BIOFUELS SUPPLEMENT
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A bi-weekly summary of world developments on biofuels, produced by the Global Knowledge Center on Crop Biotechnology, International Service for the Acquisition of Agri-biotech Applications SEAsiaCenter (ISAAA)
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November 18, 2011

In This Issue:

News and Trends
- NMR Spectroscopy for Assessing Pretreatment Effects of Switchgrass 
- "Chemostat Evolution" Used to Develop an Ethanol-fermenting Yeast in Lignocellulosic Hydrolyzates at High Solids Loading 

Energy Crops and Feedstocks for Biofuels Production
- Direct Analysis and Profiling of Intact Triglycerides in Microalgal Oil by UHPLC-MS 

Biofuels Processing
- Review of Biomass Pretreatment Methods for Biofuel Production: "Fundamentals Toward Application" 

Biofuels Policy and Economics
- Life Cycle Assessment of Biofuel Production/Use in Argentina Incorporating "Water Footprint" 
- Approach for Assessing Climate Change Mitigation and Fossil Fuel Replacement by Biofuel Technologies 





* NEWS AND TRENDS *

NMR Spectroscopy for Assessing Pretreatment Effects of Switchgrass
(complete access to journal article may require subscription or payment)
http://www.sciencedirect.com/science/article/pii/S0141391011002849

Researchers from the University of Georgia report the use of nuclear magnetic resonance spectroscopy (NMR)to assess the chemical changes in switchgrass (Panicum virgatum, a potential biofuel [grass] feedstock) after subjecting it to different types of pretreatment methods. (Pretreatment by physicochemical or biological methods "deconstructs" the plant cell wall; this is the first step in biofuel ethanol production from lignocellulosics). NMR is a technique in chemistry which relies on nuclear magnetic resonance properties of certain atoms for the structural analysis of substances, including its chemical environment.

The traditional method of pretreatment assessment of lignocellulosic biomass usually involves time-consuming procedures of extraction/purification prior to spectroscopic characterization. In the NMR method, such time-consuming procedures are reported to be not necessary. It involves dissolving the lignocellulosic sample in a binary solvent system consisting of "1-methyl imidazole-d6/DMSO-d6". High resolution NMR spectra data can be reportedly obtained from this solvent system. The NMR spectral data allows the identification of lignin sub-units, as well as polysaccharide units in pretreated switchgrass. The methods used for the pretreatment of switchgrass include: steam explosion, lime addition, and acid pretreatment.

The NMR results from the study showed that: (1) xylan is the major hemicellulose fraction of switchgrass, (2) switchgrass lignin is composed of H, G and S monolignol subunits with significant amounts of p-coumarate and ferulate, (3) steam explosion slightly degrades lignin and partially dissolved the hemicellulose in switchgrass, (4) lime pretreatment resulted in "preferential degradation of p-hydroxy benzoyl ester, ferulate and coumarate" in lignin, and less severe dissolution of hemicellulose, (4) acid treatment was the most "harsh" pretreatment"; there was significant degradation of both lignin and hemicellulose. The full paper is published in the journal, Polymer Degradation and Stability (URL above).

Related information:Basics of NMR: http://www.cis.rit.edu/htbooks/nmr/inside.htm

Lignin: http://www.dfrc.wisc.edu/dfrcwebpdfs/jr_brazil99_paper.pdf



"Chemostat Evolution" Used to Develop an Ethanol-fermenting Yeast in Lignocellulosic Hydrolyzates at High Solids Loading
http://www.biotechnologyforbiofuels.com/content/4/1/49/abstract

Scientists from the Microbiology Department, University of Georgia (United States) report the use of "chemostat evolution" for the development of a Saccharomyces cerevisiae strain which can ferment lignocellulosic-hydrolyzates at high solids loading, even in the presence of toxic levels of inhibitory compounds. Lignocellulosic-hydrolyzate is the fermentable material after the pretreatment of lignocellulosic biomass (to "deconstruct" the plant cell walls) and subsequent saccharification (to convert carbohydrate polymers in the deconstructed plant cell walls, into simple sugars).

Saccharomyces cerevisiae is the common microorganism (a type of yeast) for the fermentation of lignocellulosic-hydrolyzates into ethanol, but these organisms do not perform well at high solids loading, and are often inhibited by pretreatment by-products (substances such as fufural and acetic acid, which are present in lignocellulosic-hydrolyzates). The scientists attempted to developer an improved strain of Saccharomyces cerevisiae by subjecting the yeast to selective pressure (i.e.,lignocellulosic (softwood pine) hydrolyzate at high solids loading (17.5% w/v solids) with inhibitory compounds) in continuous culture (technically called a "chemostat").

The chemostat was repeatedly operated for many cycles. During the long operation at many cycles, the yeasts are considered to "adaptively evolve" under these conditions of selective pressure. The scientists reported that they obtained an evolved strain of Saccharomyces cerevisiae (AJP50), which possessed "greater fermentation capability than its parent in both rich liquid media supplemented with various combinations of inhibitory compounds, and in pretreated pine fermentations at high solids loadings." The full paper can be accessed in the open-access journal, Biotechnology for Biofuels (URL above).




* ENERGY CROPS AND FEEDSTOCKS FOR BIOFUELS PRODUCTION *

Direct Analysis and Profiling of Intact Triglycerides in Microalgal Oil by UHPLC-MS
(complete access to journal article may require subscription or payment)
http://www.springerlink.com/content/6371311377016383/

Oleagenous (oil-bearing) microalgae have been reported to be a potential feedstock for biodiesel production. These microorganisms are usually cultivated in large ponds, then harvested and extracted for their oils (mainly triglycerides). The extracted oils are then made to undergo a "trans-esterification"reaction with methanol in order to obtain a mixture of methyl esters (collectively known as "biodiesel"). The screening of microalgae that can be potentially cultivated as biodiesel feedstocks usually involves a determination of how much oil the algae can produce, and what type of triglycerides are present in the oil.

The common method of triglyceride analysis is to break-down the triglycerides into their component fatty acids and then "derivatize" the fatty acids into their corresponding methyl esters for injection into a gas chromatograph. However, according to the researchers from the National Research Council of Canada, GC (gaschromatographic) detection of fatty acid methyl esters (FAME) in the oil samples "offers fragmentation data of fatty acid identification", and no information is provided regarding the intact lipids. Consequently, "GC analysis could be misleading if relying solely on the fatty acid profile for strain selection in biofuels applications." An alternative method for the direct measurement of intact lipids was attempted using Ultra High Pressure Liquid Chromatography-Mass Spectrometry (UHPLC-MS). They tested the method using six different types of algal strains for possible use in biofuel applications: Botryococcus braunii, Nannochloropsis gaditana, Neochloris oleoabundans, Phaeodactyl umtricornutum, Porphyridium aerugineum,and Scenedesmus obliquus.

The following are some highlights of the study: (1) the procedure allowed the identification of more than 100 unique triglycerides from the six algal strains, (2) the "most comprehensive [triglyceride] profile" with a high abundance of oleic acid were from Botryococcus braunii and Scenedesmusobliquus. The complete paper is published in the journal, Analytical and Bioanalytical Chemistry (URL above).




* BIOFUELS PROCESSING *

Review of Biomass Pretreatment Methods for Biofuel Production: "Fundamentals Toward Application"
(complete access to journal article may require subscription or payment)
http://www.sciencedirect.com/science/article/pii/S0734975011000607

A recent review by Valery Agbor and colleagues from the University of Manitoba (Canada) provides an overview of the methods (and latest trends) being used in the pretreatment of lignocellulosic biomass for the production of "second generation" biofuel-ethanol. Pretreatment is the first of three steps in the conversion of lignocellulosic biomass to ethanol. The next two sequential steps are saccharification and fermentation. The main objective of pretreatment is to "deconstruct" the plant cell walls, by destroying the tight lignin wrapping surrounding the biomass, resulting in the exposure of the carbohydrate polymers (mainly cellulose). The exposed carbohydrate polymers can then be easily converted to simple sugars in the second-stage saccharification step,and these sugars can then be fermented to ethanol. 

The pretreatment technologies can be broadly classified as: (1) physical, (2) chemical, (3) biological, and (4) multiple/combinatorial treatments. Physical pretreatments include: comminution (grinding to particle size) and gamma-ray treatment. Chemical pretreatment methods include the use of acids, alkali, organic acids and ionic liquids. Biological methods involve the use of microorganisms (usually fungi) for the degradation of lignin and hemicellulose. Combinatorial pretreatments (usually physical plus chemical) include: steam(explosion)treatment,  the use of liquid hot water (170oC to 230oC), treatment with dilute acid (less than 4% by weight) plus heat, ammonia fiber/freeze explosion (AFEX), lime plus wet oxidation pretreatment, and organosolve pretreatment. A brief process description is presented for each method, followed by advantages/disadvantages, and the potential for large-scale application.

The review concludes that biomass pretreatment  "remains a key bottleneck in the bioprocessing of lignocellulosics for biofuels and other bioproducts." Also, the diversity of biofuel feedstocks makes it unlikely, that "one method will become the method of choice for all biomass" feedstocks. The full review is published in the journal, Biotechnology Advances.




* BIOFUELS POLICY AND ECONOMICS *

Life Cycle Assessment of Biofuel Production/Use in Argentina Incorporating "Water Footprint"
(complete access to journal article may require subscription or payment)
http://www.springerlink.com/content/d27613wx05436617/

Life Cycle Assessment (LCA) has been used as a tool for assessing sustainability of biofuel feedstocks. Essentially, LCA tracks certain parameters as the bioenergy crop goes through the stages of cultivation, processing and eventual use (combustion) of the biofuel. The analysis usually focuses on the aspects of (1) net greenhouse gas (GHG) emissions and (2) net energy yield. If the net GHG emission is high and the net energy yield (the energy output from the burning of the biofuel minus the energy input to produce the biofuel) is low, the particular biofuel feedstock might have "sustainability issues". The net GHG emission and net energy yield are related to what are known as the ‘carbon footprint" and the "energy footprint", respectively.

There have been calls to also consider the net water consumption (something like a "water footprint") in the LCA of biofuel feedstocks. This is due to the fact that water is a scarce resource in many countries, and it is possible that while net GHG emissions and net energy yields are good for, a particular biofuel feedstock, it may require large amounts of water for bioenergy crop cultivation. Thus, in water-scarce countries, large water consumption may "cancel out" the benefits of a particular biofuel feedstock. Researchers from the Technology and Society Lab (Dübendorf, Switzerland), Institute of Environmental Engineering (Zurich, Switzerland), and the Universidad Tecnologica Nacional (Buenos Aires, Argentina) report the incorporation of water consumption in its comparative life cycle assessment of biodiesel production from irrigated and non-irrigated rapeseed in Argentina.

Among the highlights of their study are: (1) there are no large variations in water consumption in biofuel production chains based on non-irrigated crops, and the water consumption range is said to be at the same level as fossil fuels, (2) "agricultural water consumption dominates the overall results of all irrigated crops", (3) aggregated "Eco-Indicator 99 scores" for biodiesel production from irrigated rapeseed were doubled by water consumption". The full paper is published in the International Journal of Life Cycle Assessment (URL above)

Related information on LCA: http://www.lcafood.dk/LCA/LCA.htm.  



Approach for Assessing Climate Change Mitigation and Fossil Fuel Replacement by Biofuel Technologies
http://www.sciencedirect.com/science/article/pii/S0306261911001711

Scientists from Vienna University of Technology (Austria) report the application of a "techno-economic" approach for assessing the costs of greenhouse gas (GHG) mitigation and fossil fuel replacement of existing and future bioenergy technologies for heat, electricity and transport-fuel production in Austria. Using data specific to Austria, they applied "sensitivity analysis and projections up to 2030 to illustrate the effect of dynamic parameters on specific abatement costs.

The methodology essentially consisted of the following steps: (1) definition of "default reference systems" for each cluster of bioenergy technologies (which include small scale heating systems, large process heat plants, combined heat/power plants (CHP), liquid/gaseous biofuels), (2) compilation of a default set of technology data, representative fuel prices, and other data, (3) calculation of GHG mitigation costs of bioenergy technologies as incremental generation costs per unit decrease in GHG emissions. The comparative cost results indicated that heat generation and combined heat/power (CHP) generation (assuming favorable conditions) are "the most cost-efficient options for reducing GHG emissions and fossil fuel dependence in Austria". The higher quantities of abatement per unit biomass are said to be the core advantage of CHP. The full paper is published in the journal, Applied Energy (URL above).




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