As a low temperature alternative to steam reforming, Aqueous Phase Reforming (APR) has emerged as a valuable means of converting organic compounds of biological origin to value- added chemicals and fuel components. Due to its feature of low temperature operation, the energy required for water and oxygenated hydrocarbon evaporation is eliminated, leading to the notable reduction of overall energy input, which overcomes the evaporation difficulty of some organic compounds with high boiling point required for steam reforming. In order to keep all reactants in the liquid phase at operation temperature (typically ~500 K), certain pressure (typically 15~50 bar) has to be applied to the whole reactor system. Such operation temperature and pressure benefit the happening of water-gas shift reaction, making it possible to produce hydrogen with low amounts of CO in a single reactor. Undesirable organic compound decomposition can also be minimized under such low reaction temperature. Furthermore, the relatively high pressure operation will also favour the downstream gas separation and purification, and even subsequent gas compression, storage, and delivery. This process is exclusively suitable for the biomass derived organic compounds with relatively longer carbon chain such as sorbitol, which has been comprehensively reviewed by the researchers in Dumesic’s group [20]. For smaller organic compounds like ethanol discussed in this chapter, APR process for hydrogen generation is less favourable from the overall energy utilization viewpoint, which is concluded by Tokarev, et al. in their recent publication [21]. Moreover, the relatively high pressure requirement raises the concerns on safety and operation cost. Hydrogen selectivity is another big challenge APR has to face, because H2 and CO2 produced are thermodynamically unstable and methane formation is favourable at such low temperature.

Cassava Bioethanol

Klanarong Sriroth1, Sittichoke Wanlapatit2 and Kuakoon Piyachomkwan2

1Dept. of Biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok 2Cassava and Starch Technology Research Unit, National Center for Genetic Engineering and Biotechnology (BIOTEC)

Thailand

1. Introduction

1.1 Cassava

Cassava (Manihot esculenta Crantz) is a shrubby perennial crop in the Family of Euphorbiaceae. It is also named others, depending upon geographic regions such as yucca in Central America, mandioca or manioca in Brazil, tapioca in India and Malaysia and cassada or cassava in Africa and Southeast Asia. Cassava is mostly cultivated in tropics of Africa, Latin America and Asia, located in the equatorial belt, between 30° north and 30° south. The crop produces edible starch-reserving roots which have long been employed as an important staple food for millions of mankind as well as animal feed. Due to the fact of ease of plantation and low input requirement, cassava is mostly cultivated in marginal land by poor farmers and is sometimes named as the crop of the poor. In these planting areas, cassava plays an essential role not only as food security, but also income generation. In addition to a primary use for direct consumption and animal feed, starch-rich roots are good raw materials for industrial production of commercial tapioca starch, having excellent characteristics of high whiteness, odorless and tasteless and when cooked, yielding high paste viscosity, clarity and stability. The distinct attributes of extracted cassava starch, either as native or modified form, are very attractive for a broad range of food and non-food application including paper, textile, pharmaceutical, building materials and adhesives. Furthermore, cassava starch is extensively utilized for a production of sweeteners and derivatives including glucose syrup, fructose syrup, sugar alcohols (e. g. sorbitol, mannitol), and organic acids (e. g. lactic acid, citric acid). The application of cassava as renewable feedstock is now expanded to biorefinery, i. e. a facility that integrates processes and equipment to produce fuels, power, chemicals and materials from biomass (Fernando et al., 2006). With this regard, cassava is signified as a very important commercial crop that can have the value chain from low-valued farm produces to high-valued, commercialized products.

4.1.1 Genetic modified sorghum

Nowadays, advances in transformation and genetic modification in plants make the development of special sorghum cultivars one of the best tactics to overcome the various known factors that reduce ethanol yields. Previous research works have concluded that fermentation efficiencies and ethanol yields are influenced by genotype and chemical composition (Wu et al., 2007, 2008; Zhao et al., 2008). These investigations have determined important traits that enhance or reduce yields. Starch, protein and tannins are the principal components related to ethanol production from sorghum grain and these characteristic can be associated to genotype and also, in the case of starch and protein, to environmental factors as sowing season and location (Wu et al., 2008). Starch composition, specifically the amylose:amylopectin ratio, is related to fermentation efficiency. Raw materials with less amylose are more efficiently converted into ethanol (Wu et al., 2006). The improvement is related to digestibility of starch, reported as higher in waxy types (Rooney & Pflugfelder, 1986). Wu et al. (2006) also attributed the increased efficiency to the lower content of amylose-lipid complexes in mashes.

The DSC thermograms of starches from waxy sorghum and waxy maize are essentially the same: both display a single, smooth endothermic peak, with approximately the same onset, peak, and ending temperatures in the range of 60-80°C. However, in normal sorghum a second peak appears around 85 to 105°C corresponding to an amylose-lipid complex that reduces the availability of starch. Waxy starches are thereby easily gelatinized and hydrolyzed, giving high conversion efficiencies (Wu et al., 2007). Thus, the waxy characteristics improved the susceptibility of the endosperm matrix for low-energy gelatinization, enzymatic hydrolysis and total ethanol production (Wu et al., 2010a).

In the case of proteins, Wu et al. (2010a) indicate that high-lysine, high-protein-digestibility (HD) sorghum lines have been developed. These genotypes have several potential advantages for their use as feedstocks in biorefineries. First, the starch granules swells and pastes more easily at lower temperatures; second, the proteins have improved feed value with higher bioavailability even for monogastrics. Interestingly, these high-lysine genotypes can contain 60% more of this essential amino acid compared to regular counterparts and similar content compared to quality protein maize (QPM) genotypes (Wu et al., 2010a). The enhanced protein digestibility of these lines is attributed to an improved kafirin digestibility as a result of the unique, abnormal and highly invaginated protein bodies. Segregated progeny with HD population lack the kafirin protein body matrix that surround starch granules and restrict swelling and pasting.

While modification in starch and protein digestibility affects ethanol production, one of the most important traits in starch conversion is total starch harvested per area. The primary goal of sorghum breeding programs has been and continues to be the development of high — yielding, drought-tolerant and pest-resistant hybrids. This effort will continue and additional gains in yield can be expected which will result in higher ethanol production from each hectare dedicated to sorghum (Rooney et al., 2007).

Ethanol is the most common renewable fuel recognized as a potential alternative to petroleum-derived transportation fuels. It can be produced from lignocellulosic materials in various ways characterized by common steps: hydrolysis of cellulose and hemicellulose to monomeric sugars, fermentation and product recovery (fig 3). The main differences lie in the hydrolysis phase, which can be performed by dilute acid, concentrated acid or enzymatically (Galbe & Zacchi, 2002).

The hybrid cybernetic approach (Kim et al., 2008; Song et al., 2009; Song & Ramkrishna, 2009) is used for modeling of recombinant yeast consuming glucose and xylose. The hybrid cybernetic model (HCM) incorporates the concept of elementary modes (EMs) (Schuster et al., 2000) into the cybernetic framework. EM is a metabolic pathway (or subnetwork) composed of a minimal set of reactions supporting a steady state operation of metabolism. Any feasible metabolic state can be represented by nonnegative combinations of EMs. HCM views EMs as cell’s metabolic options, the choice of which is optimally modulated under dynamic environmental conditions such that a prescribed metabolic objective (such as the total carbon uptake flux) is maximized.

Ethanol from biomass (Bioethanol) is one of these alternative sources. Despite polemics for biomass uses i. e. biofuels vs food and some alarmist politicians’ declarations, this alternative is really promising considering (i) the ability to satisfy a significant part of the demand for energy and (ii) biomass renewability. Polemics and limitations could have been almost rational when first generation of biofuels was concerned, "noble parts" of plants, the same used for food, being transformed. It is not the case anymore for any modern project. Another problem raised is the part of the cultivated surfaces to be reserved to biofuels, but in fact, realistic scenario is not to replace all fossil fuels volumes, but only part of them by using wastes preferentially.

The technical advantage of ethanol steam reforming over direct ethanol combustion for power generation is the improvement of thermal efficiency through hydrogen production exclusively used for fuel cell. In addition to stationary electricity generation, fuel cell is also designed for powering portable devices such as automobile. It is unsafe to travel around with compressed hydrogen tank on board. Therefore, there is a necessity for on-board steam reformer development where liquid ethanol rather than compressed hydrogen gas is fed into the storage tank. In order to get better mileage per gallon ethanol fed, the very important requirement of on-board steam reformer development is its light weight, which generates great demands on size reduction of on-board reformer. To fulfill the miniaturization and compactness requirements, various types of micro-structured reactors have been developed in recent years, which is typically composed of stacks of channeled blocks. Each micro-channel coated with active catalyst acts as the steam reformer for hydrogen production. Partial ethanol is combusted in the other side of the channel to supply heat required for reforming. Such design provides many technical advantages including rapid mass and heat transport due to large surface area to volume ratios, lower pressure drop, good structural and thermal stability, and precise control of reaction conditions leading to higher hydrogen yield [159, 160]. The main challenges faced by this technique before it becomes final commercialization are system integration, reactor fabrication process, and catalyst regeneration or replacement.

Combinatorial method originally developed for drug discovery has been introduced into the catalyst discovery field in the last decade to accelerate the catalyst screening process. By using this high-throughput approach, large and diverse libraries of inorganic materials can be prepared, processed, and tested simultaneously under the same reaction conditions for quickly obtaining potential candidates with desirable catalytic performance, which is beneficial for significant reduction of time and money spent on catalyst development [161, 162]. However, the relatively complicated algorithms for testing matrix determination, expensive testing instrument, and representability of the screening results should be better handled before it can be widely accepted as a standard catalyst development strategy.

The influence of external field (e. g., electric and magnetic field) on catalytic performance during BESR could be another interesting area to study. Because any chemical reaction involves electron transfer and rearrangement facilitated by the addition of catalyst, the application of external field which can exert impact on electron movement is expected to have influence on catalytic reactivity. Such effect has been recently evidenced by L. Yuan, et al. that hydrogen yield and selectivity were significantly enhanced when an AC current passed through Ni/Al2O3 catalyst [163].

According to LeChatelier’s Principle, referring to Reaction (1), continuous removal CO2 from product stream can shift the reaction equilibrium toward products side, leading to the improvement of hydrogen production. Based on literature review, there are mainly two methods for CO2 in-situ removal: addition of CO2 sorbent and CO2 selective membrane. The CO2 sorbent used for this purpose has to be regenerated at temperature higher than reaction temperature for reuse. For doing so, the high temperature CO2 sorbent has to be circulated between reactor and regenerator [164]. The CO2 sorbent is usually regenerated under the hot air environment and has good resistance to high temperature and attrition. According to literature reporting, CaO and lithium silicate are among the most commonly used CO2 sorbents for hydrogen production. For CO2 selective membrane, CO2 is either rejected by the membrane and stays in the retentate side, or diffuses through the membrane and swept out as permeate. In order to in-situ remove CO2 or perform hydrogen purification within the reformer, various types of membrane reactors have been developed in recent years to obtain hydrogen rich gas stream. Moreover, catalytic membrane reactor has also been invented to perform water-gas shift (WGS) and separation simultaneously through applying certain catalyst onto the membrane surface, among which Pd — impregnated membrane is the most reported one for getting purfied hydrogen product [165, 166]. Nevertheless, many technical problems including cost reduction, selectivity and permeation efficiency improvement, and rigidity enhancement have to be solved before it becomes economically attractive.

The high cost of ethanol feedstock for steam reforming mainly comes from the downstream distillation and purification steps of the crude ethanol obtained from fermentation. If the crude ethanol can be directly used as the feedstock for hydrogen production from BESR, the large amount of energy wasted during distillation for water and other impurities removal can be eliminated, leading to the significant cost reduction of ethanol feedstock and in turn hydrogen produced from BESR. In addition, other oxygenated hydrocarbons contained in the fermentation broth can also be steam reformed to generate extra 7% hydrogen if crude ethanol is employed compared to steam reforming of pure ethanol. Although this approach sounds promising for final commercialization of BESR technique, the challenge still remains at the catalyst’s tolerance to the impurities present in the crude ethanol solution. According to related publications, several researchers have conducted such study to evaluate the impact of impurities on catalytic performance toward hydrogen production. A. Akande and his coworkers investigated the influence of crude ethanol simulated through adding small amount of lactic acid, glycerol, and maltose to ethanol aqueous solution on the catalytic performance of Ni/ Al2O3 [128, 167]. Initial catalyst deactivation was observed followed by stable run within 12 hours test. Similar study has also been performed by our group over Co/CeO2. ~90% hydrogen yield is achieved and well maintained within 100 hours run. A more systematic research has been recently implemented by A. Valant, et al. over Rh/MgAl2O4 [168]. More oxygenated hydrocarbons including esters, aldehydes, amine, acetic acid, methanol, and linear or branched alchols have been tested for its influences on catalytic performance of BESR. Catalyst deactivation is observed for certain impurity additions. Through catalyst modification, much better stability has been achieved using Rh — Ni/Y-Al2O3.

Although high pressure operation will result in inhibition of hydrogen production, as predicted thermodynamically referring to Section 2.7, it is still worth investigating, because high pressure operation will significantly lower down the hydrogen compression cost for storage and transporation. In order to compensate the hydrogen production loss, hydrogen selective membrane reactor has been recently proposed in combination with high pressure operation by Argonne National Laboratory [169]. By doing so, the formed hydrogen can be continuously removed leading to the thermodynamic equilibrium shift toward hydrogen production.

2. Acknowledgment

We gratefully acknowledge funding from the U. S. Department of Energy through grant DE- FG36-05GO15033. The Ohio Supercomputer Center (OSC) is also acknowledged for generous computational support of this research.

[1] News on: Sci Focus Direct on Catalysts, 2005

[2] Genencor, relations, 21 October 2004, avaible from: http:/genencor. com/cms/connect/ genencor/media_relations/news/archive/2004/gen_211004_en. htm

[3] Genencor, products, 14 January 2010, avaible from: http:// www. genencor. com/ wps/ wcm/ connect/ genencor/ genencor/ products and services/ business development/ biorefineries/ products/ accellerase product line en. htm

[4] Novozyme, brochure, 29 January 2010, Viable from: http:// www. bioenergy. novozymes. com/ files/ documents/ Final%20Cellic%20Product%20Brochure_ 29Jan2010.pdf

Whilst the slop from molasses is very dark in color, cassava liquid waste has a light yellowish color with a lower COD (40,000-60,000) and BOD (15,000-30,000) values. The characteristics of waste water from the ethanol factories using cassava and molasses as feedstock are shown in Table 9. In consideration of this, the waste water from cassava-based process is much easier to handle than the waste obtained from molasses. This implies less investment and operational costs. In China, cogeneration of biogas obtained from waste water treatment in ethanol factory operating with cassava is reported to be able to cover all electricity needs in ethanol production process and still have some excess to supply to the grid (Dai et al., 2006). The practice for using thin stillage in Thailand is also for biogas production.

Source: Sriroth et al., 2006; n. a. = not applicable

Table 9. Characteristics of stillage obtained from ethanol factories in Thailand.

Acid hydrolysis, the most common fiber pretreatment method (Ban et al., 2008), generates significant amounts of sugars from hemicellulose. Besides it is a process relatively cheap (Gnansounou et al., 2005). Sulfuric, hydrochloric, hydrofluoric or acetic acids have been tested as catalysts (Herrera et al., 2003). The process consists on the addition of diluted aqueous acid solution (0.1 to 10 % w/ v) to the ground raw material and hydrolyzing in an autoclave. A solid residue, rich in cellulose and lignin, is formed after acid hydrolysis and subsequently treated with enzymes in order to increase the amounts of fermentable sugars (Tellez-Luis et al., 2002). Kurian et al. (2010) achieved extract with 92 g/L of total sugars from sweet sorghum bagasse treated with sulfuric acid at a concentration of 5 g/kg and treated at 140°C for 30 minutes. Ban et al. (2008) treated the same raw material at a solid — liquid mass ratio of 10% with 80 g phosphoric acid/L at 120°C for 80 minutes. These authors reported 302 g reducing sugars/kg with this pretreatment.

The composition of LCB depends on the plant species and consists primarily of cellulose, hemicelluloses and lignin, which are the integral part of cell wall in plant tissues (Fig. 3) (Fengel et al. 2003). Lignin is an amorphous aromatic biopolymer composed of phenyl propane structural units linked by ether and/or carbon-carbon bonds, supplying tissues stiffness, antiseptic, and hydrophobic properties amongst others (Fig. 4). The types of lignin structural units (p-hydroxyphenyl, guaiacyl and syringyl units), their abundance, types and frequency of inter-unit linkages vary significantly from plant to plant (Fengel et al. 2003). Lignin contributes to 15-30% of plant biomass and is the principal non-hydrolysable residue of LCB.

Cellulose and hemicelluloses are hydrolysable structural polymers of cell wall and the main sources of fermentable sugars (Lawford et al. 1993; Sanchez et al. 2008). Hemicelluloses contribute to 10-40% of plant material and are essentially heteropolysaccharides constituted

by pentoses, mainly D-xylose and L-arabinose, and hexoses, mainly D-mannose, D-galactose and D-glucose. These monosaccharides result from pentosans with a main backbone built by pentoses, and hexosans with a main backbone built by hexoses. Hemicelluloses possess an irregular structure and are chemically linked to lignins in the cell wall (Sjostrom 1993). The structure and the composition of hemicelluloses vary significantly among plant species. The most abundant hemicelluloses are xylans followed by mannans and galactans (Fig. 5). Hemicelluloses play an important structural role in cell wall regulating the spatial

distribution of principal macromolecular components (cellulose and lignin) and providing their compatibility.

Cellulose, the most abundant structural polysaccharide (30-50% abundance in the cell wall), is comprised by repeated P-D-glucopyranose units linked by P(1^4)-glycosidic bonds. In plant cell walls, cellulose chains aggregate into elementary fibrils (EF) which, in turn, are assembled into microfibrils (MF). MF are embedded into a matrix of lignin and hemicelluloses, thus becoming isolated of each other (Fig. 3). Plant cells assembled in different tissues are also separated by a layer enriched in lignin (middle lamella). This structural hierarchy hinders either chemical or enzymatic hydrolysis of cellulose, being the last one particularly difficult. Cellulose, the amorphous-crystalline polymer, is poorly accessible to hydrolysis due to the predominance of crystalline domains.

.OCH3

CH3O.

PiR link

HOH2C

J PhC ink

CH2OH

CH3O OCH3 OH

O-acetyl-4-O-methylglucuronoxylan

O-acetyl-galactoglucomannan

OH

arabinogalactan

Fig. 5. Schematic representation of major hemicelluloses in lignocellulosics