Feasibility of using biochar as buffer and mineral nutrients replacement for acetone-butanol-ethanol production from non-detoxified switchgrass hydrolysate.

Biochar can be an inexpensive pH buffer and source of mineral and trace metal nutrients in acetone-butanol-ethanol (ABE) fermentation. This study evaluated the feasibility of replacing expensive 4-morpholineethanesulfonic acid (MES) P2 buffer and mineral nutrients with biochar made from switchgrass (SGBC), forage sorghum (FSBC), redcedar (RCBC) and poultry litter (PLBC) for ABE fermentation. Fermentations using Clostridium beijerinckii ATCC 51743 in glucose and non-detoxified switchgrass hydrolysate media were performed at 35 °C in 250 mL bottles for 72 h. Medium containing buffer and minerals without biochar was the control. Similar ABE production (about 18.0 g/L) in glucose media with SGBC, FSBC and RCBC and control was measured. However in non-detoxified switchgrass hydrolysate medium, SGBC, RCBC and PLBC produced more ABE (about 18.5 g/L) than the control (10.1 g/L). This demonstrates that biochar is an effective buffer and mineral supplement for ABE production from lignocellulosic biomass without costly detoxification process.

Catalytic co-pyrolysis of red cedar with methane to produce upgraded bio-oil.

Fast pyrolysis is a promising route to transform biomass into bio-oil for further refining into transportation fuels and chemicals. However, bio-oil applications suffer from several challenges due to its adverse properties. This study reports improving bio-oil properties through co-pyrolysis of biomass with methane over molybdenum/zinc (MoZn/ZHSM-5) and HZSM-5 catalysts that promote deoxygenation, decarbonylation, hydrogen transfer and aromatization reactions. The co-pyrolysis was conducted at 650 °C and 750 °C in a micro-scale reactor and a bench-scale reactor. The highest bio-oil yield, energy content, and energy yield of 53.4%, 10.2 MJ/kg, and 29.9%, respectively, were obtained with methane over MoZn/HZSM-5 at 650 °C. Acids, alcohols, aldehydes, benzene derivatives, BTEXs, furans, ketones, PAHs, and phenols were detected in bio-oils while phenols dominated under most conditions. Oxygenated compounds decreased using MoZn/HZSM-5 with methane at 750 °C. The results demonstrate that methane used with catalysts can reduce oxygenated compounds and improve properties and yield of bio-oil.

Ethanol production during semi-continuous syngas fermentation in a trickle bed reactor using Clostridium ragsdalei.

An efficient syngas fermentation bioreactor provides a mass transfer capability that matches the intrinsic kinetics of the microorganism to obtain high gas conversion efficiency and productivity. In this study, mass transfer and gas utilization efficiencies of a trickle bed reactor during syngas fermentation by Clostridium ragsdalei were evaluated at various gas and liquid flow rates. Fermentations were performed using a syngas mixture of 38% CO, 28.5% CO2, 28.5% H2 and 5% N2, by volume. Results showed that increasing the gas flow rate from 2.3 to 4.6sccm increased the CO uptake rate by 76% and decreased the H2 uptake rate by 51% up to Run R6. Biofilm formation after R6 increased cells activity with over threefold increase in H2 uptake rate. At 1662h, the final ethanol and acetic acid concentrations were 5.7 and 12.3g/L, respectively, at 200ml/min of liquid flow rate and 4.6sccm gas flow rate.

Butanol and hexanol production in Clostridium carboxidivorans syngas fermentation: Medium development and culture techniques.

Clostridium carboxidivorans was grown on model syngas (CO:H2:CO2 [70:20:10]) in a defined nutrient medium with concentrations of nitrogen, phosphate and trace metals formulated to enhance production of higher alcohols. C. carboxidivorans was successfully grown in a limited defined medium (no yeast extract, no MES buffer and minimal complex chemical inputs) using an improved fermentation protocol. Low partial pressure of CO in the headspace, coupled with restricted mass transfer for CO and H2, was required for successful fermentation. In the absence of substrate inhibition (particularly from CO), growth limitation increased production of alcohols, especially butanol and hexanol. Concentrations of butanol (over 1.0g/L), hexanol (up to 1.0g/L) and ethanol (over 3.0g/L) were achieved in bottle fermentations. Minimal medium and controlled supply of CO and H2 should be used in characterizing candidate butanol and hexanol producing strains to select for commercial potential.

Effects of torrefaction and densification on switchgrass pyrolysis products.

The pyrolysis behaviors of four types of pretreated switchgrass (torrefied at 230 and 270°C, densification, and torrefaction at 270°C followed by densification) were studied at three temperatures (500, 600, 700°C) using a pyroprobe attached to a gas chromatogram mass spectroscopy (Py-GC/MS). The torrefaction of switchgrass improved its oxygen to carbon ratio and energy content. Contents of anhydrous sugars and phenols in pyrolysis products of torrefied switchgrass were higher than those in pyrolysis products of raw switchgrass. As the torrefaction temperature increased from 230 to 270°C, the contents of anhydrous sugars and phenols in pyrolysis products increased whereas content of guaiacols decreased. High pyrolysis temperature (600 and 700°C as compared to 500°C) enhanced decomposition of lignin and anhydrous sugars, leading to increase in phenols, aromatics and furans. Densification enhanced depolymerization of cellulose and hemicellulose during pyrolysis.

Continuous syngas fermentation for the production of ethanol, n-propanol and n-butanol.

Syngas fermentation to fuels is a technology on the verge of commercialization. Low cost of fermentation medium is important for process feasibility. The use of corn steep liquor (CSL) instead of yeast extract (YE) in Alkalibaculum bacchi strain CP15 bottle fermentations reduced the medium cost by 27% and produced 78% more ethanol. When continuous fermentation was performed in a 7-L fermentor, 6g/L ethanol was obtained in the YE and YE-free media. When CSL medium was used in continuous fermentation, the maximum produced concentrations of ethanol, n-propanol and n-butanol were 8 g/L, 6 g/L and 1 g/L, respectively. n-Propanol and n-butanol were not typical products of strain CP15. A 16S rRNA gene-based survey revealed a mixed culture in the fermentor dominated by A. bacchi strain CP15 (56%) and Clostridium propionicum (34%). The mixed culture presents an opportunity for higher alcohols production from syngas.

Mixed culture syngas fermentation and conversion of carboxylic acids into alcohols.

Higher alcohols such as n-butanol and n-hexanol have higher energy density than ethanol, are more compatible with current fuel infrastructure, and can be upgraded to jet and diesel fuels. Several organisms are known to convert syngas to ethanol, but very few can produce higher alcohols alone. As a potential solution, mixed culture fermentation between the syngas fermenting Alkalibaculum bacchi strain CP15 and propionic acid producer Clostridium propionicum was studied. The monoculture of CP15 produced only ethanol from syngas without initial addition of organic acids to the fermentation medium. However, the mixed culture produced ethanol, n-propanol and n-butanol from syngas. The addition of propionic acid, butyric acid and hexanoic acid to the mixed culture resulted in a 50% higher conversion efficiency of these acids to their respective alcohols compared to CP15 monoculture. These findings illustrate the great potential of mixed culture syngas fermentation in production of higher alcohols.

Scenario optimization modeling approach for design and management of biomass-to-biorefinery supply chain system.

The aim of this study was to develop a scenario optimization model to address weather uncertainty in the Biomass Supply Chain (BSC). The modeling objective was to minimize the cost of biomass supply to biorefineries over a one-year planning period using monthly time intervals under different weather scenarios. The model is capable of making strategic, tactical and operational decisions related to BSC system. The performance of the model was demonstrated through a case study developed for Abengoa biorefinery in Kansas. Sensitivity analysis was done to demonstrate the effect of input uncertainty in yield, land rent and storage dry matter loss on the model outputs. The model results show that available harvest work hours influence major cost-related decisions in the BSC.

Development of low cost medium for ethanol production from syngas by Clostridium ragsdalei.

The development of a low cost medium for ethanol production is critical for process feasibility. Ten media were formulated for Clostridium ragsdalei by reduction, elimination and replacement of expensive nutrients. Cost analysis and effects of medium components on growth and product formation were investigated. Fermentations were performed in 250 mL bottles using syngas (20% CO, 15% CO2, 5% H2 and 60% N2). The standard medium M1 cost is $9.83/L, of which 93% is attributed to morpholinoethane sulfonic acid (MES) buffer. Statistical analysis of the results showed that MES removal did not affect cell growth and ethanol production (P>0.05). Based on cells' elemental composition, a minimal mineral concentration medium M7 was formulated, which provided 29% higher ethanol yield from CO at 3% of the cost compared to medium M1. Ethanol yield from CO in the completely defined medium M9 was 36% higher than while at 5% the cost of medium M1.

A comparison of mass transfer coefficients between trickle-bed, hollow fiber membrane and stirred tank reactors.

Trickle-bed reactor (TBR), hollow fiber membrane reactor (HFR) and stirred tank reactor (STR) can be used in fermentation of sparingly soluble gasses such as CO and H2 to produce biofuels and bio-based chemicals. Gas fermenting reactors must provide high mass transfer capabilities that match the kinetic requirements of the microorganisms used. The present study compared the volumetric mass transfer coefficient (K(tot)A/V(L)) of three reactor types; the TBR with 3 mm and 6 mm beads, five different modules of HFRs, and the STR. The analysis was performed using O2 as the gaseous mass transfer agent. The non-porous polydimethylsiloxane (PDMS) HFR provided the highest K(tot)A/V(L) (1062 h(-1)), followed by the TBR with 6mm beads (421 h(-1)), and then the STR (114 h(-1)). The mass transfer characteristics in each reactor were affected by agitation speed, and gas and liquid flow rates. Furthermore, issues regarding the comparison of mass transfer coefficients are discussed.

Effects of cellulose, hemicellulose and lignin on thermochemical conversion characteristics of the selected biomass.

The objective of this study was to investigate effects of biomass constituents (cellulose, hemicellulose and lignin) on biomass thermal decomposition and gas evolution profiles of four biomass materials. Switchgrass, wheat straw, eastern redcedar and dry distilled grains with solubles (DDGS) were selected as the biomass materials. No significant difference was observed in the weight loss profiles of switchgrass, wheat straw and eastern redcedar even though their cellulose, hemicellulose and lignin contents were considerably different. The weight loss kinetic parameters were also not significantly different except for activation energy of the eastern redcedar. However, biomass composition did significantly affect gas evolution profiles. The higher contents of cellulose and hemicellulose in switchgrass and wheat straw may have resulted in their higher CO and CO(2) concentrations as compared to eastern redcedar. On the other hand, higher lignin content in eastern redcedar may have resulted in significantly its high CH(4) concentration.

Fermentative production of ethanol from syngas using novel moderately alkaliphilic strains of Alkalibaculum bacchi.

Ethanol production from syngas using three moderately alkaliphilic strains of a novel genus and species Alkalibaculum bacchi CP11(T), CP13 and CP15 was investigated in 250 ml bottle fermentations containing 100ml of yeast extract medium at 37 °C and pH 8.0. Two commercial syngas mixtures (Syngas I: 20% CO, 15% CO(2), 5% H(2), 60% N(2)) and (Syngas II: 40% CO, 30% CO(2), 30% H(2)) were used. Syngas I and Syngas II represent gasified biomass and coal, respectively. The maximum ethanol concentration (1.7 g l(-1)) and yield from CO (76%) were obtained with strain CP15 and Syngas II after 360 h. CP15 produced over twofold more ethanol with Syngas I compared to strains CP11(T) and CP13. In addition, CP15 produced 18% and 71% more ethanol using Syngas II compared to strains CP11(T) and CP13, respectively. These results show that CP15 is the most promising for ethanol production because of its higher growth and ethanol production rates and yield compared to CP11(T) and CP13.

Reduction of acetone to isopropanol using producer gas fermenting microbes.

Gasification-fermentation is an emerging technology for the conversion of lignocellulosic materials into biofuels and specialty chemicals. For effective utilization of producer gas by fermenting bacteria, tar compounds produced in the gasification process are often removed by wet scrubbing techniques using acetone. In a preliminary study using biomass generated producer gas scrubbed with acetone, an accumulation of acetone and subsequent isopropanol production was observed. The effect of 2 g/L acetone concentrations in the fermentation media on growth and product distributions was studied with "Clostridium ragsdalei," also known as Clostridium strain P11 or P11, and Clostridium carboxidivorans P7 or P7. The reduction of acetone to isopropanol was possible with "C. ragsdalei," but not with P7. In P11 this reaction occurred rapidly when acetone was added in the acidogenic phase, but was 2.5 times slower when added in the solventogenic phase. Acetone at concentrations of 2 g/L did not affect the growth of P7, but ethanol increased by 41% and acetic acid concentrations decreased by 79%. In the fermentations using P11, growth was unaffected and ethanol concentrations increased by 55% when acetone was added in the acidogenic phase. Acetic acid concentrations increased by 19% in both the treatments where acetone was added. Our observations indicate that P11 has a secondary alcohol dehydrogenase that enables it to reduce acetone to isopropanol, while P7 lacks this enzyme. P11 offers an opportunity for biological production of isopropanol from acetone reduction in the presence of gaseous substrates (CO, CO2, and H2).

Effect of nutrient limitation and two-stage continuous fermentor design on productivities during "Clostridium ragsdalei" syngas fermentation.

The effect of three limiting nutrients, calcium pantothenate, vitamin B(12) and cobalt chloride (CoCl(2)), on syngas fermentation using "Clostridium ragsdalei" was determined using serum bottle fermentation studies. Significant results from the bottle studies were translated into single- and two-stage continuous fermentor designs. Studies indicated that three-way interactions between the three limiting nutrients, and two-way interactions between vitamin B(12) and CoCl(2) had a significant positive effect on ethanol and acetic acid formation. In general, ethanol and acetic acid production ceased at the end of 9 days corresponding to the production of 2.01 and 1.95 gL(-1) for the above interactions. Reactor studies indicated the three-way nutrient limitation in two-stage fermentor showed improved acetic acid (17.51 gg(-1) cells) and ethanol (14.74 gg(-1) cells) yield compared to treatments in single-stage fermentors. These results further support the hypothesis that it is possible to modulate the product formation by limiting key nutrients during C. ragsdalei syngas fermentation.

Ethanol production from syngas by Clostridium strain P11 using corn steep liquor as a nutrient replacement to yeast extract.

The feasibility of replacing yeast extract (YE) by corn steep liquor (CSL), a low cost nutrient source, for syngas fermentation to produce ethanol using Clostridium strain P11 was investigated. About 32% more ethanol (1.7 g L(-1)) was produced with 20 g L(-1) CSL media in 250-mL bottle fermentations compared to media with 1 g L(-1) YE after 360 h. Maximum ethanol concentrations after 360 h of fermentation in a 7.5-L fermentor with 10 and 20 g L(-1) CSL media were 8.6 and 9.6 g L(-1), respectively, which represent 57% and 60% of the theoretical ethanol yields from CO. Only about 6.1 g L(-1) of ethanol was obtained in the medium with 1 g L(-1) YE after 360 h, which represents 53% of the theoretical ethanol yield from CO. The use of CSL also enhanced butanol production by sevenfold compared to YE in bottle fermentations. These results demonstrate that CSL can replace YE as the primary medium component and significantly enhance ethanol production by Clostridium strain P11.

Effect of temperature, pH and buffer presence on ethanol production from synthesis gas by "Clostridium ragsdalei".

Fermentation pH, incubation temperature, and presence or absence of media buffer can alter the activity of microorganisms. For instance, carbon monoxide and hydrogen components of syngas show decreased solubility with increasing temperature, Clostridium species preferentially switch from acetogenesis to solventogenesis phase at pH below 5.0, and morpholinoethanesulfonic acid (MES) added as media buffer has been shown to increase lag time for ethanol production. The objective of the present study was to determine the effects of temperature, pH and MES buffer on ethanol production by "Clostridium ragsdalei". This study showed syngas fermentation using "Clostridium ragsdalei" at 32°C with media without buffer was associated with higher ethanol concentration and reduced lag time in switching to solventogenesis. Temperature above 40°C and pH below 5.0 were outside the optimal range for growth and metabolism of the bacteria.

Sequencing of multiple clostridial genomes related to biomass conversion and biofuel production.

Modern methods to develop microbe-based biomass conversion processes require a system-level understanding of the microbes involved. Clostridium species have long been recognized as ideal candidates for processes involving biomass conversion and production of various biofuels and other industrial products. To expand the knowledge base for clostridial species relevant to current biofuel production efforts, we have sequenced the genomes of 20 species spanning multiple genera. The majority of species sequenced fall within the class III cellulosome-encoding Clostridium and the class V saccharolytic Thermoanaerobacteraceae. Species were chosen based on representation in the experimental literature as model organisms, ability to degrade cellulosic biomass either by free enzymes or by cellulosomes, ability to rapidly ferment hexose and pentose sugars to ethanol, and ability to ferment synthesis gas to ethanol. The sequenced strains significantly increase the number of noncommensal/nonpathogenic clostridial species and provide a key foundation for future studies of biomass conversion, cellulosome composition, and clostridial systems biology.

Feasibility of incorporating cotton seed extract in Clostridium strain P11 fermentation medium during synthesis gas fermentation.

Biomass gasification followed by fermentation of syngas to ethanol is a potential process to produce bioenergy. To make this process more economical, the complexity of media should be reduced while using less costly components. In this study, the feasibility of incorporating cotton seed extract (CSE) as a media component for syngas fermentation to produce ethanol using Clostridium strain P11 was evaluated. A factorial experiment was conducted to screen and evaluate the effect of different media components, in relation to CSE, on ethanol production. Also, different CSE concentrations as well as the presence of MES buffer were tested to determine their effect on ethanol production. Bottle fermentations with media containing only 1.0 gL(-1) CSE produced more ethanol after 15 d (1.17 gL(-1)) than fermentation using any other media. Further bottle experiments showed that media containing only 0.5 gL(-1) CSE produced more ethanol after 15 days (2.67 gL(-1)) than a control media (0.6 gL(-1)) and media containing only 1.0 gL(-1) CSE (2.16 gL(-1)). Fermentations in 5- and 7.5-L stirred fermentors with 0.5 gL(-1) CSE media achieved ethanol concentrations similar to those observed in bottle studies. These results indicate that CSE can replace all the vitamin and mineral media components generally used for fermentation of syngas to ethanol by Clostridium strain P11, thereby improving the process economics.

Syngas fermentation in a 100-L pilot scale fermentor: design and process considerations.

Fermentation of syngas offers several advantages compared to chemical catalysts such as higher specificity of biocatalysts, lower energy costs, and higher carbon efficiency. Scale-up of syngas fermentation from a bench scale to a pilot scale fermentor is a critical step leading to commercialization. The primary objective of this research was to install and commission a pilot scale fermentor, and subsequently scale-up the Clostridium strain P11 fermentation from a 7.5-L fermentor to a pilot scale 100-L fermentor. Initial preparation and fermentations were conducted in strictly anaerobic conditions. The fermentation system was maintained in a batch mode with continuous syngas supply. The effect of anaerobic fermentation in a pilot scale fermentor was evaluated. In addition, the impact of improving the syngas mass transfer coefficient on the utilization and product formation was studied. Results indicate a six fold improvement in ethanol concentration compared to serum bottle fermentation, and formation of other compounds such as isopropyl alcohol, acetic acid and butanol, which are of commercial importance.

Alkalibaculum bacchi gen. nov., sp. nov., a CO-oxidizing, ethanol-producing acetogen isolated from livestock-impacted soil.

Phenotypic and phylogenetic studies were performed on three strains of an acetogenic bacterium isolated from livestock-impacted soil. The bacterium stained Gram-negative and was a non-spore-forming rod that was motile by peritrichous flagella. The novel strains had an optimum pH for growth of 8.0-8.5 and utilized H2 : CO2, CO : CO2, glucose, fructose, mannose, turanose, ribose, trimethylamine, pyruvate, methanol, ethanol, n-propanol and n-butanol as growth substrates. Acetate was produced from glucose. Acetate, CO2 and ethanol were produced from CO : CO2. 16S rRNA gene sequence analysis indicated that the novel strains formed a new subline in the family Eubacteriaceae (rRNA cluster XV) of the low G+C-containing Gram-positive bacteria of the class Clostridia. The DNA G+C base composition was 34 mol%. Cell wall analysis revealed the existence of a novel B-type peptidoglycan similar to the B2α-type (B4) configuration with a variation containing aspartic acid. Based on phylogenetic and phenotypic evidence, it is proposed that the new isolates represent a novel genus and species, for which the name Alkalibaculum bacchi gen. nov., sp. nov. is proposed. The type strain of the type species is CP11(T) (=ATCC BAA-1772(T)=DSM 22112(T)).