Pretreatment is required to alter the physical and chemical properties of the biomass to make it easier to process. The methods of pretreatment are similar for either enzy­matic or microbial cellulosic ethanol processing. Removing or altering the lignin allows access to carbohydrates in the biomass. Higher lignin sources require chemi­cal treatment to reduce the level to below 12% to enhance digestibility [50]. To gain access to the cellulose fiber, de-crystallization of the hemicellulose that is cova­lently bound with the lignin via hydrolysis is required [52]. The conversion of all the sugars derived from hemicellulose is highly desired to increase efficiency and minimize by-products. Pretreatment of the biomass is also required to increase the surface area and pore size, thus making it easier to digest. The increase in surface area is from the combination of hemicellulose solubilization, lignin solubilization, and lignin redistribution caused by various methods of pretreatment [53].

There are several methods by which pretreatment is performed: physical, chem­ical, and biological. Physical methods include ball and compression milling that shear or shed the biomass to de-crystallize the cellulose and increase the surface area and digestibility. However, these processes do very little to degrade hemicellulose and lignin polymers. Milling also requires long processing times with high capital and operating costs, thus it is not economical and has not been pursued in scale-up operations [50, 54]. Radiation pretreatment utilizes gamma rays, electron beams, or microwaves to react to weaken and break the chemical bonds between hemicellu — lose and lignin through chemical reactions such as chain scission [55]. However, the high consumption of energy and capital costs makes this process economically unviable.

Dilute-acid pretreatment is a chemical process that increases the solubility of hemicellulose to 80-100%, extensively redistributes the lignin, and depolymerizes some of the cellulose [53]. The process soaks the biomass in a dilute solution of sulfuric, hydrochloric, or nitric acid and then raises the temperature by injecting steam to enhance the pretreatment method [50]. Autohydrolysis generates acids by the introduction of saturated steams into the biomass to breakdown the hemicellu — lose and lignin [50]. The pressure is rapidly released resulting in the breakup of the biomass due to the instant vaporization of the trapped water. This process is known as steam explosion pretreatment and results in 80-100% solubilization into a mix­ture of monomers and oligomers of hemicellulose. It also redistributes the lignin, and depolymerizes some of the cellulose [53]. Similar to steam explosion, ammonia fiber explosion pretreatment (AFEX) uses high temperature and pressure ammonia to de-crystallize cellulose, and increase the solubility of lignin by 10-20%, and of hemicellulose up to 60% while hydrolyzing about 90% to oligomers [53].

Other chemical pretreatment methods include “hydrothermal” processes using liquid hot water, supercritical carbon dioxide, “organosolv” processes that involve organic solvents in an aqueous medium, concentrated phosphoric or peracetic acid treatment, and strong alkali processes using sodium hydroxide or lime [50, 53]. A biological pretreatment process utilizes fungi, such as white rot, brown rot, and soft rot, to hydrolyze the cellulose component of biomass. Filamentous fungi, typically Trichoderma and Penicillium species, can be used directly for cellulose hydroly­sis because of the greater capacity for extracellular protein production than that of cellulolytic bacteria [56]. However, it requires a three-fold reduction in cost for com­mercialization and the reaction rates for the hydrolysis of cellulose are relatively low in comparison to chemical pretreatment methods [56].

Enzymatic saccharification utilizes enzyme blends for recovering carbohydrates from the hydrolyzate generated after pretreatment [51]. Commonly, cellulase and hemicellulase enzymes are used as a “cocktail” with other enzymes to enhance yields and reduce enzyme costs. The products of enzymatic saccharification — the process of breaking a complex carbohydrate into its monosaccharide components — severely inhibit cellulases and hemicellulases [57]. To overcome this difficulty, Simultaneous Saccharification and Fermentation (SSF) of the pretreated hydrolyzate is preferred. Once the structure of the biomass is disrupted, the cellulose and hemicellulose is enzymatically converted to sugars by the saccharification process. During the fermentation process, yeasts such as Saccharomyces cerevisiae, con­vert the sugars to ethanol. The advantage of SSF over Separate Hydrolysis and Fermentation (SHF) is higher yields of ethanol but SSF requires more than dou­ble the fermentation time [58]. However, the hydrolyzate also contains acetic acid and other toxic compounds. Together with increasing ethanol concentrations, this can inhibit the enzymes and fermentation organisms, thus lowering yields. New developments in enzymatic saccharification and fermentation have been developed by Iogen Energy Corporation and the NREL to develop effective “cocktails” of enzymes along with modified strains of yeast that can break down complex sugar molecules, which conventional fermentation yeast cannot.

Recently, Royal Dutch Shell (Shell) announced a partnership with Iogen Energy Corporation to advance cellulosic ethanol from agriculture residues such as cereal straw and corn cobs and stalks. And just recently, Iogen Energy shipped 100,000 L (26,417 gal) to Shell, which is the first installment of the initial order of 180,000 L (47,550 gal) of cellulosic ethanol. Iogen’s demonstration facility located in Ottawa, which first began producing cellulosic ethanol in 2004, is being purchased by Shell for use in upcoming fuel applications [59].

Cellulolytic microorganisms, an alternative to yeast, utilize ethanol fermenting microbes that both hydrolyze and ferment the sugars into ethanol from a milder pretreatment process. Gram-negative bacteria, such as Escherichia coli, Klebsiella oxytoca, and Zymomonas mobilis, are being investigated as potential microor­ganisms for industrial production of ethanol [52]. Using genetic and metabolic engineering, NREL has developed a strain of Z. mobilis (Zymo) that can break down complex sugars like xylose, and tolerate higher concentrations of acetic acid [51]. Other studies have shown that the Z. mobilis strain can produce theoretical yields up to 95% and handle a wider range of feedstocks [52]. High technological costs have impeded the widespread production of cellulosic ethanol by microor­ganisms. Consolidated bio-processing or CBP has been developed to address this problem. This process utilizes cellulolytic microorganisms to perform the hydroly­sis of biomass and the fermentation of sugars into ethanol within a single process, which is a large cost reducing strategy [53]. CBP is expected to reduce overall production costs by eight-fold compared to SSF under similar conditions.

Mascoma Corporation has dedicated their research team to focus on the com­mercialization of CBP, which is seen as the lowest cost configuration for cellulosic ethanol. Mascoma Corporation is in the process of developing a cellulosic fuel pro­duction facility that will use non-food biomass to convert woodchips into fuel. They are predicting that the new facility will produce 40 million gallons of ethanol and other valuable fuel products per year [60].

3.4 Summary

Cellulosic ethanol is ethyl alcohol produced from wood, grass, or the non-edible parts of plants, and is a sustainable and renewable biofuel that is biodegradable. The promising features of cellulosic ethanol are the diverse and abundant feedstock that can utilize existing waste by-products. Iogen Energy Corporation is currently producing cellulosic ethanol for Shell using enzymatic saccharification and fermen­tation in a small-scale commercial facility. Another approach to cellulosic ethanol is via the use of cellulolytic microorganisms. As commercialization of cellulosic ethanol expands, it can be used to increase ethanol production without causing food shortages or demands, and will reduce greenhouse gas emissions and our dependence on fossil fuels.