Anne M. Ruffing

Additional information is available at the end of the chapter http://dx. doi. org/10.5772/52050

1. Introduction

The world’s supply of petroleum hydrocarbons, which serve as feedstock for the fuel and chemical industries, is rapidly diminishing to satisfy the global demand for energy and consumer goods. In response to this increasing demand and limited supply, the cost of crude oil has risen to over $100 per barrel in 2012, a 10-fold increase compared to prices in the late 1990s [1]. As fossil fuels are nonrenewable resources, the price of oil is only expected to increase in the future. This unavoidable reality necessitates the development of renewable energy sources in order to maintain the current standard of living. Among the alternative energy options under development, biofuels are anticipated to supplement and eventually replace the petroleum-based fuels that supply the transportation and chemical industries. Currently, first generation biofuels like corn-based ethanol are blended into conventional petroleum fuels, with biofuels supplying 2.7% of the world’s transportation fuel in 2010 [2]. It appears that biofuels are on their way to becoming a viable renewable energy source, yet technological and biological advancements are necessary for sustainable and economical biofuel production at the scales necessary to support the world’s energy needs.

The current practice of using food crops, like corn or soybean, as feedstocks for biofuel production is not a viable, long-term solution to the energy crisis. In fact, to replace our current petroleum usage with crop-based ethanol production, the entire surface area of land on Earth would be needed for corn production [3]. In addition to this shortcoming, first generation biofuels compete with food production for arable land, require significant nutrient resources (fertilizer and fresh water), and typically have low net energy yields due to the low energy density of the product fuel (i. e. ethanol) and the energy input required to harvest the feedstock and convert it into fuel [4]. Second and third generation biofuels address these limitations. Second generation biofuels use lignocellulosic biomass as the feedstock for fuel production.

Lignocellulose, the main component of plant biomass, is the most abundant form of renewable carbon on the Earth, making it an ideal feedstock for renewable hydrocarbon production. The cellulose and hemicellulose components of lignocellulose can be degraded into fermentable sugars to serve as the carbon source for microbial-based fuel production. The carbon feedstocks for both first and second generation biofuels are ultimately derived from carbon dioxide (CO2) fixation through the process of photosynthesis. Third generation biofuels use photo­synthetic microorganisms (i. e. microalgae) to directly convert CO2 into fuel molecules or fuel precursors, eliminating the biomass intermediate (Figure 1). While both second and third generation biofuels require land, nutrients, and energy investment for harvesting and fuel production, the fuel production yields from these processes are predicted to be capable of meeting energy needs. However, these technologies have yet to be demonstrated at scale and still require further improvement before they can be economically competitive with fossil fuels.

Both second and third generation biofuels rely on microbes to convert the carbon feedstock into the desired hydrocarbon fuels. Microorganisms have been identified that are capable of producing a range of fuel molecules and fuel precursors, yet the natural rates of microbial fuel synthesis are typically too low to support industrial-scale production. Metabolic engineering is a powerful tool to improve microbial fuel production, either through engineering the metabolic pathways within the native microorganism to encourage high fuel synthesis or though transferring the fuel production pathway into a model organism for optimization. This chapter will focus on the application of metabolic engineering to increase hydrocarbon fuel production. Within this chapter, hydrocarbon-based fuels are defined to include oxygen — containing fuel molecules with long hydrocarbon chains, such as fatty alcohols and fatty acid ethyl esters (FAEE), in addition to pure hydrocarbons like alkanes, alkenes, and isoprenoid — based molecules: hemiterpene (C5), monoterpenes (C10), and sesquiterpenes (C15). Hydro­carbon-based fuel precursors will also be considered, including free fatty acids (FFAs) and triacylglycerol (TAG). The structures of these hydrocarbon-based fuels and precursors are illustrated in Figure 2. Hydrocarbon-based fuels and precursors can be produced by both second and third generation biofuel processes. Therefore, the first section in this chapter will

discuss the metabolic pathways for hydrocarbon fuel production and common metabolic engineering strategies for improving fuel synthesis. Because second and third generation biofuel processes rely on different carbon sources, sugars and CO2 respectively, the remaining sections will focus on the use of organic carbon (heterotrophy) and inorganic carbon (auto­trophy) as feedstocks for biofuel production. This division, based on carbon source, is impor­tant from both the biofuel production and metabolic engineering perspectives. The chapter will conclude with a discussion of the future outlook for microbial-based, hydrocarbon fuel synthesis.

Figure 2. Chemical structures of hydrocarbon-based biofuels and fuel precursors. (A) Fuels derived from fatty acid bio­synthesis and (B) fuels derived from isoprenoid biosynthesis, including (1) hemiterpene, (2) monoterpenes, and (3) ses­quiterpenes.