v) bioethanol dehydration, (vi) bio-oil and biodiesel production, and (vii) algae harvesting. The advantages and limitations of membrane technologies for these applications are discussed and new membrane-based integrated processes are proposed. Finally, challenges and opportunities of membrane technologies for biorefining and bioenergy production in the coming years are addressed.     

生物能源专刊序言

发展生物能源是减轻经济和社会发展对不可再生矿物质能源依赖程度,实现CO2减排的有效措施。本期专刊包括综述报告和研究论文两部分,涉及燃料乙醇、生物柴油、生物燃气、生物氢能、微生物燃料电池和微生物电解池等主要生物能源产品和系统,比较全面地分析其基础研究、关键技术开发和产业发展现状,讨论了存在的问题和挑战,展望了发展的前景。     

Perspective on opportunities in industrial biotechnology in renewable chemicals

From biomass to renewable chemicals: while industrial biotechnology offers a clear value proposition, a number of hurdles need to be addressed to fully realize the commercial potential of bio-based products and chemicals over the coming decade. A review of an early roadmap for biological production of chemicals from renewable sugars reveals a focus on those that would provide co-products for integrated biorefineries producing biofuels and bioenergy. A growing number of companies are now focusing on specialty chemicals as an entry point to build the bio-based economy.     

Modeling product formation in anaerobic mixed culture fermentations

The anaerobic conversion of organic matter to fermentation products is an important biotechnological process. The prediction of the fermentation products is until now a complicated issue for mixed cultures. A modeling approach is presented here as an effort to develop a methodology for modeling fermentative mixed culture systems. To illustrate this methodology, a steady-state metabolic model was developed for prediction of product formation in mixed culture fermentations as a function of the environmental conditions. The model predicts product formation from glucose as a function of the hydrogen partial pressure (P(H2)), reactor pH, and substrate concentration. The model treats the mixed culture as a single virtual microorganism catalyzing the most common fermentative pathways, producing ethanol, acetate, propionate, butyrate, lactate, hydrogen, carbon dioxide, and biomass. The product spectrum is obtained by maximizing the biomass growth yield which is limited by catabolic energy production. The optimization is constrained by mass balances and thermodynamics of the bioreactions involved. Energetic implications of concentration gradients across the cytoplasmic membrane are considered and transport processes are associated with metabolic energy exchange to model the pH effect. Preliminary results confirmed qualitatively the anticipated behavior of the system at variable pH and P(H2) values. A shift from acetate to butyrate as main product when either P(H2) increases and/or pH decreases is predicted as well as ethanol formation at lower pH values. Future work aims at extension of the model and structural validation with experimental data.     

Present and potential applications of cellulases in agriculture, biotechnology, and bioenergy

Cellulase (CEL) presently constitutes a major group of industrial enzyme based on its diverse ranges of utilization. Apart from such current and well-established applications—as in cotton processing, paper recycling, detergent formulation, juice extraction, and animal feed additives—their uses in agricultural biotechnology and bioenergy have been exploited. Supplementation of CELs to accelerate decomposition of plant residues in soil results in improved soil fertility. So far, applying CELs/antagonistic cellulolytic fungi to crops has shown to promote plant growth performance, including enhanced seed germination and protective effects. Their actions are believed mainly to trigger plant defense mechanisms and/or to act as biocontrol agents that mediate disease suppression. However, the exact interaction between the enzymes/fungi and plants has not been clearly elucidated. Under mild conditions, removal of plant cell wall polysaccharides by CELs for protoplast preparation results in reduced protoplast damage and increased viability and yields. CELs have recently shown great potential in enzyme aid extraction of bioactive compounds from plant materials before selective extraction through enhancing release of target molecules, especially those associated with the wall matrix. To date, attempts have been made to formulate CEL preparation for cellulosic-based bioethanol production. The high cost of CELs has created a bottleneck, resulting in an uneconomic production process. The utilization of low-cost carbohydrates, strain improvement, and gene manipulations has been alternatively aimed at reducing the cost of CEL production. In this review, we focus on and discuss current knowledge of CELs and their applications in agriculture, biotechnology, and bioenergy.     

Influence of the pH on (open) mixed culture fermentation of glucose: a chemostat study

Catabolic products from anaerobic fermentation processes are potentially of industrial interest. The volatile fatty acids and alcohols produced can be used as building blocks in chemical processes or applied directly as substrates in a mixed culture process to produce bioplastics. Development of such applications requires a predictable and controllable product spectrum of the fermentation process. The aim of the research described in this paper was (i) to investigate the product spectrum of an open mixed culture fermentation (MCF) process as a function of the pH, using glucose as substrate, and (ii) to relate the product spectrum obtained to generalized biochemical and thermodynamic considerations. A chemostat was operated under carbon and energy limitation in order to investigate the pH effect on the product spectrum in a MCF process. A transition from CO(2)/H(2) production at lower pH values to formate production at higher pH values was observed. The ratio of CO(2)/H(2) versus formate production was found to be related to the thermodynamics of formate dehydrogenation to CO(2)/H(2). This transition was associated with a shift in the catabolic products, from butyrate and acetate to ethanol and acetate, likely due to a decrease in the oxidation state of the electron carriers in the cell. The product spectrum of the MCF process as a function of the pH could largely be explained using general biochemical considerations.     

Plants to power: bioenergy to fuel the future

Bioenergy should play an essential part in reaching targets to replace petroleum-based transportation fuels with a viable alternative, and in reducing long-term carbon dioxide emissions, if environmental and economic sustainability are considered carefully. Here, we review different platforms, crops, and biotechnology-based improvements for sustainable bioenergy. Among the different platforms, there are two obvious advantages to using lignocellulosic biomass for ethanol production: higher net energy gain and lower production costs. However, the use of lignocellulosic ethanol as a viable alternative to petroleum-based transportation fuels largely depends on plant biotechnology breakthroughs. We examine how biotechnology, such as lignin modification, abiotic stress resistance, nutrition usage, in planta expression of cell wall digestion enzymes, biomass production, feedstock establishment, biocontainment of transgenes, metabolic engineering, and basic research, can be used to address the challenges faced by bioenergy crop production.     

Characterizing a stable methane-utilizing mixed culture used in the synthesis of a high-quality biopolymer in an open system

AIMS: To characterize a methane-utilizing poly-beta-hydroxybutyrate (PHB)-producing microbial community. METHODS AND RESULTS: Three different approaches based on microbiology, analytical chemistry and molecular biology were used to determine the composition of the mixed culture. The dominant species, Methylocystis sp. GB25, represents more than 86% of the total biomass. Seven accompanying bacterial species are present in the mixed culture of which two are methylotrophic bacteria and five are utilizers of complex carbon sources. Both these groups were found to be present at the same ratio with respect to each other. Results of fatty acid analysis and PCR-DGGE fingerprints reflect the stability of the mixed-culture composition in the open system during multiple continuous growth and polymer formation processes throughout a period of 29 months. The consistently high quality of the accumulated polymer further corroborates this finding. CONCLUSION: The methane-utilizing mixed culture has the potential of self-regulation resulting in a stable composition even under non-aseptic conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: Avoiding the necessity of sterile conditions, as demonstrated in this paper, is an important step towards the development of a viable large-scale process for the production of PHB using cheap substrates like methane from natural or renewable sources. This is the first report characterizing a bacterial mixed culture being used for the biotechnological production of a high-value product in an open system.     

Next‐Generation Industrial Biotechnology‐Transforming the Current Industrial Biotechnology into Competitive Processes

The chemical industry has made a contribution to modern society by providing cost‐competitive products for our daily use. However, it now faces a serious challenge regarding environmental pollutions and greenhouse gas emission. With the rapid development of molecular biology, biochemistry, and synthetic biology, industrial biotechnology has evolved to become more efficient for production of chemicals and materials. However, in contrast to chemical industries, current industrial biotechnology (CIB) is still not competitive for production of chemicals, materials, and biofuels due to their low efficiency and complicated sterilization processes as well as high‐energy consumption. It must be further developed into “next‐generation industrial biotechnology” (NGIB), which is low‐cost mixed substrates based on less freshwater consumption, energy‐saving, and long‐lasting open continuous intelligent processing, overcoming the shortcomings of CIB and transforming the CIB into competitive processes. Contamination‐resistant microorganism as chassis is the key to a successful NGIB, which requires resistance to microbial or phage contaminations, and available tools and methods for metabolic or synthetic biology engineering. This review proposes a list of contamination‐resistant bacteria and takes Halomonas spp. as an example for the production of a variety of products, including polyhydroxyalkanoates under open‐ and continuous‐processing conditions proposed for NGIB.     

Production of ethanol,organic acids and hydrogen: an opportunity for mixed culture biotechnology?

Anaerobic fermentation of biodegradable organic materials is usually carried out to obtain the final product, methane, a valuable energy source. However, it is also well known that various intermediates are produced in this process, e.g. ethanol, volatile organic acids and hydrogen. All these species have applications and value as fuels or chemicals. This paper shows a critical analysis of the potential of using anaerobic fermentation by mixed cultures to produce intermediates, e.g. ethanol, acetic, lactic and butyric acid and hydrogen, rather than methane. This paper discusses the current processes to produce these chemicals and compares them with the alternative approach of using open mixed cultures to produce them simultaneously via fermentation from renewable resources. None of these chemicals is currently produced via mixed culture fermentation: ethanol and lactic acid are usually produced in pure culture fermentation using food crops, e.g. corn or sugar cane, as starting materials; hydrogen, acetic and butyric acids are mainly produced via chemical synthesis from fossil fuel derived starting materials. A possible flow-sheet for the production of these chemicals from organic waste using mixed culture fermentation is proposed and the advantages and disadvantages of this process compared to current processes are critically discussed. The paper also discusses the research challenges which need to be addressed to make this process feasible.     

Microbial Production of Hydrogen by Mixed Culture Technologies: A Review

With its high energy content and clean combustion, hydrogen is recognized as a renewable clean energy source with enormous potential. Biological hydrogen production is a promising alternative with significant advantages over conventional petroleum‐derived chemical processes. Sustainable hydrogen production from renewable resources such as cassava, wastewater, and other agricultural waste is economically feasible for industrial applications. So far, the major bottlenecks in large‐scale biological hydrogen production are the low production rate and yield. This review discusses the various factors that affect the metabolic pathways of dark hydrogen production, and highlights the state‐of‐the‐art development of mixed culture technology. The aim of this review is to provide suggestions for the future directions of mixed culture technology, as well as by‐product valorization in dark fermentation.     

Biotechnology in the wood industry

Wood is a natural, biodegradable and renewable raw material, used in construction and as a feedstock in the paper and wood product industries and in fuel production. Traditionally, biotechnology found little attention in the wood product industries, apart from in paper manufacture. Now, due to growing environmental concern and increasing scientific knowledge, legal restrictions to conventional processes have altered the situation. Biotechnological approaches in the area of wood protection aim at enhancing the treatability of wood with preservatives and replacing chemicals with biological control agents. The substitution of conventional chemical glues in the manufacturing of board materials is achieved through the application of fungal cultures and isolated fungal enzymes. Moreover, biotechnology plays an important role in the waste remediation of preservative-treated waste wood.     

Recent advances in biodegradation in China: New microorganisms and pathways,biodegradation engineering,and bioenergy from pollutant biodegradation

We are facing serious environmental challenges, and environmental biotechnology is an enabling technology to reduce or eliminate pollution. In recent years, environmental pollution in China has been receiving great attention, and this paper provides an up-to-date review on progress in biodegradation research in China. This progress includes the isolation of extremophilic microorganisms for pollutant degradation in extreme conditions and the study of genes and enzymes related to biodegradation pathways. Biodegradation engineering has potential as an interesting and powerful platform, where genetic engineering, process engineering, and signal transduction engineering are applied together. In addition, pollutant treatment combined with the production of renewable sources of bioenergy by microorganisms is attractive.     

Impact of dissolved hydrogen partial pressure on mixed culture fermentations

Mixed culture fermentations are of interest for the low-cost production of organic acids from complex agricultural waste streams. Models are developed for these processes in order to predict the product spectrum as a function of the environmental process conditions. An important assumption in many existing models for anaerobic mixed culture fermentations is that the NADH/NAD⁺ ratio is directly coupled to the dissolved hydrogen partial pressure (pH_{2, liquid}). In this study, this assumption was tested experimentally with mixed culture chemostats operated at dilution rates of 0.05 and 0.125 h^?1 for a wide range of calculated dissolved hydrogen partial pressures (0.04–6.8 atm). No correlation was found between pH_{2, liquid} and the NADH/NAD⁺ ratio. This result, together with thermodynamic calculations, suggests that additional electron carriers such as ferredoxin and formate should be included in models predicting product formation by mixed cultures.     

Atmospheric impact of bioenergy based on perennial crop (reed canary grass,Phalaris arundinaceae,L.) cultivation on a drained boreal organic soil

Marginal organic soils, abundant in the boreal region, are being increasingly used for bioenergy crop cultivation. Using long‐term field experimental data on greenhouse gas (GHG) balance from a perennial bioenergy crop [reed canary grass (RCG), Phalaris arundinaceae L.] cultivated on a drained organic soil as an example, we show here for the first time that, with a proper cultivation and land‐use practice, environmentally sound bioenergy production is possible on these problematic soil types. We performed a life cycle assessment (LCA) for RCG on this organic soil. We found that, on an average, this system produces 40% less CO₂‐equivalents per MWh of energy in comparison with a conventional energy source such as coal. Climatic conditions regulating the RCG carbon exchange processes have a high impact on the benefits from this bioenergy production system. Under appropriate hydrological conditions, this system can even be carbon‐negative. An LCA sensitivity analysis revealed that net ecosystem CO₂ exchange and crop yield are the major LCA components, while non‐CO₂ GHG emissions and costs associated with crop production are the minor ones. Net bioenergy GHG emissions resulting from restricted net CO₂ uptake and low crop yields, due to climatic and moisture stress during dry years, were comparable with coal emissions. However, net bioenergy emissions during wet years with high net uptake and crop yield were only a third of the coal emissions. As long‐term experimental data on GHG balance of bioenergy production are scarce, scientific data stemming from field experiments are needed in shaping renewable energy source policies.     

Microalgae immobilization: current techniques and uses

Information about advances in immobilization techniques and biotechnology use of freshwater and marine microalgae is scattered. This work aims to bring together the main recent research about the topic. Passive and active immobilization techniques used on microalgae are listed and described in the text. Effect of immobilization on growth and metabolism of the cells is also reviewed. Current uses of immobilized microalgae include metabolite production, culture collection handling, obtaining of energy and removing of undesired or valuable substances from media (nutrients, metals and different pollutant agents). Applications of immobilized microalgae in environmental aquatic research have been recently increased: novel immobilization techniques as well as the use of living microalgae as biosensors in electronic devices designed to measure toxicity of substances and effluents demonstrated to be a very promising topic in biotechnology research. Recent research pointed out the advantages of mixed bacterial-algal co-immobilized systems in water treatment plants. Application of immobilized systems to the production of non-contaminant energy (as H(2) obtained from algal cultures) is also an important topic to be explored in the next years.