Nonedible oils that may be used in biofuel production include Jatropha, Pongamia, jojoba, linseed and cotton seed oil. Nonedible oils are not suitable for human consumption due to the presence of toxic compounds in the oils, for example, curicin present in Jatropha oils is a toxic lectin. Biofuels from nonedible lipids have many advantages over the edible alternative including the ability of these organisms to grow in harsh nutrient — and moisture-limiting conditions and the reduction in carbon emissions. Nonedible oils are generally more cost-effective as they do not have applications in food production and thus are lower value oils, containing low sulfur concentrations and low aromatic compound concentrations and the lipids produced are biodegrad­able (No, 2011). A disadvantage of using nonedible oils is the large amounts of free fatty acids (FFAs) that cannot be converted into biodiesel using an alkaline catalyst (Demirbas et al., 2011).

Jatropha is one of the most widely used nonedible oils due to the high potential yield of 0.5—12 tons per hectare per year; the yield is highly effected by the conditions in which it is produced, and the ability of the organism to grow in harsh environmental conditions of low water availability and low nutrient content (Francis et al., 2005). The oil produced by Jatropha has good cold flow properties due to the composition of the oil. The Jatropha plant is a small tree and produces seeds with high lipid content. In addition to the drought resistance within the plant it is also pest tolerant and unpalatable to animals and grows rapidly with a lifetime of 30 years; each of these factors makes it a suitable choice for the produc­tion of biofuels. The ability of the plant to grow in harsh conditions led to Jatropha being considered a revolution­ary plant that could provide the solution for the produc­tion of large volume of lipids without competing with the food industry. However, when grown in marginal lands studies revealed that the number of seeds pro­duced by the plant was quite low and although the tree is capable of growing in low nutrient conditions, the lipid production is low (Pandey et al., 2012). There­fore, the economic returns of Jatropha grown on marginal lands is low; however, growing the crop in developing areas with poor land may be a viable method of produc­tion of oil on a small scale. The energy balance from the crop is also low if only the seeds are used for the produc­tion of biofuel; however, the value is increased if all components, for example, the husks are also utilized (Prueksakorn and Gheewala, 2006).

and Chemicals

Lin Mei Wu1, Chun Hui Zhou1’2’*, Dong Shen Tong1, Wei Hua Yu1

^Research Group for Advanced Materials & Sustainable Catalysis (AMSC), Breeding Base of State Key Laboratory
of Green Chemistry Synthesis Technology, College of Chemical Engineering and Materials Science,
Zhejiang University of Technology, Hangzhou, Zhej’iang, China,

2The Institute for Agriculture and the Environment, University of Southern Queensland, Queensland, Australia
*Corresponding author email: clay@zjut. edu. cn, Chun. Zhou@usq. edu. au

OUTLINE

Introduction 243

Pyrolysis of Biomass 244

Fast Pyrolysis 244

Catalytic Pyrolysis 244

Reactors 245

Entrained-Fow Reactors 245

Ablative Reactors 245

Bubbling Fluid Bed Reactor and Circulating Fluidized Beds 245

Rotating Cone Reactor 246

New Systems 247

Gasification of Biomass 247

INTRODUCTION

Thermochemical processing usually refers to the one in which solid reactants are heated at high tempera­tures for a certain period to yield the desired products. In modern times, the thermochemical processing has often been used in industry for the production of fuels, chemicals and materials. Today, the production of fuels, chemicals and materials from biomass become attrac­tive because it has renewability, one of the advantages
over the fossil oil sources (Zhou et al., 2011; Wu et al.,

2013) . In a sense, thermochemical processing of biomass is not a new technique. Wood combustion for heating and cooking, a method that humanity have been using since prehistoric time can be regarded as a thermochemical processing of biomass. However, to­day’s need for thermal processing of biomass is far beyond combustion. The combination of thermal pro­cessing and catalysis is bringing about new opportu­nities for using biomass to produce renewable fuels,

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chemicals and materials (Brown, 2011). The past three decades have witnessed rapid progress in catalytically thermochemical technologies (Zhou et al., 2008; Huber et al., 2006; Fan et al., 2009). Pyrolysis, gasification and hydrothermal liquefaction are major methods frequently tested for the catalytically thermochemical conversion of biomass (Zhou et al., 2011). Other ther­mochemical technologies could be regarded as modifi­cation, in more or less ways, of these three methods. Relevant studies and progress have shown that these technologies are promising alternatives to process diverse biomass feedstocks to yield fine chemicals and biofuels.

Lignin offers a significant opportunity for enhancing the operation of a lignocellulosic biorefinery. It is an extremely abundant raw material contributing as much as 30% of the weight and 40% of the energy con­tent of lignocellulosic biomass (Holladay et al., 2007). Lignin’s native structure suggests that it could play a central role as a new chemical feedstock, particularly in the formation of supramolecular materials and aro­matic chemicals (Holladay et al., 2007; Hatakeyama and Hatakeyama, 2010). Up to now the vast majority of industrial applications have been developed for

lignosulfonates. These sulfonates are isolated from acid sulfite pulping and are used in a wide range of lower value applications where the form but not the quality is important. The solubility of this type of lignin in water is an important requirement for many of these applica­tions. Around 67.5% of world consumption of lignosul­fonates in 2008 was for dispersant applications followed by binder and adhesive applications at 32.5%. Major end-use markets include construction, mining, animal feeds and agriculture uses. The use of lignin for chemical production has so far been limited due to contamination from salts, carbohydrates, particulates, volatiles and the molecular weight distribution of lignosulfonates. The only industrial exception is the limited production of vanillin from lignosulfonates (Evju, 1979). Besides ligno — sulfonates, kraft lignin is produced as commercial prod­uct at about 60 kton/year. New extraction technologies, developed in Sweden, will lead to an increase in kraft lignin production at the mill side for use as external energy source and for the production of value-added applications (Ohman et al., 2009).

The production of bioethanol from lignocellulosic feedstocks could result in new forms of higher quality lignin becoming available for chemical applications. The Canadian company Lignol Energy has announced the production of cellulosic ethanol at its continuous pi­lot plant at Burnaby, British Columbia. The process is based on a wood pulping process using Canadian wood species but the pilot plant will test a range of feed­stocks while optimizing equipment configurations, enzyme formulations and other process conditions (Lignol Energy. 2013). The Lignol Energy process pro­duces a lignin product (HP-L lignin) upon which the company is developing new applications together with
industrial partners. Also other lignin types will result from the different biomass pretreatment routes under development and unfortunately there is not one lignin macromolecule that will fit all applications. However, if suitable cost-effective and sustainable conversion technologies can be developed, a lignocellulosic bio­refinery can largely benefit from the profit obtained from this side stream lignin (Gosselink, 2011).

The production of more value-added chemicals from lignin (e. g. resins, composites and polymers, aromatic compounds, carbon fibers) is viewed as a medium — to long­term opportunity that depends on the quality and func­tionality of the lignin that can be obtained (Figure 17.3, Table 17.8). The potential of catalytic conversions of lignin (degradation products) has been recently reviewed (Zakzeksi et al., 2010).

The main chemical building blocks can be organized by their carbon number, i. e. C1— Cn. In the following sec­tions, examples of biobased chemicals are discussed with respect to their current status and the companies that are pursuing the development of these new chemicals.

PGA was first observed by Ivanovics et al. produced by Bacillus anthracis strain during 1937. The traditional Japanese food natto (fermented product made from soyabean) consists of PGA and a fructan, and is pro­duced through fermentation using the strain Bacillus natto. PGA produced can be essentially of three types,

a polymer purely made of either D-glutamic acid, L-glu — tamic acid, or d and L-glutamic acid. Besides Bacillus, other microbial species from Archaea (Natrialba aegyp — tiaca, Natronococcus occultus), eubacteria as well as eu­karyotes (hydra) were found to produce PGA (Kocianova et al., 2005; Hezayen et al., 2001). PGA is one of the most utilized poly(amino acid)s, with multi­tude of applications ranging from agriculture to cos­metics, food industry and even pharmaceuticals (Buescher and Margaritis, 2007).

Gustavo B. Leite, Patrick C. Hallenbeck*

Departement de Microbiologie et Immunologie, Universite de Montreal, Montreal, Quebec, Canada
Corresponding author email: patrick. hallenbeck@umontreal. ca

OUTLINE

Introduction 389

Engineering Cyanobacteria 392

Strains, Tools and Methods 392

Cyanobacteria as a Production System for Biofuels: Current Status 393

Hydrogen 393

Hydrogen Bioproduction 394

Hydrogen-Evolving Enzymes 394

Hydrogen Bioproduction 395

Ethanol 398

Ethylene 398

Microbial Production of Ethylene 399

Bioproduction of Ethylene Using efe 399

Isoprene 400

Butyraldehyde and Butanol 401

Photosynthetic Production of Aliphatic Alcohols and Alkanes 402

Conclusion and Outlook 403

References 403

INTRODUCTION

Paleontological and geochemical data as well as molecular analysis of the plastid genome point to a single prokaryote as the origin of several groups of organisms scattered throughout the tree of life, including the entire kingdom of Plantae (Knoll, 2008; Yoon, 2004). A cyano- bacterial ancestor is believed to be the only organism ever to couple together two photosystems, harvesting electrons from water to produce energy-rich molecules such as adenosine triphosphate (ATP) and reduced nico­tinamide adenine dinucleotide phosphate (NADPH) (Knoll, 2008) (Figure 22.1). These molecules provide the necessary chemical energy, protons and electrons for cellular reactions and the synthesis of other molecules, most importantly powering CO2 fixation through the Calvin-Benson-Bassham cycle. This event is thought to have happened between the mid-Archean and early Pro­terozoic eras (2000—3000 millions of years ago). The
atmosphere was poor in oxygen and rich in CO2, and the oceans were rich in salts and minerals; perfect conditions for the first algal blooms. The invention of oxygenic photosynthesis conferred a great advantage to this ancient cyanobacterium, starting widespread speciation and changing the composition of the atmo­sphere through the oxidation of water into protons and molecular oxygen (Figure 22.1). This was probably the first universally relevant instance of primary production and established a food chain by transforming inorganic nutrients into organic molecules that could be used by heterotrophic organisms (Knoll, 2008). The role of primary producers, so important in fully establishing life on earth, is still equally important today, when cyano­bacteria are thought to be responsible for 25% of all carbon dioxide fixation and together with eukaryotic microalgae sustain most of oceanic life, fixing CO2 and carrying out important steps in various biogeochemical nutrient cycles (Field et al., 1998).

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Humanity is totally dependent on photosynthesis for food and fuel. As well as a source of organic carbon, mankind relies on photosynthesis as energy source, through the use of fossil fuels, ancient photosynthetic products stored and cooked under pressure for millions of years, the burning of readily available biomass, or more recently through the use of biofuel crops as a new source of liquid fuels. Sugarcane or corn ethanol and biodiesel have been produced from crops for more than 40 years, with a greatly increased role the last two decades. These first-generation biofuels are presently being produced at large scale, with worldwide produc­tion of ethanol and biodiesel of 50 billion and 9 billion liters, respectively, in 2007. Even though these seem like significant quantities, biofuels still represent a miniscule fraction of the world’s primary energy use; in 2011, 161 tons per day of renewable liquid biofuels were produced, whereas 12 million tons per day of crude oil were consumed (BP, 2012).

Humans have been constantly perfecting agricultural technology since the dawn of civilization, and with the green revolution, food crop yields have shown consider­able increases decade after decade, although this progress is now stagnating in many food producing areas (Ray et al., 2012). At any rate, given the enormous demand for energy and the predicted increase in the world’s population to 9 billion by 2050, it is evident that there is not enough arable land to satisfy both nutritional and energy demands through food and fuel crops. Of course, in addition to renewable energy derived through photosynthesis, other sources of
sustainable energy exist: solar, wind, geothermal, hydro­electric, etc., but together these energy sources cannot supply the quantity and types of energy demanded worldwide since electricity is not suitable for all applica­tions. Modern society is built around liquid and gaseous fuels, which are very efficient energy carriers suitable for a variety of applications, in particular mobile power. Liquid biofuels are essentially photosynthetically derived compounds, at present sustainably produced through the cultivation of energy crops, but as discussed above, this directly competes with the production of food crops.

A possible and promising alternative for sustainable energy production system is intimately related to crude oil formation over the previous millions of years. Before the appearance of vascular land plants on earth, ancestral cyanobacteria were already occupying a large variety of environments and now, after a long period of evolution, cyanobacteria and the microalgae formed through endosymbiosis of cyanobacteria, can be isolated from virtually any natural water sample, from extremely fresh water to hypersaline lakes, from snow in the Arctic Circle to hot or relatively dry environments. The richness of this speciation over billions of years can be appreciated through the variety of morphological forms that are found. These organisms show themselves to be a promising system for the production of hydrocarbons and other desirable products. Cultivation can be carried out using nonarable land; seawater and wastewater have been shown to support growth, bioremediating effluents while fixing atmospheric carbon dioxide into
possible commercial products. The rather simple nutri­tion requirements of these organisms highlight the capa­bility of their metabolism to produce all the molecules needed for cellular growth. Their pathways frequently contain metabolites with commercial interest that can be readily used or easily processed into a final product (Figure 22.2).

Although cyanobacteria and eukaryotic algae share these attributes, cyanobacteria have the additional advantage of being relatively easily manipulated genet­ically. Thus, using cyanobacteria, if a desired product is not naturally produced, genetic engineering techniques allow the insertion of genes or even entire pathways to make novel products, either high-value compounds or

commodity chemicals such as biofuels. Of course any molecule that is produced using cyanobacteria could be produced in other microorganisms, especially fermentation workhorses such as Saccharomyces cerevi — siae or Escherichia coli, but these are heterotrophs requiring carbon compounds previously fixed through photosynthesis, i. e. agriculturally produced.

Thus, the cyanobacteria are uniquely positioned to carry out CO2 fixation driven by solar energy capture while at the same time being amenable of genetic engi­neering to produce a wide variety of liquid and gaseous biofuels. In this chapter the current achievements on research toward the production of biofuels and crude oil substitutes using cyanobacteria as a model organism are reviewed. As will be seen, although much has already been achieved in terms of engineering toward the production of biofuels, in most cases productivity is the greatest bottleneck, although some steps in down­stream processing also present many challenges. Thus, at present, use of a cyanobacterial system for commer­cial production of biofuels at cost-effective levels still faces significant hurdles.

Raw materials (containing saccharides) include sweet sorghum, sugar beet, banana, mango, watermelon, sug­arcane and other fruits; these are examples of sugar feedstock. The wastes of these sugar-containing sources can be fermented by using different microorganisms of interest. However, the use of these materials for
bioethanol production is highly expensive and humans use them as food (Kahn et al., 2011).

The first research about bioethanol showed in Table 3.1 reports the production of biofuel using resi­dues of fruits and vegetables. These are an important source of sugar for ethylic fermentation because the pro­cessing of fruits have a great potential to generate resi­dues that can be used. The authors report the use of enzymes in the process; this is necessary because these residues have a lot of fiber that can be hydrolyzed (Patle and Lal, 2007). The use of wastes is the main aspect of two other studies (Gouvea et al., 2009; Ge et al., 2011). Gouvea et al. (2009) reported the use of a residue very abundant in Brazil, that is, the coffee husk. As well as in the case of fruit and vegetable residues the floating seaweed wastes have other kinds of carbohydrates that were hydrolyzed by enzymes (Wu et al., 2011). Ge et al. (2011) reported the use of a principal sugar source for ethylic fermentation, the stalk juice of sugarcane. This juice has great quantity of glucose in its composi­tion and has a low cost. More studies about sugarcane juice were omitted since the main subject of this chapter is agroindustrial residues.

Our knowledge about cell wall biosynthesis and biodegradation has steadily increased in the past decades, although there is still a long way to go to fully understand these extremely complex processes. So far our knowledge is at best fragmented and characterized genes are often from various organisms and dispersed in the literature. Therefore, dedicated, centralized and frequently updated databases of bioenergy-related genes are crucial to guide the development of transgenic biofuel crops and the annotation of newly sequenced metagenomes/genomes to look for novel enzymes. Like other biology research areas, bioenergy research has also been benefiting from bioinformatics. Table 6.2 provides a list of bioinformatics databases and web — based resources developed specifically for bioenergy — related enzymes as well as some general bioinformatics web resources that are valuable for bioenergy research. Here we offer a summary of a few selected resources that are particularly useful.

The intended final biofuel product defines successful microalgae cultivation. If biodiesel is the final product, algal strains should be selected and cultured to produce maximal saturated fatty acids. If biocrude is the desired product, high organic content, or a simple abundance of biomass, is required. Whatever the target product, suc­cessful cultivation requires specific environmental condi­tions to drive the production of specific fuel precursors. Major parameters that influence biomass production include adequate light (wavelength and intensity), tem­perature, CO2 concentration, nutrient composition, salinity, contaminants, and mixing conditions.

PHOTOTROPHIC MICROALGAE

Phototrophic microalgae use carbon dioxide (carbon source), sunlight (energy source), and nutrients to prolif­erate. Two properties of light energy are important for algal growth and metabolism: quality of the light spec­trum and quantity of the light photons. As phototrophs, light-harvesting pigments (chlorophyll and carotenoids) absorb light at specific wavelengths to drive the photo­synthetic process. Light absorption, however, is hin­dered both by light scattering through increasing depths of the culture medium and by mutual shading as the culture increases in density. Antenna structures of microalgae are excessively efficient at harvesting light energy, absorbing all the photons that hit them even though only a fraction of those photons are used for photosynthesis. This deprives nearby algae from absorbing photons and consequently leads to low pro­ductivity. Aggressive mixing of the culture mitigates some of these effects, but cannot completely overcome the light penetration limitations inherent in a photosyn­thetic system.

HETEROTROPHIC MICROALGAE

Several wild-type and genetically modified species of microalgae have been reported capable of growing pho — totrophically, heterotrophically or both (mixotrophi — cally). Unlike phototrophic algae that require light energy, heterotrophic algae have no such requirement. Instead, these algae utilize organic carbons supplied in the media to drive cellular proliferation and lipid accu­mulation. Without the limitations imposed by inefficient light harvesting due to mutual shading and light scat­tering in the medium, the densities of heterotrophic cul­tures can far exceed the densities achieved in phototrophic systems. Increased densities can translate to higher biofuel precursor yields. For example, when Chlorella protothecoides was grown heterotrophically us­ing an organic carbon source, oil accumulation far exceeded that seen in corresponding autotrophic cells (Miao and Wu, 2004). Hence, heterotrophic production has several advantages over phototrophic systems including increased densities that eliminate the need for dewatering, and increased process control that facil­itates the maintenance and rapid growth of monocul­tures and the creation of a consistent product. The primary limitation for commercial-scale heterotrophic production of biofuel oils in microalgae is the cost of the organic carbon source. Sugars such as glucose and acetate have been utilized as the primary carbon source at the bench scale, but become cost-prohibitive at pro­duction scale. It is therefore unsurprising that increased efforts to identify microalgal species that can thrive on waste sugars, such as bagasse or cellulosic waste, are underway.

Following extraction and regardless of the process described above, the end product will generally be a rather impure biolipid that contains undesirable contents such as FFAs, tocopherols, waxes and possibly phosphatides. The latter, if not removed before storage, will produce a thick gum over time. Gums are formed when the biolipid absorbs water, which causes some of the phosphatides (such as phosphocholine) to become hydrated and thereby lipid insoluble. Accordingly, hydrating the gums and removing the hydrated gums from the oil before storage can prevent the formation of a gum deposit. This treat­ment is called water degumming and involves the addi­tion of water at 60—90 °C before the phase is separated. An optimum temperature is sought, as it must not be so high as to increase the solubility of phosphatides in oil. A temperature that is too low will increase the viscosity, making phase separation more difficult. It is never applied to fruit oils like olive oil and palm oil, since these oils have already had considerable water contact during their pro­duction. The removal of nonhydratable phosphatides (such as phosphatidic acid) requires the addition of an acid, usually citric or phosphoric, which will form a sludge that can be easily removed (Dijkstra and Van Opstal, 1989). This addition of acid is proportional to the amount of phosphorous already contained in the sample. In addi­tion, this acid also reduces any iron salts and decreases chlorophyll contamination. Enzymatic degumming fo­cuses on the use of lipases, which convert nonhydratable lipids to more hydratable forms. Although the process has been tried at a larger scale for 20 years, it has not made the advancement toward widespread use (Dijkstra, 2010; Yang et al., 2008).

Two different clay minerals (namely BBn and VLn), mined in Boa Vista District of Paralba (Brazil), were acid-activated using a 10 wt% suspension of clay in 4mol/l sulfuric acid at 90 °C for 2 h to produce the BBa and VLa solids, respectively. The solids were filtered under reduced pressure and washed with distilled water until the washing water had the same pH of the original material. The material was dried in
an oven at 110 °C for 24 h and finally ground until pas­sage through a 60 mesh sieve. The surface area of the acid-activated clays was improved from 71 m2/g to 238 m2/g for the VLn sample and from 123 m2/g to 170 m2/g for the BBn sample. The catalytic activity of these compounds was evaluated in the esterification of different organic acids, using different acylating agents and reflux conditions. In the case of methyl and hexyl esterification of lauric acid (Figure 16.6), the sulfuric acid activation of both clays greatly improved their cat­alytic activity and this was also valid for other acids and acylating agents. As reported in Case 2, acid activation led to catalytic activities higher than the standard Lewis acid catalyst (K10).

In conclusion, clay minerals are cheap inorganic ma­terials that are readily available worldwide, environ­mentally friendly and suitable for the development of reusable acid catalysts for the esterification of fatty acids and the transesterification of oils and fats. The natural acidity can be improved by thermal treatment and selec­tive acid activation. Depending on the clay minerals’ origin and genesis, different chemical compositions are possible and different acid treatments are needed to optimize the acidic properties. Normally, the catalysts can be used in several consecutive reaction cycles and, after deactivation, the residual solids can be easily disposed of or even incorporated in native clays for the production of ceramic materials, bricks and roofs, as well as in the production of porcelains.