Chapter 6 – Oil production

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6.1 Oil substitutes from biomass
6.2 Microalgae as biological sources of lipids and hydrocarbons
6.3 Thermochemical liquefaction of microalgae
6.4 Algal hydrogenation
6.5 Future prospects
References

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6.1 Oil substitutes from biomass

It is believed that the majority of oil and natural gas originates from algae in ancient oceans. Oil (petroleum) consists of liquid hydrocarbons which arc compounds composed of carbon and hydrogen. At least 80% w/w of oil is carbon. The remainder is principally hydrogen, but sulfur and oxygen may each account for up to 5% of the weight of oil. The burning heating volume of oil is relatively high owing to its liquid state, and is comparable to that of coal.

Total proven oil reserves worldwide are estimated to be worth the equivalent of 40 years of consumable oil, based on a 1988 worldwide oil production rate of 64.2 million barrels per day. Proven oil reserves include residual oil in oil fields where production drilling has already begun. Projected oil reserves are slightly greater than proven oil reserves, but are not however infinite. Furthermore, oil reserves are not equally distributed globally (Fig: 6-1). The Middle East has by far the world’s greatest proven oil reserves. Oil production in other nations will decrease as their reserves decline, putting the Middle East in a dominant position in the oil market. The Organization of Petroleum Exporting Countries (OPEC) is in control of approximately 60% of the world’s oil, and exercises a strong influence on oil prices worldwide. The lack of stability of future energy supplies has motivated the development of alternative energy sources in order to eliminate the possibility of a future energy shortage.

Figure 6.1 – Global distribution of proven oil reserves

Solar energy is renewable, whereas all other fuels including those of fossil and nuclear origin, are limited in amount and are exhaustible. One efficient method of capturing solar energy is through the use of the photosynthetic process to produce biomass (a renewable raw material resource for the production of food, fuel and chemicals) through appropriate conversions.

This Chapter examines developments in microalgal oil production. Microalgae posses several attractive characteristics:

1) Costs associated with the harvesting and transportation or microalgae are relatively low, in comparison with those of other biomass materials such as trees, crops, etc.2) By virtue of their relatively small sizes, microalgae can be easily chemically treated.

3) Algae can be grown under conditions which are unsuitable for conventional crop production.

4) Microalgae are capable of fixing CO₂ in the atmosphere, thus facilitating the reduction of increasing atmospheric CO₂ levels, which are now considered a global problem.

Microalgal oils are produced through either biological conversion to lipids or hydrocarbons or thermochemical liquefaction of algal cells.

6.2 Microalgae as biological sources of lipids and hydrocarbons

Microalgae contain lipids and fatty acids as membrane components, storage products, metabolites and sources of energy. The chemical compositions of various microalgae are shown in Table 6-1. Algal fatty acids and oils have a range of potential applications. Algal oils posses characteristics similar to those offish and vegetable oils, and can thus be considered as potential substitutes for the products of fossil oil (2).

Direct extraction of microalgal lipids appears to be a more efficient methodology for obtaining energy from these organisms, than is the fermentation of algal biomass to produce either methane or ethanol. The lipid and fatty acid contents of microalgae vary in accordance with culture conditions In some cases, lipid content can be enhanced by the imposition of nitrogen starvation or other stress factors. In the late 1940s, lipid fractions as high as 70 to 85% on a dry weight basis were reported in microalgae. Such high lipid contents, exceed that of most terrestrial plants. The effect of nitrogen on the lipid fraction and on cell growth of the strain Nannochlolis cultured under saline conditions is summarized in Table 6-2.

Nutrient deficiencies (other than nitrogen deficiency) may also lead to an increase in cellular lipid content. Coomls, et al. (3) reported that the lipid content of the diatom Navioua pelliculosa increased by about 60% during a 14-hour silicon starvation period.

Table 6-1 Chemical Composition of Algae Expressed on A Dry Matter Basis (%))

Strain	Protein	Carbohydrates	Lipids	Nucleic acid
Scenedesmus obliquus	50-56	10-17	12-14	3-6
Scenedesmus quadricauda	47	–	1.9	–
Scenedesmus dimorphus	8-18	21-52	16-40	–
Chlamydomonas rheinhardii	48	17	21	–
Chlorella vulgaris	51-58	12-17	14-22	4-5
Chlorella pyrenoidosa	57	26	2	–
Spirogyra sp.	6-20	33-64	11-21	–
Dunaliella bioculata	49	4	8	–
Dunaliella salina	57	32	6	–
Euglena gracilis	39-61	14-18	14-20	–
Prymnesium parvum	28-45	25-33	22-38	1-2
Tetraselmis maculata	52	15	3	–
Porphyridium cruentum	28-39	40-57	9-14	–
Spirulina platensis	46-63	8-14	4–9	2-5
Spirulina maxima	60-71	13-16	6-7	3-4.5
Synechoccus sp.	63	15	11	5
Anabaena cylindrica	43-56	25-30	4-7	–

Source: Becker, (1994).

Table 6-2 Effect of Nitrogen Concentration (as KNO₃) on Microalgal Lipid Content

KNO3 cone. (mM)	Cell growth (g/L)	Internal lipid content (g/L)
0.9	0.39	42.4
9.9	2.5	32.9
9.9 + feeding	2.6	33.6

Similarly, Werner (4) also reported an increase in cellular lipids during a 24 hours silicon starvation period. The switch from carbohydrate accumulation to lipid accumulation in these diatoms occurs very rapidly, though mechanisms involved are not yet understood.

The effects of growth conditions, growth stage of algal cultures and the taxonomic position of algae, on both lipid content and lipid type, arc discussed in recent reports by Piorreck and coworkers (5, 6). According to these reports, during early stages of growth, green algae produced relatively large amounts of polar lipids and polyunsaturated C₁₆ and C₁₈ fatty acids. On approaching the stationary phase of growth, however, the dominant lipids produced by these algae were neutral, and consisted primarily of saturated 18:1 and 16:0 fatty acids. In the case of blue-green algae, the lipid and fatty acid composition showed relatively little change during the growth cycle.

In addition to nutrition, fatty acid and lipid composition and content are influenced by a number of other factors. Light enhances the formation of polyunsaturated C₁₆ and C₁₈ fatty acids as well as mono- and di-galactosyl-diglycerides, sphingolipids and phosphoglycerides in Euglena gracils and Chlorella vulgaris (7-10). Low temperatures increase the synthesis of polyunsaturated C₁₈ fatty acids by Monochrysis lutheri (11), and also cause changes in the fatty acid composition of Dunaliella salina (12).

Glycerol content is also influenced by culture conditions, particularly NaCl concentrations. Figure 6-2 shows the relationship between NaCl concentration, cell growth and glycerol accumulation in Dunaliella tertiolecta. The highest yield of glycerol was obtained at a NaCl concentration of approximately 2M, while maximal algal growth occurred at a lower NaCl concentration.

Figure 6.2 – Relationship between NaCl concentration and intracellular-extracellular glycerol concentration (Tanisho, 1995)

Botryococcus is a well known hydrocarbon producer. Under unfavorable conditions of growth, Botryococcus enters a stage in which its unsaponifiable lipid content increases to a level of 90 %. The bulk of the Botryococcus hydrocarbon (ca. 95%) is extracellularly located in the colony matrix and in occluded globules (13). While small amounts of hydrocarbons are produced by various algae, Botryococcus is the only alga that has been extensively investigated for its hydrocarbon-producing ability. Almost all the information collected on Botryococcus is based upon indoor algal cultivation; mass cultivation and bulk production of hydrocarbons outside of laboratory conditions, have not yet been reported (14). There are two main reasons for this. The first is the low doubling time of algae (which exceeds seven days). Owing to the energy-intensive nature of hydrocarbons, their mass accumulation in microalgae is less than that which occurs in typical unicellular green algae. However, increased hydrocarbon productivity may be feasible by modifying culture conditions, and/or the bacterial strains used; modifications which according to the literature, result in doubling times of two to three days. Hence, it seems worthwhile to examine a variety of strains for better growth performance and productivity. The second reason being that high hydrocarbon concentrations exist only in non-growing, senescent and even decaying cultures; the hydrocarbon content of growing algae is fairly low.

6.3 Thermochemical liquefaction of microalgae

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6.3.1 Liquid fuels from microalgal biomass
6.3.2 Cultivation of microalgae
6.3.3 Liquefaction of microalgae

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6.3.1 Liquid fuels from microalgal biomass

It is well known that microalgae can assimilate CO₂ gas as a carbon source for growth. However, if the resulting cell mass is not suitably treated, CO₂ will be evolved and diluted into the environment by decomposition, thus preventing CO₂ fixation from contributing to a reduction in atmospheric CO₂.

Petroleum is widely believed to have its origins in kerogen, which is easily converted to an oily substance under conditions of high pressure and temperature (15-17). Kerogen is formed from algae, biodegraded organic compounds, plankton, bacteria, plant material, etc., by biochemical and/or chemical reactions such as diagenesis and catagenesis. Several studies have been conducted to simulate petroleum formation by pyrolysis, some of which used the marine alga Fucus sp. as the base material. Recently, activated sludge and fungi were converted to oily substances at relatively low temperatures as compared with those used in previous experimental simulations. On the basis of these findings, it is assumed that algae grown in CO₂-enriched air can be converted to oily substances, and that such an approach can contribute to solving two major problems: air pollution resulting from CO₂ evolution, and future crises due to a shortage of energy sources. Use of thermochemical liquefaction of organisms in the production of alternative fuels, would reduce CO₂ evolution into the atmosphere since such fuels would indeed be produced from CO₂.

Apart from the experimental simulation discussed above, other work has also been conducted with the objective of producing fuel from microalgae. Feinberg (18) reported that diesel fuel and gasoline were produced through the transesterification and catalytic cracking of lipids accumulated in algal cells. However, the raw material utilized in their work was restricted to microalgae of high lipid content. A process for the production of fuel oil from microalgae by pyrolysis has been proposed. The pyrolysis usually requires a drying procedure in which large amounts of energy are required to vaporize water. An alternative technique involving the direct thermochemical liquefaction of biomass of high moisture content, such as wood and sewage sludge, has been proposed and applied to the production of fuel oils from microalgae.

This liquefaction is carried out in an aqueous solution of either alkali or NaCl at a temperature of about 300 C and pressure of 10 MPa in the absence of reducing gases such as hydrogen and/or carbon monoxide. Since drying is not required, energy consumption for water vaporization is avoided. Microalgal cell precipitates derived from centrifugation, which are of a high moisture content, are thus good raw materials for liquefaction.

6.3.2 Cultivation of microalgae

The cultivation and liquefaction of microalgae has been recently investigated (19,20). The green microalga Dunaliella tertiolecta ATCC 30929 was cultured in a medium of the following composition: NaCl 58.5g; MgCl₂•6H₂O 1.5g; KNO₃ 1.0 g; MgSO₄ •7H₂O 0.5 g; KCl 0.2g; CaCl₂ •2H₂O 0.2 g; NaHCO₃ 43mg; KH₂PO₄, 40.8 mg; K₂HPO₄ 0.495 g; FeCl₃ solution 1.0 ml; metal solution 1.0 ml. The FeCl₃ solution consisted of the following (per liter): FeCl₃ 0.03 g; EDTA •2Na 5.84 g. The metal solution consisted of the following (per liter): H₃BO₄ 0.61 g; MnCl₂ •4H₂O 23mg; ZnSO₄ •7H₂O 87mg; CuSO₄ •7H₂O 0.06 g; (NH₄)₆Mo₇O₂₄ •4H₂O 21mg; CoCl₂ •5H₂O 15mg; EDTA •2Na 1.89g. The main culture was conducted in 3 L of medium at 27°C, using a 5-L fermenter (Fig. 6-3). The aeration rate and speed of agitation were maintained constant at 1 vvm and 200 rpm, respectively. Sparging gas was produced by mixing CO₂ with air at a fixed ratio of between 0 and 0.1. Four fluorescent lamps were used for external irradiation of the fermenter; the degree of irradiation at the inner surface of the fermenter vessel was 10,000 1x.

Algae were also cultured in 20-L box-type water tanks (366 x 216 x 250 mm) containing 12 L of medium as illustrated in Fig. 6-4. The medium was not sterilized prior to cultivation. The aeration rate was maintained constant at 0.25 vvm and the CO₂ content of the aeration gas was fixed at approximately 3 %, throughout the culture experiment. The water tanks were placed either in a bio-photo chamber (max 20,000 Ix, type LX-2100, Taitec Co. Ltd., Tokyo), or on shelves with fluorescent lamps (Iwasaki Co. Ltd., Tokyo), enabling the irradiation intensity to be altered, as shown in Fig. 6-4. The initial pH of the culture broth was adjusted to 8.0 in the main culture. The innoculum volume of 400 ml, was equivalent to 3.3% of the working volume of the main culture.

The effect of NaCl concentration on growth was examined using 5-L jar fermentors. Optimal growth was obtained at an NaCl concentration of 1.0 M. In order to ascertain the effect of contamination under saline conditions, algae were also cultured at a 1M NaCl concentration in unsterilized box-type water tanks containing unsterilized medium. The vessels were not airtight, thus exposing the culture broth to contaminating microorganisms. D. tertiolecta grew to levels of 1.2 g/L and was not affected by the non-sterility of growth conditions.

Figure 6-5 shows cell growth under 5,000 and 10,000 Ix illumination in the fermenter. Growth under 10,000 Ix was almost twice that under 5,000 Ix, indicating that growth is limited by insufficient intensities of light irradiation. Light intensity is thus an important factor in scale up of the cultivation process. Cell growth within the fermenter occurred normally within a CO₂ concentration range of 3 to 10 % (Fig. 6-6). However, limited cell growth occurred at very low CO₂ concentrations (0.03% CO₂).

6.3.3 Liquefaction of microalgae

Liquefaction was performed using a conventional stainless steel autoclave of 100-ml capacity with mechanical mixing (Fig. 6-7). The autoclave was charged with algal cells (about 20 g), following which nitrogen was introduced to purge the residual air. The nitrogen pressure was then elevated to 3 MPa, in order to prevent water present from vaporizing. The reaction was initiated by heating the autoclave to a fixed temperature, using an electric heater. The temperature of the heated autoclave was maintained constant for a 5 to 60 minute period, following which it was cooled with the use of an electric fan.

Figure 6.3 – Fermentation apparatus (5 L fermentor) used in the culture of microalgae

The separation procedure is schematically presented in Figure 6-8. The autoclave was opened, and the reaction mixture which was removed for separation and analysis consisted of a tar-like material, floating on the surface of a water phase. This reaction mixture was extracted with dichloromethane in order to separate out the oil fraction. The dichloromethane extract was filtered from the reaction mixture, following which residual dichloromethane was evaporated at 35/C under reduced pressure, yielding a dark-brown viscous material (hereafter referred to as the oil). The aqueous phase resulting after dichloromethane extraction (i.e. the dichloromethane insoluble fraction) was washed with water and filtered, in order to remove insoluble dichloromethane. The product yield in thermo-chemical liquefaction was thus calculated using the following equation:

Figure 6.4 – Box-type vessel (20-L capacity) for the culture of microalga

Figure 6.5 – Effect of light intensity on cell growth within a fermentor

Figure 6.6 – Effect of CO₂ concentration (as a sparged gas) on cell growth within a fermentor

Figure 6.7 – Schematic of autoclave equipment used for thermo-chemical liquefaction reactions

A heavy oil yield of 35.6% was obtained. This heavy oil consisted of carbon (73%), hydrogen (9%), nitrogen (5%), and oxygen (13%). The heating volume of the heavy oil was 34.7 kJ/g, which is almost the same as that of C heavy oil. This heavy oil had a viscosity of 860 cps, which was similar to that of castor oil. This heavy oil was of a higher nitrogen content than ordinary petroleum, thus necessitating requirements for flue gas treatment in order to prevent the formation of NOx.

These experimental results indicate that oil can be produced from CO2 gas using microalgae. However, two problems remain to be solved before such a system can be put to practical use. The first is the existence of a severe growth limiting factor, namely, light intensity. The effect of light intensity on growth is clearly shown in Figure 6-5; light could not penetrate deeply into the culture broth, owing to its obstruction by the microalgal body. In order to scale up algal cultivation while maintaining good growth levels, an extremely powerful light source and/or large surface area for irradiation will be required. The second problem is the high nitrogen content of the heavy oil which could be a potential source of Nox synthesis if the oil were used as a fuel. These two problems need to be addressed in the scale-up of CO₂ fixation and/or oil production using microalgae.

6.4 Algal hydrogenation

A hydrogenation process which permitted the conversion of algae (Chlorella pyrenoidosa) to liquid products (21) was also developed. Algal hydrogenation was performed batch wise, using an autoclave under high temperature and pressure conditions in the presence of a catalyst and a solvent. Major process variables in algal hydrogenation include reaction temperature, hydrogen pressure, reaction time, the nature of the catalyst, and process solvent. The effect of temperature in the reactor was investigated between 340 and 430°C, and reaction times were varied from 0 to 210 minutes. Algal hydrogenation is a three-phase operation in which contact must be established between the gaseous phase (hydrogen and hydrocarbon gases), liquid phase (mixture of solvent and liquid products), and solid-particle phase (algae and catalyst) in order to achieve algal conversion and to promote the transfer of momentum, heat, and mass.

A stirred slurry reactor, illustrated schematically in Figure 6-9, was employed to carry out the three-phase operation. The gaseous reactant was bubbled through the liquid from a sparger at the bottom of the reactor, and the solid particles slurried with the liquid were fed to the reactor. The gaseous reactant and the solid reactant (if any) first dissolve in the liquid phase and diffuse with the liquid reactant towards the catalyst. All these reactants then interact at the surface of the catalyst. The entire system was mechanically agitated by an impeller in order to enhance the rates of heat and mass transfer, to disperse bubbles throughout the agitated liquid phase, and to keep solid particles suspended in the liquid medium. The liquid products, solid particles and unreacted materials were continuously drawn off to maintain a constant liquid level within the reactor, and a constant composition in the liquid phase. Gaseous products and unreacted gaseous reactant entered the space above the liquid surface and flowed out through a gas pipe.

Figure 6.8 – Separation scheme for liquefied microalgal cells

Algae can be converted to liquid hydrocarbons at temperatures between 400 and 430/C, and operating hydrogen pressures of 1025-2250 psig, in the presence of a cobalt molybdate catalyst. The highest oil yield obtained was 46.7 wt % on the basis of algae charged. In addition, up to 10 wt% liquid products, and 34 wt% hydrocarbon-rich gases were obtained. In general, higher temperatures and longer reaction times increase the degree of conversion and oil yield and decrease the asphaltene yield in the overall hydrogenation of algae. The oil yield and the degree of conversion also increase proportionally with hydrogen pressure to a maximum of about 1200 psig, and then level off. A catalyst is required for the algal liquefaction process, and the yield distribution from algal hydrogenation is greatly influenced by the nature of the catalyst employed. The algal hydrogenation process offers a useful means of producing liquid hydrocarbons for use as fuels, feedstocks, and chemicals.

6.5 Future prospects

Petroleum supplies will be exhausted in the future, and the development of technologies for mass production of petroleum alternatives is desired. Mechanisms of crude oil formation by natural phenomena have been partially elucidated and technology for crude oil synthesis has been developed. Several studies relevant to the production of oil using microalgae have been reported. These include hydrocarbon production by Botryococcus, thermochemical liquefaction of microalgae, and algal hydrocarbon processes, all of which were introduced in this chapter.

Petroleum is not only used as fuel, but is also a raw material for the production of a variety of chemicals. Petroleum alternatives should be developed prior to the exhaustion of petroleum supplies. The majority of petroleum has its origins in algae, which were grown using CO; as a sole carbon source. Research into the production of petroleum alternatives using microalgae is important to the future of mankind.

Figure 6.9 – Apparatus used for hydrogenation within an autoclave

References

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