Biofuel Production Process Research Articles

Thermochemical valorization of camelina straw waste via fast pyrolysis

The present work investigates the thermochemical valorization of camelina straw, which is a waste generated during the harvesting of Camelina sativa, an oilseed crop for the production of biodiesel or hydrotreated vegetable oil (HVO). In particular, it is focused on obtaining bio-oil via thermal or catalytic fast pyrolysis, which would be the first stage on a sequence of chemical processes for biofuel production. The catalytic interference of the inorganic matter present in the biomass was studied by preparing a batch of de-ashed camelina straw by washing with diluted nitric acid. Chemical analysis revealed this treatment effectively removed alkaline (K and Na) and alkaline earth (Ca and Mg) metals. Pyrolysis of de-ashed camelina straw led to higher mass and energy yields of bio-oil in water-free basis (bio-oil*), but with higher oxygen concentration. Catalytic pyrolysis over HZSM-5 was also studied in both raw and de-ashed feedstocks. This catalyst promoted mainly decarbonylation and decarboxylation reactions of the pyrolysis vapors, leading to much higher gas yields and lower of bio-oil*, but with better quality. Catalytic pyrolysis of untreated camelina straw exhibited a synergetic effect between both the inorganic matter and the external HZSM-5 catalyst, so that bio-oil* yield was the lowest (20 wt%) due to an extensive deoxygenation (18 wt% oxygen content), which resulted in the highest HHV obtained (37.3 MJ/kgdb). Significant differences were also found on the molecular composition of the bio-oils* with larger proportion of anhydro sugars when the biomass was de-ashed, while HZSM-5 strongly promoted the formation of oxygenated aromatics and aromatic hydrocarbons.

Toward high solids loading process for lignocellulosic biofuel production at a low cost.

High solids loadings (>18 wt%) in enzymatic hydrolysis and fermentation are desired for lignocellulosic biofuel production at a high titer and low cost. However, sugar conversion and ethanol yield decrease with increasing solids loading. The factor(s) limiting sugar conversion at high solids loading is not clearly understood. In the present study, we investigated the effect of solids loading on simultaneous saccharification and co-fermentation (SSCF) of AFEX™ (ammonia fiber expansion) pretreated corn stover for ethanol production using a xylose fermenting strain Saccharomyces cerevisiae 424A(LNH-ST). Decreased sugar conversion and ethanol yield with increasing solids loading were also observed. End-product (ethanol) was proven to be the major cause of this issue and increased degradation products with increasing solids loading was also a cause. For the first time, we show that with in situ removal of end-product by performing SSCF aerobically, sugar conversion stopped decreasing with increasing solids loading and monomeric sugar conversion reached as high as 93% at a high solids loading of 24.9 wt%. Techno-economic analysis was employed to explore the economic possibilities of cellulosic ethanol production at high solids loadings. The results suggest that low-cost in situ removal of ethanol during SSCF would significantly improve the economics of high solids loading processes. Biotechnol. Bioeng. 2017;114: 980-989. © 2016 Wiley Periodicals, Inc.

Enhanced ethanol fermentation by engineered Saccharomyces cerevisiae strains with high spermidine contents.

Construction of robust and efficient yeast strains is a prerequisite for commercializing a biofuel production process. We have demonstrated that high intracellular spermidine (SPD) contents in Saccharomyces cerevisiae can lead to improved tolerance against various fermentation inhibitors, including furan derivatives and acetic acid. In this study, we examined the potential applicability of the S. cerevisiae strains with high SPD contents under two cases of ethanol fermentation: glucose fermentation in repeated-batch fermentations and xylose fermentation in the presence of fermentation inhibitors. During the sixteentimes of repeated-batch fermentations using glucose as a sole carbon source, the S. cerevisiae strains with high SPD contents maintained higher cell viability and ethanol productivities than a control strain with lower SPD contents. Specifically, at the sixteenth fermentation, the ethanol productivity of a S. cerevisiae strain with twofold higher SPD content was 31% higher than that of the control strain. When the SPD content was elevated in an engineered S. cerevisiae capable of fermenting xylose, the resulting S. cerevisiae strain exhibited much 40-50% higher ethanol productivities than the control strain during the fermentations of synthetic hydrolysate containing high concentrations of fermentation inhibitors. These results suggest that the strain engineering strategy to increase SPD content is broadly applicable for engineering yeast strains for robust and efficient production of ethanol.

Medium-low temperature hydrothermal hydrolysis kinetic characteristics of concentrated wet microalgae biomass.

To improve microalgae biomass utilization efficiency during biofuel production process, medium-low temperature hydrothermal hydrolysis pretreatment was adopted in this study. The pretreatment kinetic characteristics of concentrated wet microalgae Chlorella vulgaris biomass (50 g/L) under medium-low temperature hydrolysis (100°C-200°C) were experimentally investigated. The hydrothermal hydrolysis kinetics describing the coupled effects of temperature, initial pressure and retention time then were proposed using response surface methodology (RSM). The maximum carbohydrate yield reached 327.3 mg/g dried biomass under initial pressure of 4 MPa at reaction temperature of 150°C for 120 min. The maximum protein yield (321.5 mg/g dried biomass) was obtained under initial pressure of 4 MPa at reaction temperature of 200°C for 60 min. Based on the hydrothermal hydrolysis kinetic models, it was confirmed that temperature was the most important factor affecting both carbohydrate and protein release during hydrothermal hydrolysis process. Hydrothermal initial pressure and retention time were significant to carbohydrate release, but not to protein release. While, lipid was mainly distributed in microalgae residual and almost did not exist in supernatant (about 8.03 mg/g). And with assistance of mixed hexane and methanol (the ratio of hexane to methanol was 7:3), 67.69% of microalgae lipid was extracted out from hydrothermal hydrolysed microalgae residual (123.3 mg/g dried biomass). Keywords: hydrothermal hydrolysis, kinetic characteristics, microalgae, medium-low temperature, biofuel, response surface methodology DOI: 10.3965/j.ijabe.20171001.2699 Citation: Ding X J, Huang Y, Liao Q, Fu Q, Xia A, Xiao C, et al. Medium-low temperature hydrothermal hydrolysis kinetic characteristics of concentrated wet microalgae biomass. Int J Agric & Biol Eng, 2017; 10(1): 154–162.

Comparative LCA of Flocculation for the Harvesting of Microalgae for Biofuels Production

In recent years, the use of Life Cycle Analysis (LCA) to evaluate environmental benefits resulting from the production of biofuel from microalgae has continued to evolve. Literature in this field shows that one of the main challenges associated with the effect of biofuel production on the environment is the high energy consumption necessary in the microalgae harvesting phase to achieve the level of dewatering required to the next steps. Moreover, detailed LCAs specifically focused on the assessment of alternative technologies for the harvesting of microalgae have yet to be presented. As such, the aim of this paper is to analyze the potential environmental benefits and shortcomings arising from the use of flocculation for the harvesting of microalgae in the biofuel production process, with particular attention to the Canadian context.The method employed is a comparative LCA, where two alternative scenarios based on the application of two harvesting technologies are taken into account: (1) flocculation and centrifugation and (2) direct centrifugation (without flocculation). The calculations of environmental impact and the sensitivity analysis are performed with the SimaPro software.

Enhanced microalgal lipid extraction using bio-based solvents for sustainable biofuel production

Bio-based solvents are efficient replacements for traditional fossil-derived solvents in microalgal lipid extraction towards developing environmentally friendly biofuel production processes.

Application, Deactivation, and Regeneration of Heterogeneous Catalysts in Bio-Oil Upgrading

The massive consumption of fossil fuels and associated environmental issues are leading to an increased interest in alternative resources such as biofuels. The renewable biofuels can be upgraded from bio-oils that are derived from biomass pyrolysis. Catalytic cracking and hydrodeoxygenation (HDO) are two of the most promising bio-oil upgrading processes for biofuel production. Heterogeneous catalysts are essential for upgrading bio-oil into hydrocarbon biofuel. Although advances have been achieved, the deactivation and regeneration of catalysts still remains a challenge. This review focuses on the current progress and challenges of heterogeneous catalyst application, deactivation, and regeneration. The technologies of catalysts deactivation, reduction, and regeneration for improving catalyst activity and stability are discussed. Some suggestions for future research including catalyst mechanism, catalyst development, process integration, and biomass modification for the production of hydrocarbon biofuels are provided.

Improved sugar yields from biomass sorghum feedstocks: comparing low-lignin mutants and pretreatment chemistries.

BackgroundFor biofuel production processes to be economically efficient, it is essential to maximize the production of monomeric carbohydrates from the structural carbohydrates of feedstocks. One strategy for maximizing carbohydrate production is to identify less recalcitrant feedstock cultivars by performing some type of experimental screening on a large and diverse set of candidate materials, or by identifying genetic modifications (random or directed mutations or transgenic plants) that provide decreased recalcitrance. Economic efficiency can also be increased using additional pretreatment processes such as deacetylation, which uses dilute NaOH to remove the acetyl groups of hemicellulose prior to dilute acid pretreatment. In this work, we used a laboratory-scale screening tool that mimics relevant thermochemical pretreatment conditions to compare the total sugar yield of three near-isogenic brown midrib (bmr) mutant lines and the wild-type (WT) sorghum cultivar. We then compared results obtained from the laboratory-scale screening pretreatment assay to a large-scale pretreatment system.ResultsAfter pretreatment and enzymatic hydrolysis, the bmr mutants had higher total sugar yields than the WT sorghum cultivar. Increased pretreatment temperatures increased reactivity for all sorghum samples reducing the differences observed at lower reaction temperatures. Deacetylation prior to dilute acid pretreatment increased the total sugar yield for all four sorghum samples, and reduced the differences in total sugar yields among them, but solubilized a sizable fraction of the non-structural carbohydrates. The general trends of increased total sugar yield in the bmr mutant compared to the WT seen at the laboratory scale were observed at the large-scale system. However, in the larger reactor system, the measured total sugar yields were lower and the difference in total sugar yield between the WT and bmr sorghum was larger.ConclusionsSorghum bmr mutants, which have a reduced lignin content showed higher total sugar yields than the WT cultivar after dilute acid pretreatment and enzymatic hydrolysis. Deacetylation prior to dilute acid pretreatment increased the total sugar yield for all four sorghum samples. However, since deacetylation also solubilizes a large fraction of the non-structural carbohydrates, the ability to derive value from these solubilized sugars will depend greatly on the proposed conversion process.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0667-y) contains supplementary material, which is available to authorized users.

Bioconversion of hemicelluloses of lignocellulosic biomass to ethanol: an attempt to utilize pentose sugars

ABSTRACTBiofuels are the imperative commodities influential in future bioeconomy that can be made sustainable by utilizing hemicelluloses which are the primary unutilized residues obtained from a lignocellulosic ethanol refinery. The abundant fraction in these hemicelluloses include pentoses (mainly xylose). The rationale behind the huge loss of hemicellulosic fraction is their heteropolymeric nature, low fermentability, lack of pentose specific transporters and enzyme cascades required for pentose fermentation by natural yeast strains. These lacunae lead to lower overall yield and productivity of ethanol thus rendering the entire process uneconomical. However, improvements in the conventional cellulose to ethanol know-how can trigger evolution towards bioeconomy. For industrial implementation of process technology, hemicellulosic stream should be integrated as a primary fraction along with celluloses for converting into ethanol. Thus, a novel hexose and pentose co-fermenting yeast strain is essential for a consolidated bioprocess aiming to achieve process economization. Considering a range of value added platform chemicals other than ethanol, such as xylitol, sorbitol, furfurals, hydroxyl methyl furfurals, ethylene glycol, glycolic acid, acetic acid, acetone, ethylene, and other residual biomass based products, such as biomanure, can render the biofuel production process more economical as multiple products may be obtained from the same biomass making the production process a comprehensive one. Therefore, the present review showcases the various technical impediments, breakthroughs, and bioeconomic aspects in the field of pentose to ethanol research which are instrumental in the future of the biofuel sector to address escalating oil prices and diminishing oil reserves.

Biofuels Are Not the Full Answer, but They Can Be Part of a Solar-Oriented Solution to Our Energy-Climate Dual Crisis

Biofuels are part of a trend towards increased util ization of local resources for energy generation, especially those with greenhouse gases’ reduction b enefits. In this paper, we argue that photosynthesi s dependent biofuels ‐ with a practical maximum of 2% energy conversion efficiency ‐ can only satisfy a fraction of the world’s liquid fuel needs. Even tha t cannot be achieved without the deployment of 2 nd generation biofuel production processes ‐ still und er development ‐ which accept cellulosic and microalgal feed stocks, not food crops. For these f eed stocks to be sustainably cultivated, they must be grown on non-arable lands using limited amounts of irrigation water and fossil-fuel based agrichemical s. Additionally, biofuels pose their own unique set of issues to consider such arable land use, water consumption among other metrics. Finally, biofuels should be viewed as just one way of converting sunlight into energy besides other means such as so lar heating and photovoltaics, with the latter opti on realizing an energy conversion efficiency exceeding 10%. By reviewing the potential alternatives to oi l and petrol based fuels, it can be seen that biofuel s are only a viable replacement when used in conjun ction with other renewable energy sources.

Modelling and thermodynamic assessment of biofuel production process from biomass with the SAFT-VR approach

Two reliable methods for fuel production from biomass including supercritical water gasification (SCWG) and hydrothermal upgrading (HTU) have been studied. The Gibbs free energy minimization method was used to calculate equilibrium mole fraction of the reaction products for glucose and cellulose supercritical water gasification and hydrothermal upgrading of wood as feed biomass. The phase behavior and phase equilibrium of the separators used for the target product separation (hydrogen in SCWG and biocrude in HTU processes) was carried out successfully using the SAFT-VR equation of state for the first time. The effect of pressure and temperature on SCWG process was investigated and turned out that temperature increase causes more hydrogen production while the pressure effect was insignificant. Finally, the exergy analysis and efficiency calculations were performed respectively for HTU and SCWG processes. According to the outcomes, the exergy efficiency and SCWG efficiency for glucose were obtained as 83.63% and 88.28%.

Highly Efficient Process for Production of Biofuel from Ethanol Catalyzed by Ruthenium Pincer Complexes.

A highly efficient ruthenium pincer-catalyzed Guerbet-type process for the production of biofuel from ethanol has been developed. It produces the highest conversion of ethanol (73.4%, 0.02 mol% catalyst) for a Guerbet-type reaction, including significant amounts of C4 (35.8% yield), C6 (28.2% yield), and C8 (9.4% yield) alcohols. Catalyst loadings as low as 0.001 mol% can be used, leading to a record turnover number of 18 209. Mechanistic studies reveal the likely active ruthenium species and the main deactivation process.

Economic implications of incorporating emission controls to mitigate air pollutants emitted from a modeled hydrocarbon‐fuel biorefinery in the United States

AbstractThe implementation of the US Renewable Fuel Standard is expected to increase the construction and operation of new biofuel facilities. Allowing this industry to grow without adversely affecting air quality is an important sustainability goal sought by multiple stakeholders. However, little is known about how the emission controls potentially required to comply with air quality regulations might impact biorefinery cost and deployment strategies such as siting and sizing. In this study, we use a baseline design for a lignocellulosic hydrocarbon biofuel production process to assess how the integration of emission controls impacts the minimum fuel selling price (MFSP) of the biofuel produced. We evaluate the change in MFSP for two cases as compared to the baseline design by incorporating (i) emission controls that ensure compliance with applicable federal air regulations and (ii) advanced control options that could be used to achieve potential best available control technology (BACT) emission limits. Our results indicate that compliance with federal air regulations can be achieved with minimal impact on biofuel cost (~$0.02 per gasoline gallon equivalent (GGE) higher than the baseline price of $5.10 GGE−1). However, if air emissions must be further reduced to meet potential BACT emission limits, the cost could increase nontrivially. For example, the MFSP could increase to $5.50 GGE−1 by adopting advanced emission controls to meet potential boiler BACT limits. Given tradeoffs among emission control costs, permitting requirements, and economies of scale, these results could help inform decisions about biorefinery siting and sizing and mitigate risks associated with air permitting. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. Biofuels, Bioproducts and Biorefining published by Society of Industrial Chemistry and John Wiley & Sons Ltd.

Effect of aging on lignin content, composition and enzymatic saccharification in Corymbia hybrids and parental taxa between years 9 and 12

Corymbia (a eucalypt) is an important forestry genus and a potential lignocellulosic bioenergy feedstock. The composition of the lignocellulosic cell wall significantly impacts pretreatment efficiency and conversion to biofuel but is variable and changes with age. In this study, we estimated Klason lignin content, composition, and monosaccharide (glucose and xylose) release after enzymatic saccharification of untreated and hydrothermally pretreated biomass from Corymbia parental species Corymbia torelliana (CT), Corymbia citriodora subsp. variegata (spotted gum; CCV), and interspecific F1 hybrids (CT × CCV) at ages 9 and 12 years from planting. Analysis of lignin composition derived from syringyl/guaiacyl monolignols (S/G) found significant differences among taxa, with CT S/G ratios (2.2 and 2.0) being significantly lower than CCV (2.6 and 2.3) or hybrids (2.5 and 2.3) at ages 9 and 12 respectively. In general, enzymatic saccharification yields from untreated biomass were significantly different among taxa, with CT (113 and 75 mg g−1) and hybrids (108 and 81 mg g−1) yielding significantly higher glucose from untreated biomass than CCV (82 and 56 mg g−1) at ages 9 and 12 respectively. Comparison of traits within taxa between ages 9 and 12 found S/G ratios and glucose yields from untreated biomass were significantly lower in CT, CCV and hybrid taxa. In conclusion, the formation of lignocellulosic cell walls is complex, influenced by genetics and age of material, requiring optimization of rotation age for biofuel production and other industrial processes.

Microalgae Recovery from Water for Biofuel Production Using CO2-Switchable Crystalline Nanocellulose.

There is a pressing need to develop efficient and sustainable approaches to harvesting microalgae for biofuel production and water treatment. CO2-switchable crystalline nanocellulose (CNC) modified with 1-(3-aminopropyl)imidazole (APIm) is proposed as a reversible coagulant for harvesting microalgae. Compared to native CNC, the positively charged APIm-modified CNC, which dispersed well in carbonated water, showed appreciable electrostatic interaction with negatively charged Chlorella vulgaris upon CO2-treatment. The gelation between the modified CNC, triggered by subsequent air sparging, can also enmesh adjacent microalgae and/or microalgae-modified CNC aggregates, thereby further enhancing harvesting efficiencies. Moreover, the surface charges and dispersion/gelation of APIm-modified CNC could be reversibly adjusted by alternatively sparging CO2/air. This CO2-switchability would make the reusability of redispersed CNC for further harvesting possible. After harvesting, the supernatant following sedimentation can be reused for microalgal cultivation without detrimental effects on cell growth. The use of this approach for harvesting microalgae presents an advantage to other current methods available because all materials involved, including the cellulose, CO2, and air, are natural and biocompatible without adverse effects on the downstream processing for biofuel production.

Effect of phosphate concentration on exergetic-based sustainability parameters of glucose fermentation by Ethanolic Mucor indicus

In this work, a holistic exergetic-based framework was developed to assess the sustainability and productivity of a batch bioreactor used for ethanol production by ethanolic fungus Mucor indicus. Analyses were carried out in order to identify the most exergetically-sustainable concentration of phosphorous compound to produce ethanol and biomass. The microorganisms were aerobically cultivated using glucose as carbon source at various phosphate concentrations ranging from 0.0 to 7.5 g/L. The results obtained showed that the exergetic parameters of the fermentation process were remarkably influenced by the concentration of phosphate. Generally, the findings achieved revealed 3.5 g/L phosphate concentration as the most optimal fermentation condition from the exergetic point of view. Under this condition, the process exergetic efficiency and normalized exergy destruction as decision making parameters were found to be 53.42% and 0.48 kJ/kJ product, respectively. Moreover, the rational and process sustainability indexes for this concentration were determined at 3.92 and 2.15, respectively. The developed framework could be easily transplanted to evaluate the renewability of various lab-scale biofuel production processes to meet the goals laid forth for sustainable development.

Performance of hydrothermal liquefaction (HTL) of biomass by multivariate data analysis

Hydrothermal liquefaction (HTL) is one of the most promising biomass reforming processes for production of drop-in biofuels. The technique has been under development for a number of years, and yet, due to its complexity, it has always been difficult to generalize information about the optimal conditions. The main issue regards to the limited knowledge available from batch studies evaluating HTL by a finite number of process conditions in certain combinations. In this study, multivariate statistical methods were applied for investigation of HTL data available in the literature. The aim was to determine a set of generally valid rules for prediction of the output from the process (yields and the energy content) on the basis of relatively few process parameters. The results have shown that multivariate data analysis can be used to make predictions about HTL and increase our understanding of the process, despite the fact that the input data constituted a very broad spectrum of values. In general, biomass type and properties were the most significant parameters controlling both the obtained yields and the energy content in the produced biocrude. Regression models calculated for all groups of biomass were relatively poor, due to the lack of common trends. However, a number of statistically sound models were obtained for selected combinations of biomass and responses. The drawn conclusions not only supported the pre-understood axioms of HTL, but also indicated a number of new associations. It was shown that the overall conversion rates are governed by biomass properties and the applied heating velocities, while the amount of homogeneous catalyst and the reaction time control the distribution of the products between the water phase and the biocrude. The energy content in the biocrude produced from lignocellulose was dependent mostly on the biomass content and properties, and not the process conditions.

Lifecycle impacts of ethanol production from spruce wood chips under high-gravity conditions.

BackgroundDevelopment of more sustainable biofuel production processes is ongoing, and technology to run these processes at a high dry matter content, also called high-gravity conditions, is one option. This paper presents the results of a life cycle assessment (LCA) of such a technology currently in development for the production of bio-ethanol from spruce wood chips.ResultsThe cradle-to-gate LCA used lab results from a set of 30 experiments (or process configurations) in which the main process variable was the detoxification strategy applied to the pretreated feedstock material. The results of the assessment show that a process configuration, in which washing of the pretreated slurry is the detoxification strategy, leads to the lowest environmental impact of the process. Enzyme production and use are the main contributors to the environmental impact in all process configurations, and strategies to significantly reduce this contribution are enzyme recycling and on-site enzyme production. Furthermore, a strong linear correlation between the ethanol yield of a configuration and its environmental impact is demonstrated, and the selected environmental impacts show a very strong cross-correlation (r^2>0.9 in all cases) which may be used to reduce the number of impact categories considered from four to one (in this case, global warming potential). Lastly, a comparison with results of an LCA of ethanol production under high-gravity conditions using wheat straw shows that the environmental performance does not significantly differ when using spruce wood chips. For this comparison, it is shown that eutrophication potential also needs to be considered due to the fertilizer use in wheat cultivation.ConclusionsThe LCA points out the environmental hotspots in the ethanol production process, and thus provides input to the further development of the high-gravity technology. Reducing the number of impact categories based only on cross-correlations should be done with caution. Knowledge of the analyzed system provides further input to the choice of impact categories.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0468-3) contains supplementary material, which is available to authorized users.

Accounting for all sugars produced during integrated production of ethanol from lignocellulosic biomass

Accurate mass balance and conversion data from integrated operation is needed to fully elucidate the economics of biofuel production processes. This study explored integrated conversion of corn stover to ethanol and highlights techniques for accurate yield calculations. Acid pretreated corn stover (PCS) produced in a pilot-scale reactor was enzymatically hydrolyzed and the resulting sugars were fermented to ethanol by the glucose–xylose fermenting bacteria, Zymomonas mobilis 8b. The calculations presented here account for high solids operation and oligomeric sugars produced during pretreatment, enzymatic hydrolysis, and fermentation, which, if not accounted for, leads to overestimating ethanol yields. The calculations are illustrated for enzymatic hydrolysis and fermentation of PCS at 17.5% and 20.0% total solids achieving 80.1% and 77.9% conversion of cellulose and xylan to ethanol and ethanol titers of 63g/L and 69g/L, respectively. These procedures will be employed in the future and the resulting information used for techno-economic analysis.

Using global transcription machinery engineering (gTME) to improve ethanol tolerance of Zymomonas mobilis.

BackgroundWith the increasing global crude oil crisis and resulting environmental concerns, the production of biofuels from renewable resources has become increasingly important. One of the major challenges faced during the process of biofuel production is the low tolerance of the microbial host towards increasing biofuel concentrations.ResultsHere, we demonstrate that the ethanol tolerance of Zymomonas mobilis can be greatly enhanced through the random mutagenesis of global transcription factor RpoD protein, (σ70). Using an enrichment screening, four mutants with elevated ethanol tolerance were isolated from error-prone PCR libraries. All mutants showed significant growth improvement in the presence of ethanol stress when compared to the control strain. After an ethanol (9 %) stress exposure lasting 22 h, the rate of glucose consumption was approximately 1.77, 1.78 and 1.39 g L−1 h−1 in the best ethanol-tolerant strain ZM4-mrpoD4, its rebuilt mutant strain ZM4-imrpoD and the control strain, respectively. Our results indicated that both ZM4-mrpoD4 and ZM4-imrpoD consumed glucose at a faster rate after the initial 9 % (v/v) ethanol stress, as nearly 0.64 % of the initial glucose remained after 54 h incubation versus approximately 5.43 % for the control strain. At 9 % ethanol stress, the net ethanol productions by ZM4-mrpoD4 and ZM4-imrpoD during the 30–54 h were 13.0–14.1 g/l versus only 6.6–7.7 g/l for the control strain. The pyruvate decarboxylase activity of ZM4-mrpoD4 was 62.23 and 68.42 U/g at 24 and 48 h, respectively, which were 2.6 and 1.6 times higher than the control strain. After 24 and 48 h of 9 % ethanol stress, the alcohol dehydrogenase activities of ZM4-mrpoD4 were also augmented, showing an approximate 1.4 and 1.3 times increase, respectively, when compared to the control strain. Subsequent quantitative real-time PCR analysis under these stress conditions revealed that the relative expression of pdc in cultured (6 and 24 h) ZM4-mrpoD4 increased by 9.0- and 12.7-fold when compared to control strain.ConclusionsCollectively, these results demonstrate that the RpoD mutation can enhance ethanol tolerance in Z. mobilis. Our results also suggested that RpoD may play an important role in resisting high ethanol concentration in Z. mobilis and manipulating RpoD via global transcription machinery engineering (gTME) can provide an alternative and useful approach for strain improvement for complex phenotypes.