Bio-based Processes Research Articles

New waves underneath the purple strain.

SummarySuccessful merging of chemical and biotechnological operations is essential to achieve cost‐efficient industrialization of bio‐based processes. The demonstration of the use of syngas, derived from microwave assisted pyrolysis of municipal solid waste, for the improved growth and poly‐3‐hydroxybutyrate production in Rhodospirillium rubrum, stands out as an example of the synergistic contribution of chemical engineering and applied microbiology to sustainable biomaterial manufacturing, paving the way to similar applications for other syngas derived bioproducts.

Systematic development of biomass overproducing Scheffersomyces stipitis for high-cell-density fermentations

The development of economically feasible bio-based process requires efficient cell factories capable of producing the desired product at high titer under high-cell-density fermentation. Herein we present a combinatorial approach based on systems metabolic engineering and metabolic evolution for the development of efficient biomass-producing strain. Systems metabolic engineering guided by flux balance analysis (FBA) was first employed to rationally design mutant strains of Scheffersomyces stipitis with high biomass yield. By experimentally implementing these mutations, the biomass yield was improved by 30% in GPD1, 25% in TKL1, 30% in CIT1, and 44% in ZWF1 overexpressed mutants compared to wild-type. These designed mutants were further fine-tuned through metabolic evolution resulting in the maximal biomass yield of 0.49 g-cdw/g-glucose, which matches well with predicted yield phenotype. The constructed mutants are beneficial for biotechnology applications dealing with high cell titer cultivations. This work demonstrates a solid confirmation of systems metabolic engineering in combination with metabolic evolution approach for efficient strain development, which could assist in rapid optimization of cell factory for an economically viable and sustainable bio-based process.

Isolation and characterization of a hydrocarbonoclastic bacterial enrichment from total petroleum hydrocarbon contaminated sediments: potential candidates for bioaugmentation in bio-based processes.

Seven hydrocarbonoclastic new bacterial isolates were isolated from dredged sediments of a river estuary in Italy. The sediments were contaminated by shipyard activities since decades, mainly ascribable to the exploitation of diesel oil as the fuel for recreational and commercial navigation of watercrafts. The bacterial isolates were able to utilize diesel oil as sole carbon source. Their metabolic capacities were evaluated by GC-MS analysis, with reference to the depletion of both the normal and branched alkanes, the nC18 fatty acid methyl ester and the unresolved complex mixture of organic compounds. They were taxonomically identified as different species of Stenotrophomonas and Pseudomonas spp. by the combination of amplified ribosomal DNA restriction analysis (ARDRA) and repetitive sequence-based PCR (REP-PCR) analysis. The metabolic activities of interest were analyzed both in relation to the single bacterial strains and to the combination of the latter as a multibacterial species system. After 6days of incubation in mineral medium with diesel oil as sole carbon source, the Stenotrophomonas sp. M1 strain depleted 43-46% of Cn-alkane from C28 up to C30, 70% of the nC18 fatty acid methyl ester and the 46% of the unresolved complex mixture of organic compounds. On the other hand, the Pseudomonas sp. NM1 strain depleted the 76% of the nC18 fatty acid methyl ester, the 50% of the unresolved complex mixture of organic compounds. The bacterial multispecies system was able to completely deplete Cn-alkane from C28 up to C30 and to deplete the 95% of the unresolved complex mixture of organic compounds. The isolates, either as single strains and as a bacterial multispecies system, were proposed as candidates for bioaugmentation in bio-based processes for the decontamination of dredged sediments.

Model-based optimization of Scheffersomyces stipitis and Saccharomyces cerevisiae co-culture for efficient lignocellulosic ethanol production

The utilization of both C6 and C5 sugars is required for economical lignocellulosic bio-based processes. A co-culture system containing multiple strains of the same or different organisms holds promise for conversion of the sugar mixture available in different lignocellulosic feedstock into ethanol. Herein a co-culture kinetic model has been developed which can describe the co-cultivation of S. stipitis and S. cerevisiae for ethanol fermentation in mixed C6/C5 sugars. The predicted fermentation kinetics and ethanol production performance agreed well with experimental results, thus validating the model. The co-culture kinetic model has been implemented to design the optimal cell ratio for efficient conversion of rice straw or sugarcane bagasse feedstock into ethanol. The results reveal that the optimal co-culture system could enhance ethanol titer by up to 26 %, and ethanol productivity by up to 29 % compared to a single-strain culture. The maximum ethanol titer and productivity reached by the optimized co-culture was 46 and 0.49 g/l h, respectively. The co-culture model described here is a useful tool for rapid optimization of S. stipitis/S. cerevisiae co-culture for efficient and sustainable lignocellulosic ethanol production to meet the economic requirements of the lignocellulosic ethanol industry. The developed modeling tool also provides a systematic strategy for designing the optimal cell ratio of co-culture, leading to efficient fermentation of the C6/C5 sugars available in any biomass feedstock.

Removing chiral contamination of lactate solutions by selective metabolism of the d-enantiomer

A bio-based process is appealing for purification of L-lactic acid, the major enantiomer of polylactic acid syrup, generated by thermochemical processes at the end of life of PLA-based plastics, from its chiral impurity, D-lactic acid, before reuse. Polylactic acid (PLA), a renewable alternative to petroleum-derived plastics, contains a mixture of L- and D-lactic acid (LA) isomers with the L-isomer dominating (up to 95%). A novel bio-based process was developed to produce chirally pure L-LA from syrup produced during recycling of PLA-plastics. This process utilizes an engineered Escherichia coli (strain DC1001) containing novel gene deletions (lld, ykg) that eliminated the oxidative metabolism of L-lactate, leaving the membrane-bound D-lactate dehydrogenases to selectively metabolize the D-isomer. Strain DC1001 removed 8.7g D-lactate l(-1) from a PLA-syrup containing 135g total lactic acid l(-1) in 24h. Average rates of removal of D-lactic acid were 0.25g D-lactate h(-1) (g cell dry weight)(-1) and 0.36g D-lactate l(-1) h(-1). Bio-based purification of PLA-syrup utilizing E. coli strain DC1001 is an attractive process step during recycling of PLA-plastics. This selective oxidation process can also be used to remove chiral contamination of L-lactate in medical applications.

A New Approach for the Design and Assessment of Bio-based Chemical Processes toward Sustainability

Substituting biomass for fossil-based feedstock is a potential scenario of chemical production. To achieve sustainable production, various issues must be considered, such as the constraints of limited biomass resources, feasibility of production technologies, and competitiveness of bio-based chemical products. In this study, we developed a new framework for the design and assessment of bio-based chemical processes. Both monetary and nonmonetary indicators including production cost, energy consumption, environmental impact, and safety hazards are considered. The framework was applied to the design and assessment of both existing and emerging new bio-based processes producing potential platform chemicals from bioethanol. On the basis of the design and assessment results, the suitable biomass feedstock, a sustainable synthesis process, a competitive bio-based product, and an appropriate production scenario can be selected. Using data available at the conceptual design stage, the framework can be applied to synthesize high-potential bio-based chemical processes for future sustainability development.

New biofuel alternatives: integrating waste management and single cell oil production.

Concerns about greenhouse gas emissions have increased research efforts into alternatives in bio-based processes. With regard to transport fuel, bioethanol and biodiesel are still the main biofuels used. It is expected that future production of these biofuels will be based on processes using either non-food competing biomasses, or characterised by low CO2 emissions. Many microorganisms, such as microalgae, yeast, bacteria and fungi, have the ability to accumulate oils under special culture conditions. Microbial oils might become one of the potential feed-stocks for biodiesel production in the near future. The use of these oils is currently under extensive research in order to reduce production costs associated with the fermentation process, which is a crucial factor to increase economic feasibility. An important way to reduce processing costs is the use of wastes as carbon sources. The aim of the present review is to describe the main aspects related to the use of different oleaginous microorganisms for lipid production and their performance when using bio-wastes. The possibilities for combining hydrogen (H2) and lipid production are also explored in an attempt for improving the economic feasibility of the process.

Methodologies and perspectives of proteomics applied to filamentous fungi: from sample preparation to secretome analysis.

Filamentous fungi possess the extraordinary ability to digest complex biomasses and mineralize numerous xenobiotics, as consequence of their aptitude to sensing the environment and regulating their intra and extra cellular proteins, producing drastic changes in proteome and secretome composition. Recent advancement in proteomic technologies offers an exciting opportunity to reveal the fluctuations of fungal proteins and enzymes, responsible for their metabolic adaptation to a large variety of environmental conditions. Here, an overview of the most commonly used proteomic strategies will be provided; this paper will range from sample preparation to gel-free and gel-based proteomics, discussing pros and cons of each mentioned state-of-the-art technique. The main focus will be kept on filamentous fungi. Due to the biotechnological relevance of lignocellulose degrading fungi, special attention will be finally given to their extracellular proteome, or secretome. Secreted proteins and enzymes will be discussed in relation to their involvement in bio-based processes, such as biomass deconstruction and mycoremediation.

Development and validation of an rDNA operon based primer walking strategy applicable to de novo bacterial genome finishing.

Advances in sequencing technology have drastically increased the depth and feasibility of bacterial genome sequencing. However, little information is available that details the specific techniques and procedures employed during genome sequencing despite the large numbers of published genomes. Shotgun approaches employed by second-generation sequencing platforms has necessitated the development of robust bioinformatics tools for in silico assembly, and complete assembly is limited by the presence of repetitive DNA sequences and multi-copy operons. Typically, re-sequencing with multiple platforms and laborious, targeted Sanger sequencing are employed to finish a draft bacterial genome. Here we describe a novel strategy based on the identification and targeted sequencing of repetitive rDNA operons to expedite bacterial genome assembly and finishing. Our strategy was validated by finishing the genome of Paenibacillus polymyxa strain CR1, a bacterium with potential in sustainable agriculture and bio-based processes. An analysis of the 38 contigs contained in the P. polymyxa strain CR1 draft genome revealed 12 repetitive rDNA operons with varied intragenic and flanking regions of variable length, unanimously located at contig boundaries and within contig gaps. These highly similar but not identical rDNA operons were experimentally verified and sequenced simultaneously with multiple, specially designed primer sets. This approach also identified and corrected significant sequence rearrangement generated during the initial in silico assembly of sequencing reads. Our approach reduces the required effort associated with blind primer walking for contig assembly, increasing both the speed and feasibility of genome finishing. Our study further reinforces the notion that repetitive DNA elements are major limiting factors for genome finishing. Moreover, we provided a step-by-step workflow for genome finishing, which may guide future bacterial genome finishing projects.

Microbial production of short chain diols.

Short chain diols (propanediols, butanediols, pentanediols) have been widely used in bulk and fine chemical industries as fuels, solvents, polymer monomers and pharmaceutical precursors. The chemical production of short chain diols from fossil resources has been developed and optimized for decades. Consideration of the exhausting fossil resources and the increasing environment issues, the bio-based process to produce short chain diols is attracting interests. Currently, a variety of biotechnologies have been developed for the microbial production of the short chain diols from renewable feed-stocks. In order to efficiently produce bio-diols, the techniques like metabolically engineering the production strains, optimization of the fermentation processes, and integration of a reasonable downstream recovery processes have been thoroughly investigated. In this review, we summarized the recent development in the whole process of bio-diols production including substrate, microorganism, metabolic pathway, fermentation process and downstream process.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-014-0165-5) contains supplementary material, which is available to authorized users.

Combination of algae and yeast fermentation for an integrated process to produce single cell oils.

Economic and ecological reasons cause the industry to develop new innovative bio-based processes for the production of oil as renewable feedstock. Petroleum resources are expected to be depleted in the near future. Plant oils as sole substituent are highly criticized because of the competitive utilization of the agricultural area for food and energy feedstock production. Microbial lipids of oleaginous microorganisms are therefore a suitable alternative. To decrease production costs of microbial lipids and gain spatial independence from industrial sites of CO2 emission, a combination of heterotrophic and phototrophic cultivation with integrated CO2 recycling was investigated in this study. A feasibility study on a semi-pilot scale was conducted and showed that the cultivation of the oleaginous yeast Cryptococcus curvatus on a 1.2-L scale was sufficient to supply a culture of the oleaginous microalgae Phaeodactylum tricornutum in a 21-L bubble column reactor with CO2 while single cell oils were produced in both processes due to a nutrient limitation.

Rational design of ornithine decarboxylase with high catalytic activity for the production of putrescine

Putrescine finds wide industrial applications in the synthesis of polymers, pharmaceuticals, agrochemicals, and surfactants. Owing to economic and environmental concerns, the microbial production of putrescine has attracted a great deal of attention, and ornithine decarboxylase (ODC) is known to be a key enzyme in the biosynthetic pathway. Herein, we present the design of ODC from Escherichia coli with high catalytic efficiency using a structure-based rational approach. Through a substrate docking into the model structure of the enzyme, we first selected residues that might lead to an increase in catalytic activity. Of the selected residues that are located in the α-helix and the loops constituting the substrate entry site, a mutational analysis of the single mutants identified two key residues, I163 and E165. A combination of two single mutations resulted in a 62.5-fold increase in the catalytic efficiency when compared with the wild-type enzyme. Molecular dynamics simulations of the best mutant revealed that the substrate entry site becomes more flexible through mutations, while stabilizing the formation of the dimeric interface of the enzyme. Our approach can be applied to the design of other decarboxylases with high catalytic efficiency for the production of various chemicals through bio-based processes.

Fuels and plastics from lignocellulosic biomass via the furan pathway; a technical analysis

Biorefineries convert biomass into bio-based products, which have the potential to replace typical products produced by petroleum refineries. They provide a technology platform to reduce anthropogenic greenhouse gas emissions, increase security of supply and reduce the dependency on crude oil. The biorefinery concept presented in this paper focuses on a combination of (1) organosolv fractionation to produce carbohydrates from lignocellulosic biomass and (2) the furan technology to convert carbohydrates into polyethylene furanoate (PEF), a bio-based alternative to polyethylene terephthalate (PET), and furfuryl ethyl ether (FEE), a bio-based transportation fuel component. The goal of this paper is to determine the mass and energy balances of the production of PEF and FEE from lignocellulosic biomass and indicate the benefits, as well as potential bottlenecks in the coupling of organosolv and furan chemistry as a biorefinery concept. Three cases are defined, modeled and analyzed, each focusing on a different approach to combine the organosolv and furan conversion technologies and determine the possibility and degree of integration. Modeling results based on experimental data and expert judgments show that wheat straw, as an example of lignocellulosic biomass, can be converted into PEF and FEE at yields between 20 and 40 w/w%, based on total input, while energetic efficiencies are between 30 and 40%. This is comparable or even better compared to other upcoming bio-based processes, e.g. 15–35% yield for second generation bio-ethanol production and 25–50% energy efficiency. The conclusion is that in each of the three cases presented bio-based fuels and plastics can be produced via the furan pathway at efficiencies that constitute a viable option from a technological point of view.

Microbial production of short chain diols

Short chain diols (propanediols, butanediols, pentanediols) have been widely used in bulk and fine chemical industries as fuels, solvents, polymer monomers and pharmaceutical precursors. The chemical production of short chain diols from fossil resources has been developed and optimized for decades. Consideration of the exhausting fossil resources and the increasing environment issues, the bio-based process to produce short chain diols is attracting interests. Currently, a variety of biotechnologies have been developed for the microbial production of the short chain diols from renewable feed-stocks. In order to efficiently produce bio-diols, the techniques like metabolically engineering the production strains, optimization of the fermentation processes, and integration of a reasonable downstream recovery processes have been thoroughly investigated. In this review, we summarized the recent development in the whole process of bio-diols production including substrate, microorganism, metabolic pathway, fermentation process and downstream process.

Sustainability metrics of 1-butanol

The production of chemicals from renewable resources often has to compete with a petrochemical process, optimized during several decades. Four sustainability metrics were used to compare the production of 1-butanol via the petrochemical and biobased processes: material efficiency, energy efficiency, land use and total costs. The selected petrochemical process is the oxo synthesis, i.e., the hydroformylation of propene, followed by the hydrogenation of the formed aldehydes with a yield of 95% 1-butanol. The selected biobased process is the anaerobic continuous acetone–butanol–ethanol (ABE) fermentation on glucose substrate from maize starch and a product recovery via distillation with a yield of 0.42gABEg−1 glucose. The petrochemical process has significantly higher material and energy efficiencies, compared to the biobased process. For the biobased process, land is used to produce the biomass (0.29haton−1), while no land is used for the petrochemical process. Production costs are higher for the biobased process (1041EURton−1), compared to the petrochemical process (915EURton−1). Based on the four metrics, the petrochemical process is preferable to the biobased process. However, biomass for sustainable fuels and chemicals will be the only resource for future generations. The efficiency of the biobutanol production can be improved by altering upstream processes, by metabolic engineering, by decreasing byproduct formation and by improving in situ product recovery techniques.

Metabolix launches bio-based additive to enhance recycled PVC performance

Biomaterials firm Metabolix, Inc of Cambridge, MA, USA, has launched I6003rp, a new bio-based polymeric performance additive for recycled polyvinyl chloride (PVC). The certified bio-based process aid lowers processing torque and improves properties, the company claims. I6003rp also reportedly demonstrates excellent miscibility in PVC when used in combination with PVC recyclate.

Green upgrading of succinate using dimethyl carbonate for a better integration with fermentative production

In this work we present a new process alternative comprising bio-based succinate production, its recovery by ion exchange and further upgrading to dimethyl succinate via O-alkylation by direct downstream catalysis. The use of dimethyl carbonate as alkylating agent permits the in-process generation of a mobile bicarbonate anion, playing the role of counter-ion during succinate capture and then transported upstream where is used as inorganic carbon source and neutralizing agent required for most bacterial succinate fermentations. Succinate recovery using a strong anion exchange resin in the bicarbonate form presented good equilibrium characteristics and column performance. Subsequently, dimethyl succinate was produced in high yield (96%) using dimethyl carbonate as a solvent and reagent and the kinetics of the system were described by a pseudo-first order model. Throughout the reaction and in the presence of water, it was verified that the resin is effectively regenerated to its bicarbonate form maintaining its ion exchange capacity. As a main advantage, the validated concept leads to an improved bio-based process for succinate to ester derivatives where no stoichiometric waste is produced, contrary to conventional acidification–esterification operations.

Early‐Stage Comparative Sustainability Assessment of New Bio‐based Processes

Our increasing demand for materials and energy has put critical roadblocks on our path towards a sustainable society. To remove these roadblocks, it is important to engage in smart research and development (R&D). We present an early-stage sustainability assessment framework that is used to analyze eight new bio-based process alternatives developed within the CatchBio research consortium in the Netherlands. This assessment relies on a multi-criteria approach, integrating the performance of chemical conversions based on five indicators into an index value. These indicators encompass economics, environmental impact, hazards and risks thereby incorporating elements of green chemistry principles, and techno-economic and life cycle assessments. The analyzed bio-based options target the production of fuels and chemicals through chemical catalysis. For each bio-based process, two R&D stages (current laboratory and expected future) are assessed against a comparable conventional process. The multi-criteria assessment in combination with the uncertainty and scenario analysis shows that the chemical production processes using biomass as feedstock can provide potential sustainability benefits over conventional alternatives. However, further development is necessary to realize the potential benefits from biomass gasification and pyrolysis processes for fuel production. This early stage assessment is intended as an input for R&D decision making to support optimal allocation and utilization of resources to further develop promising bio-based processes.

Process Development for Functionalization of Cotton with Silver Nanoparticles Synthesized by Bio-based Approaches

Cotton is a widely used raw material for textiles but drawbacks regarding their poor mechanical properties often limit their applications as functional materials. The present investigation involved process development for one step coating of cotton with silver nanoparticles (SNP) synthesized using Azadirachta indica and Citrus limon extract to develop functional textiles. Addition of starch to functional textiles led to efficient binding of nanoparticles to fabric and led to drastic decrease in release of silver from fabricated textiles after ten washing cycles enhancing their environment friendliness. Differential scanning calorimetry, scanning electron microscopy, FTIR analysis and mechanical studies demonstrated efficient binding of nanoparticles to fabric through bio-based processes. The functionalized textiles developed by the bio-based methods showed significant antibacterial activity against E. coli and S. aureus (with 99% microbial reduction). Present work offers a simple procedure for coating SNP using bio-based approaches with promising applications in specialized functions. Keywords: Silver nanoparticle, A. indica, C. limon, textile, binding, antimicrobial activity.

Angling for uniqueness in enzymatic preparation of glycosides.

In the early days of biocatalysis, limitations of an enzyme modeled the enzymatic applications; nowadays the enzyme can be engineered to be suitable for the process requirements. This is a general bird’s-eye view and as such cannot be specific for articulated situations found in different classes of enzymes or for selected enzymatic processes. As far as the enzymatic preparation of glycosides is concerned, recent scientific literature is awash with examples of uniqueness related to the features of the biocatalyst (yield, substrate specificity, regioselectivity, and resistance to a particular reaction condition). The invention of glycosynthases is just one of the aspects that has thrust forward the research in this field. Protein engineering, metagenomics and reaction engineering have led to the discovery of an expanding number of novel enzymes and to the setting up of new bio-based processes for the preparation of glycosides. In this review, new examples from the last decade are compiled with attention both to cases in which naturally present, as well as genetically inserted, characteristics of the catalysts make them attractive for biocatalysis.