Optimization of a Novel Algal Oil Harvesting Strategy for Renewable

BiofuelsJames Madison University

May 2016

Written by: Hannah Aloumouati, Victoria Foster, McKinnon Langston and Alexander Macfarlane

Advisor: Dr. Chris Bachmann, Ph. D

Table of ContentsAbstract.........................................................................................................................................3

1.0 Introduction..............................................................................................................................4

1.1 Petroleum Dependence...........................................................................................................4

1.2 Algae-Based Biofuels............................................................................................................10

1.3 Environmental Concerns......................................................................................................11

1.4 Objectives.............................................................................................................................14

2.0 Methodology...........................................................................................................................15

2.1 Algae Cultivation..................................................................................................................15

2.2 Harvesting...........................................................................................................................19

2.3 Separation of End Products..................................................................................................24

2.4 Algae Oil Purification and Solvent Reclamation.....................................................................26

2.5 Algae Oil Quantification.......................................................................................................27

2.6 Determination of Energy Input vs. Output (need kilowatt meter photo)....................................28

2.7 Mathematical Modeling of Theoretical Algae Oil Yields.........................................................29

2.8 Cost Analysis........................................................................................................................29

3.0 Results....................................................................................................................................29

4.0 Discussion...............................................................................................................................34

4.1 Unintended Consequences....................................................................................................38

5.0 Conclusion..............................................................................................................................38

6.0 Current and Future Direction of Research..............................................................................39

References....................................................................................................................................42

Abstract

The United States Environmental Protection Agency has determined that the transportation sector was responsible for 27% of U.S. Greenhouse Gas Emissions in 2013. To reduce emissions released into the atmosphere, The U.S. Department of Energy (DOE) is funding research for “Advanced Biofuels” that are less harmful to the environment. The use of algae as a biofuel feedstock has attracted considerable attention because of its potential to provide abundant, clean, renewable fuel that does not interfere with the global food supply. The DOE’s National Algal Biofuels Technology Roadmap identifies two main problem areas that need to be overcome for algae biofuels to become economically viable: large-scale algae production and algae harvesting. This project focuses on the optimization of a novel, energy efficient and cost effective algae harvesting method. This method was developed through a public-private partnership between James Madison University and Wholesome Energy of Edinburgh, VA. The process uses extremely high shear mixing in the presence of a powerful non-polar solvent to breakdown algae cell membranes and facilitate migration of oil into the solvent phase. The algae oil is then purified from the solvent and the solvent is recycled back into the extraction process. Preliminary data for the harvester shows approximately 29,500 gallons of seawater culture media are needed to produce 1 gallon of algae oil. At current operating conditions, this small scale harvester would need to run for 28.5 days continuously at a cost to harvest at $0.31 per day, yielding a total cost to harvest the raw algae oil at $6.08 per gallon. Despite this process’s costly production, further investigations to improve the overall energy and cost efficiency are currently underway. This algae-oil harvesting strategy could potentially be used to facilitate the cleanup of harmful algae blooms in the Chesapeake Bay by providing an economic incentive for algae collection. When combined with offshore algae cultivation, this system has the potential to provide abundant clean renewable fuel not only to Virginia, but also to other parts of the United States. This results in significant reductions in harmful greenhouse gas emissions from the transportation sector.

1.0 Introduction

As the world population steadily increases, the rate of consumption steadily increases as well [1]. Consumption is inevitable. The consumed resources cradled from the environment range from simple food products to highly energy intensive fuels. A major stepping stone in today’s society is the need for an abundant clean energy resource that can supply the demand of the current population without impairing that of future generations [2]. Something to consider with this is that the economic feasibility of an energy source is a heavily weighted parameter in the widespread adoption of a fuel. Other parameters include social acceptance, molding to current infrastructure, stakeholders, environmental soundness, and others. Current energy resources are in favor of fuel powering sources like coal, oil, and natural gas that can each have detrimental impacts on the environment and impede the societal and environmental development of future populations. Fossil fuels are low cost, abundant, easily accessible, and have more than 100 years of development behind their extraction, refinement, distribution, and end-use. There aren’t many options that can compete, or even be able to meet demand, let alone be cost competitive. Despite the consequences learned from the utilization of these fuels, accessibility, low cost and infrastructure allow its use to continue. An intermediary solution to this could be the use of algae microorganisms to produce biodiesel that can be utilized as a fuel source, particularly the transportation sector [3]. Not only does this fuel source provide the same energy content as that of diesel fuel, but it also is almost a carbon neutral source, it does not disrupt the global food supply to produce, it has a doubling rate of about 48 hours for constant production, and most importantly, it fits the current liquid fuel infrastructure already established in the United States. This report assesses the promise and opportunity algae biodiesel has to offer for a sustainable development.

1.1 Petroleum Dependence Research of algae as a fuel source has been under development since the 1920s and currently, the 2010 National Algae Biofuels Technology Roadmap identifies algae cultivation and harvesting as two main problem areas with algae biofuels [4]. This thesis specifically focuses on optimizing a novel harvesting method for algae crude oil. It’s the United States addiction to oil that is preventing the prosperity of an alternative fuel source that is better environmentally. Oil has not always been the staple fuel source utilized predominantly in the United States by various end-use sectors. Coal was once the primary fuel to supply energy to each sector, and currently it is most widely used to generate electricity [5]. Coal’s abundance and known attributes enabled its progressive use throughout the United States. The addiction to oil sprang from the Industrial Revolution, where a cheaper, more accessible fossil fuel was needed to substitute coal [6]. Oil became this primary fuel source for many reasons. It has become the main source for fuel in civilization for reasons such as its social needs and demands, influence of corporate stakeholders, the physical traits of oil, and the dominance of oil as the premier energy source for the transportation sector. Around the mid-1800s oil became the fossil fuel that everyone began to try and locate. Due to this need the oil industry started to really succeed in the mid to late 1800s. Edwin L. Drake was hired by an American entrepreneur named George Bissel to locate this oil [6]. Drake was hired, by Bissel’s Pennsylvania Rock Oil Company of New York, to drill for oil near a well-known oil seep on Oil Creek in Titusville, Pennsylvania [6].The refined

crude oil extracted from the ground began to produce a byproduct called Kerosene [7]. One of the first American uses for oil was this byproduct [7]. Kerosene was used during the American Civil War from 1861-1865 to provide light and heat in the form of a gas lamp [7]. Thomas Edison’s invention of the light bulb soon impacted the use of Kerosene for light [8]. This allowed the paradigm to shift from oil being used as a light source, to oil being used for gasoline. As the next few decades approached, an entrepreneur by the name of John D. Rockefeller began to understand the need for oil and its impact on the American economy [8]. At the age of 16, John D. Rockefeller began to recognize the importance and dependence of oil and its impact on the American economy [9]. He was a bookkeeper for a produce company and began to save his money for future investments into the prominence of oil [9]. In 1859 Rockefeller invested into a Cleveland based oil refinery [9]. By 1865 Rockefeller had begun to recognize the successful growth of this refinery and he proceeded to take out a loan to buy out his partners [9]. Rockefeller now controlled the largest oil refinery in Cleveland [9]. In the coming years, Rockefeller expanded his entrepreneurship ideas in the oil industry. In 1870 he started his own company, Standard Oil Company. Standard Oil Company used the ability to market, produce, transport, and refine petroleum better than any competitor [9]. By 1882 Rockefeller's Standard Oil Company became a monopoly to the oil market [9]. Americans felt this threat of the monopoly and took government action to address the situation. The federal government passed a legislative act called the Sherman Anti-Trust Act in 1890 [10]. This act was passed by the United States Congress and gave them the ability based on the constitutional power of Congress to regulate interstate commerce [10}. The Sherman Anti-Trust Act was used against Rockefeller’s company and through many legal battles, Standard Oil Company began to dissipate. In 1904, Standard Oil Company still controlled 91% of oil production and 85% of its final sales [11]. It took until 1911 that Standard Oil Company was completely dissolved by the Sherman Anti-Trust Act into 34 smaller companies [9}. Companies that are around today were created due to this act such as the following: Amoco, Exxon, Marathon Oil, and Mobile. Around the 1800s, when a more effective and faster transportation method was needed to stray away from the horse and buggy, three main fuel types emerged: steam, electric, and gasoline {12}. Each of these vehicle types had the chance to prosper in the United States but the gasoline/oil based engine dominated because it was cheap abundant, reliable, was faster to refuel, and range anxiety wasn’t an issue compared to the other fuel types. The internal combustion engine forever changed the demand for coal fuel supplies as well [13]. Entrepreneurs began to see the opportunity in gasoline and oil. In the 1900s, Henry Ford established his Ford Motor Company, and five years later, the company released their Model T gasoline engine vehicle [14]. Before Ford, cars were seen as a luxury to Americans where only wealthy people could afford them. Once Henry Ford constructed and mastered the assembly line to mass produce his Model T product, the demand for the cars steadily rose, bringing the price of this vehicle down. This process provided the gasoline/oil internal combustion engine with a very advantageous position in the market, wiping other fuel types from prospering as greatly. Due to the influence of John D. Rockefeller and Henry Ford businesses, the American oil industry began to grow rapidly. Since Henry Ford’s Model T had an internal combustion engine, the oil market continued strong into the early parts of the 1900s. Ford’s assembly line and production of car changed the landscape of the oil industry. Cars were becoming more prevalent and the supply and demand for oil needed to be

met somehow. An accommodation for this need had to be met to address this rapid growth. American oil companies continued to grow and the need for an American infrastructure to support it had to be met. Not only were there oil wells and companies popping up everywhere, the ability to ship and transport this crude oil needed to be met. By the advent of World War I, crude oil pipelines were starting to transverse the United States [15]. Driven by the rapid growth of the automobile industry, pipelines were starting to form the foundation of the oil infrastructure. In 1920, there were approximately 115,000 miles of oil pipelines across the country [15]. Pipelines began expanding westward in the 1930s due to population growth west of the Mississippi. Throughout World War II the pipeline infrastructure grew rapidly. The United States had 48 oil tankers sunk during this war, mainly along the eastern seaboard [15]. America addressed this by creating land based pipelines that began to pump oil from states such as Texas and Oklahoma [15]. These were large-diameter pipelines carrying crude oil to East Coast consumer states [15]. This foundation infrastructure laid back then is still the current one we use today, with some modifications to fit current technology [15]. In 2015, according to the United States Department of Transportation, there was over 1.7 million miles of gas and oil pipelines [16]. Everything in this oil industry, down to the gas pumps that put gas in our vehicles, is a direct correlation of the global influence of oil as a fuel source. The fuel source’s base product is oil, allowing the industry to continue its growth. Up until 1910s, the United States produced between 60-70% of the world’s oil supply [17]. With the large supply, there was concern for the U.S. supply being diminished. In 1908 Theodore Roosevelt brought this concern to the forefront and appointed the National Conservation Commission (NCC) to take an inventory of America’s natural resources and to discover areas where egregious waste was occurring [18]. When the NCC delivered its findings to Congress in 1909, it reported that the nation’s petroleum supply would last a mere 25 to 30 more years [18]. This generated a fear throughout America, forcing them to turn to foreign countries. As Americans were scared the nation’s petroleum supply was diminishing, countries like Mexico, Iraq, Iran, and Venezuela were beginning to discover more oil. Shortly after this in 1924, Texas, Oklahoma, and California experienced an oil boom that mitigated America’s oil scare [19]. This oil boom actually brought oil pricing plummeting down to ten cents a barrel and regulations had to be set to raise these prices back up. From this stemmed OPEC (Organization of Petroleum Exporting Countries), which was created to help manage oil exportation and prevent oil prices from ever plummeting again. In 1954, Marion King Hubbert, working for Shell Oil at the time, developed a peak oil curve for the United States oil production [20]. This curve incorporated cumulative oil production, proven oil reserves and future discoveries all as a bell shaped curve shown in the figure below. From his findings, he predicted that the United States oil production would peak in the year 1970 [20]. During this time, a lot of people argued back and forth on whether or not the U.S. would run out of oil. Hubbert was just another opinion to add to this pot, so his findings were not taken seriously. This time period also saw a popularity of the muscle cars and gas guzzlers in America. With the combination of these vehicles and rates of consumption increasing, the U.S. did in fact reach a peak in their production in the year 1970 as Hubbert predicted. Three years later, the U.S. experienced its first oil crisis.

The 1973 Oil Crisis is most widely known as the Arab Oil Embargo. On October 6th, 1973, Egypt and Syria launched a surprise military campaign against Israel, and six days later, the U.S. provided aid to Israel. In response to the U.S. supply Israel with aid, OPEC, consisting of Saudi Arabia, Iran, Iraq, Abu Dhabi, Kuwait, and Qatar at this time, announced an oil embargo against Canada, Japan, The Netherlands, United Kingdom and the United States [21]. This raised the price of oil by about 70%, sending these countries into an oil shock [21]. When the U.S. could not manage any longer, the Nixon Administration negotiated and convinced Israel to withdraw its forces and pull back [21]. Shortly after, the Arab Oil Ministers lifted the embargo in December 1974, ending the crash of the stock market [21]. After the oil crisis, Hubbert continued his research on oil production but instead, he analyzed a global perspective. From his research, Hubbert then predicted that there would be a global oil production peak in 1995 if the current rate of consumption continued [20]. Soon after this prediction was released, a second oil crisis erupted in the United States. The 1978 oil crisis was the results of a decreased oil output from its foreign imports in the wake of the Iranian Revolution [21]. The global oil supply only decreased by about 4% but the widespread panic in the U.S. is what drove the price per barrel far higher than justified by the supply. The price per barrel more than doubled and odd-even rationing had to be implemented in the states [21]. Odd-even rationing meant that Americans could only get gas on certain days of the week based off of their license plate number. Sometimes they were even limited by how much gasoline they could purchase. This crisis clearly distinguished the United States vulnerability to foreign oil supply. A management council was needed to promote U.S. energy independence by further diversifying its energy portfolio. The ideas of creating a department to address this energy portfolio dated back to before World War I. Albert Einstein helped promote these ideas by handwriting a letter to President Theodore Roosevelt explaining the importance of nuclear chain reactions and the possibility that it might lead to the

Figure 1.1.1: Hubbert’s peak concept is demonstrated by this figure. This image illustrates the concept of increased oil discovery, followed by increased oil extraction, then a rapid decline in production as oil reserves are depleted at a maximal extraction rate. Image by www.agorafinacial.com

production of powerful bombs [22]. Many energy programs were created to address this and projects such as the Manhattan Project was formed in late 1941 [22]. The Manhattan Project was the codename for an American project that had aspirations of designing and building the first atomic bomb. Projects such as the Manhattan Project continued to arise and be completed with no form of federal government policy. This became something that America needed to address and President Jimmy Carter took this idea seriously. He was inaugurated President of the United States in January of 1977, and addressed the need to put Federal energy activities under one umbrella and provided the framework for a national energy plan [22]. Through his efforts, The Department of Energy was formed in August of 1977 [22]. This department combined most of these government energy programs, but also addressed defense responsibilities that include the design, construction, and testing of nuclear weapons. The Office of Fuels Development was one of the government energy programs that now fell under the Department of Energy umbrella. They wanted to create a program to develop renewable fuels for transportation from algae. The end result of this want was the Aquatic Species Program being started in 1978 [3]. The main focus of this program was to achieve the production of biodiesel from high lipid-content algae grown in ponds while utilizing waste carbon dioxide from coal fired power plants [3]. Researchers in the Aquatic Species Program wanted to identify specific algae strains that had high lipid content along with being able to survive in severe conditions [3]. The conditions that concerned them the most were the temperature, pH, and salinity of the water. The researchers collected over 3000 strains of algae across the continental US and Hawaii [23]. After screening, isolation, and characterization efforts the collection was reduced to 300 strains [23]. Between 1978 and 1982 the research efforts of this program were designated to use the algae strains to produce hydrogen [3]. The program then switched its efforts to transportation fuels and more specifically biodiesel for the rest of its existence. Funding began to become difficult for this program due to the federal government’s idea of downsizing. Not only did funding play a vital role on the Aquatic Species Program but so did the price of oil. In the 1980s and 1990s oil prices began to fall and the need to find an alternative fuel source was not at the forefront of Americans minds. In 1996 the Aquatic Species Program was shut down closing a program that once seemed so promising [3]. The program concluded that the production of microalgae used for fuels is not limited by engineering designs, but by the many microalgae cultivation issues [3]. The issues ranged from species control in large outdoor systems, to harvesting and lipid accumulation, to overall productivity [3]. Once this program was shut down, the focus on the bioethanol sector began to grow. At the turn of the century, oil prices were still low but due to political issues, they were steadily rising. The U.S. was importing about 75% of its total oil consumption and its oil production was declining, which is shown in the graphical figure below generated by the EIA.

The United States saw this as an energy security issue and knew they were still vulnerable to foreign oil supplies. At this time, Iraq had the second largest oil reserve in the world behind Saudi Arabia, whom was America’s ally. In 2003, the U.S. invaded Iraq ultimately because the U.S. was fearful of Iraq possessing weapons of mass destruction and the public perception of Iraq having an influence on the attack on September 11, 2001. One of the primary goals of this invasion was to secure Iraq’s oil fields and resources by ending the regime of Saddam Hussein. The U.S. wanted to give Iraq’s oil reserves back to the Iraqi citizens to not only promote the economic health of Iraq, but also create an ally in them as well. In the meantime, the U.S. reliance on foreign fuels increased with their position in war. This brought about the 2005 Energy Policy Act that was passed on July 29th by Congress and George W. Bush [24]. Some general provisions of this act included exempting all fluids used in the natural gas extraction process from the protections under the Clean Air Act, Clean Water Act, Safe Drinking Water Act, and CERCLA, it increased the amount of biofuels (usually ethanol) that must be mixed with gasoline sold in the U.S., it seeks to increase coal as an energy source while also reducing air pollution, authorizing $200 million annually for clean coal initiatives, it gives a tax credit to renewable energy producers, and as well as many other provisions [24]. One of the provisions of this act was for the U.S. to have 25% of renewable energy used by end-users in the U.S. by the year 2025 [24]. This was a goal and not necessarily a mandate, but agencies like the Department of Navy and the Department of Energy aggressively wanted to achieve this goal. This facilitated the establishment of a Renewable Energy Strategy that helped promote research and adoption of renewable energy sources. This act also influenced a larger production of ethanol, stimulating 27 new ethanol plants, 401 new E85 gas pumps installed, and in 2006 1.4 billion barrels of ethanol was produced [25].

Figure 1.1.2: This is a graph of U.S. crude oil production and oil imports for the U.S. generated by the EIA. Notice how the production in the U.S. steadily declines due to supply issues and imports increase. Then production increases around the same time hydraulic fracturing takes off and imports steadily decline.

The production of ethanol skyrocketed, while it also met its intended goals. This contributed to EISA, the Energy Independence and Security Act of 2007. This mainly encouraged a higher fuel standard for fleets of vehicles, more efficient home appliances, and it also set up regional standards for heating and cooling throughout the United States [26]. The stated purpose of this act was to “...move the United States toward a greater energy independence and security…” [26]. The original bill sought to cut subsidies on the petroleum industry in order to promote petroleum independence, but this was quickly dropped due to the industry’s stakeholders and influence. Instead the Renewable Fuel Standards was adopted, requiring transportation fuel sold in the U.S. to contain a minimum of 36 billion gallons of renewable fuels, including advanced and cellulosic biofuels and biomass-based diesel by 2022 [27]. Since ethanol production was already increasing and meetings its intended goals, stimulating its production even further was not an issue. Today, 93% of the transportation sector is dependent on oil use, 70% of all oil products goes to transportation and about 36% of the total energy demand is met from oil in the United States [28]. Since the overall need and use of oil has increased throughout the years, the United States is working diligently to increase their energy independence by finding more sources of oil nationally. There are over 4,000 offshore drilling rigs in Texas, Alaska, and California, making up approximately 35% of the total domestic oil production in the United States [29]. For the United States to achieve energy independence from foreign oil imports, the total domestic production must exceed that of the total imports. Looking closer at the U.S. and global oil supply, we have a larger supply than ever before. The image below was taken from a BP Statistical Review, where you can clearly see the supply increasing over the years. There is an even bigger supply jump from the year 2001 to 2011. This can largely be attributed to the hydraulic fracturing extraction process. This is commonly used to extract natural gas, but it is also just as effective to extracting oil supplies. Since the 2005 Energy Policy Act exempting all fluids used in this process from regulation, this process became a lot more enticing to utilize for oil harvesting. Now that the U.S. and world has a large supply of oil, the main reason to switch to an alternative fuel isn’t a matter of a diminishing supply, but more so a large concern for the health of our environment and its implications on future generations.

1.2 Algae-Based Biofuels

With the ability to adapt to circumstances along with its rapid growth and doubling rate, algae was used for many research efforts. One of the early use by American researchers of algae, was to provide a food source. In 1948, researchers had the idea that algae could end world hunger. After World War II there were impoverished and malnourished people across the world. Researched used Chlorella as this strain of algae to end world hunger. When grown in optimal conditions—sunny, warm, shallow ponds fed by simple CO2—Chlorella converted around 20% of available solar energy into plant biomass containing over 50% protein when dried [30]. The problem with Chlorella is that it was a very temperamental strain of algae that was not sustainable. Also the thick cell walls of Chlorella made it indigestible without a very cost and energy intensive mechanical process [30]. The research gave up their push to end world hunger and algae began to be used to produce other things such as a gas and biofuels [30].

The production of methane gas from microalgae dates back to 1950. A professor at the University of California-Berkeley used microalgae to produce methane gas by anaerobic digestion, better known as

fermentation [31]. The energy crisis in 1973 gave the research of microalgae into gaseous fuels (hydrogen and methane) momentum that grew the idea of turning microalgae into biofuel [31]. The Department of Energy (DOE) was formed in August of 1977 and helped microalgae research thrive. The Aquatic Species Program was started under the DOE in 1980 and research efforts were used to produce algae based biofuels for the transportation sector [3]. Initial efforts were to produce hydrogen gas, but then quickly switched to transportations fuels and more specifically biodiesel [3]. Once this program was shut down in 1996, the private sector for algae research needed to take on the responsibility of using algae as a biofuel source. During the 1990s, the Japanese government supported a major program on algae for CO 2

capture and greenhouse gas abatement, with a budget of more than $250 million dollars [31].

Currently the private sector along with the DOE are starting to realize the need for an algae based biofuel. Companies such as British Petroleum and Exxon have put aside millions of dollars for research investments [31]. The National Algal Associations and the Algal Biomass Organization are two of the main research groups in America [31]. A huge need for an alternative fuel needed to be sought out since the transportation sector is responsible for 34% of all CO2 emissions in the United States and transportation sector uses oil [32].

1.3 Environmental Concerns While the fossil fuels reserves are continuing to expand as newer technologies make it possible to access fuel that was previously thought to be unattainable, the need for alternative fuels stems from the impact that fossils fuels has on our environment. The transportation sector is responsible for 34% of all CO2

emissions in the United States and transportation sector uses oil [32]. Though abundant, these fuels are anything but clean and are responsible for negative environmental impacts throughout their lifecycle. The production and use of oil has many external and indirect costs [33]. The first being the extraction of the oil. The ability to access locations of oil has become more difficult as previous wells have dried up. This has caused oil producers to take extreme measures to retrieve oil, in turn, increasing the risk of the process. The Deepwater Horizon oil leak is an example of this associated risk. Over 200 million gallons of crude oil leaked into the Gulf of Mexico, impacting aquatic life for thousands of shoreline miles and directly affecting the local economies [34].

Next is the transportation of the fuel. Only a few countries produce a majority of the world's oil which means most of the oil is exported. This creates opportunity for disaster to occur during its transportation. An example of this disaster is the Exxon Valdez oil spill in 1989. An oil tanker on route to California, collided with an Alaskan reef, causing 38 million gallons of crude oil to be spilled. This was considered to be one of the most devastating human-caused environmental disasters of its time, which now pales in comparison to the BP oil leak [34]. All of these are examples of issues with petroleum before its consumption. But when the oil is finally combusted, toxic chemicals and carcinogenic soot is released into the environment, leading to lung disease, lung cancer, asthma, and increased mortality rates [35]. These issues all contribute to the degradation of the environment, but the largest issue with fossil fuels is by far the excessive anthropogenic CO2 released into the atmosphere, eventually giving way to Global Climate Change.

Since the Industrial Revolution began around 1750, human activities have contributed substantially to climate change by adding CO2 and other heat-trapping gases to the atmosphere through the burning of fossil fuels [36]. Currently human activities release over 30 billion tons of CO2 into the atmosphere every year. 34% of these CO2 emissions come from the transportation sector alone [36]. These greenhouse gas emissions have increased the greenhouse effect and caused Earth’s surface temperature to rise [37]. 2015 was the hottest year in recorded history and Scientists at the National Snow and Ice Data Centre said that this March the sea ice cover reached the lowest winter maximum since records began in 1979 [38]. Not only is the increase in temperature melting arctic more than has been recorded. But this massive melting is predicted to lead to a six foot sea level rise by the end of the century [39].

Current consumption of fossil fuels has caused carbon dioxide levels to increase drastically, in turn increasing the amount of greenhouse gas present in Earth’s atmosphere causing global climate change. Regardless of the known consequences and effects of global climate change, very little legal action has been taken to restrict or limit use of fossil fuels. The matter of fact is that most people who use fossil fuels are aware of the environmental consequences but refuse to take action on a personal level. That is why creating a renewable fuel source that consumes the CO2 it creates from combustion is a feasible reason to implement it on a full scale.

Figure 1.3.1: This graph displays the rate of change of sea level that occurs per year and future projections. 52% of the nation’s population lives in coastal region so sea level rise is understandably of high concern to the United States [40]. Flooding in these communities on the coast could lead to a displacement of 13 million people in the US by 2100 [41]. This massive relocation would not only cause serious tension in the nation, but would also create monumental disruptions along the shipping trade routes [42].

In addition to consuming CO2, the areas where salt water algae can be grown help save terrestrial ecosystems from being destroyed. Due to its ability to be cultured offshore, algae does not require land to be cleared like certain energy sources, like solar power, do. Not only does this benefit the environment, but the offshore culturing of algae does not take away from any land being used for agriculture. While the impact of marine ecosystems may seem an issue at first, there are many dead zones in the ocean that have little to no known life existing there and these dead zones could be a safe environment to grow algae for biofuel. The increase in CO2 levels have led to an increase in such natural disasters as fires, droughts, hurricanes, floods, and acid rain. Acid rain kills fish, alters ecosystems, and decreases biodiversity [43]. It can also harm people through the atmosphere or through the food chain by having crops grown in the toxic soil, animals consumed by humans, and by drinking water [43]. The acidifications of the oceans due to CO2

levels has dropped by 0.1 units since the industrial revolution and is predicted to drop by a further 0.3–0.4 units by 2100 [44]. This has led to major destruction and bleaching of coral reefs [44]. Currently over 93 percent of the Great Barrier Reef is bleached [44]. Due to all the negative environmental impacts spurred by the usage of fossil fuels, an environmentally friendly fuel needs to be implemented.

The DOE and DON has already began research for several of these possible transportation alternatives but to insure the success of any alternative fuel, we must address the social and environmental impacts that may come with them. Below is a list of several possible alternative fuels as well as their advantages and disadvantages.

Figure 1.3.2: This chart compares several alternative transportation fuels that the Depart of Energy and Department of Navy have considered pursuing. Biodiesel has many advantages that make it a more feasible option than other alternatives.

Using algae oil as a transportation alternative poses many advantages that trump other options. There are disadvantages to using algae but we believe that with the continuation of research into algae cultivation and harvesting methods, algae oil will become a successful alternative.

1.4 Objectives

In order to produce an image for the acceptance of algae produced oils for fuel usage, it was necessary to review all relevant information currently dedicated for algae based biofuels studies relevant to this specific report. As this report analyzes the optimization of algal oil harvesting by manipulating various parameters within the emulsification process, the solution can be seen as a conformational change to the process itself or a need for change within the mechanics of the system. Based on the research, the latest developments and future direction of this research, basic knowledge of this subject and societal issues that will influence the successful adoption of the project are evaluated. Research efforts were started by recognizing the two main issues with algae biodiesel identified in the 2010 National Algae Biofuels Technology Roadmap. These issues were the harvesting and cultivation issues with algae [4]. The Department of Energy through the report: A Look Back at the U.S. Department of Energy’s Aquatic Species Program: Biodiesel from Algae, developed a list of algae strains that had high-lipid content thus allowing us to receive high oil yields [3]. Nannochloropsis was chosen as this strain. In continuing research, there had to be a proper technique to harvest this algae. The current methods were very energy intensive and required a time consuming dewatering step. This research effort focuses on a process that does not require dewatering. The emulsification process of the Nannochloropsis did not require dewatering and thus was chosen for the optimization effort on harvesting the algae. Emulsification is a process of high-shear mixing of n-Hexane with the saltwater algae volume that makes the algae cell walls break apart releasing an oil that is collected through a reclamation device. Many efforts to produce algae biodiesel have been sought out, but the cultivation and harvesting issues identified in the 2010 National Algae Biofuels Technology Roadmap deem too large of an obstacle to make this an energy and cost efficient task. This project sought to optimize the harvesting process of algae in order to achieve a higher and more cost competitive algal oil yield. The steps implemented to optimize the system were finding the ideal flow rate and a single pass system. An energy cost analysis was conducted on the harvester to understand the energy input. A single pass system was implemented to reduce the time the system was operating and therefore lower energy input. Regulating the input of the solvent n-Hexane was accomplished through introduction of a turn valve. The rest of this report includes methodology and results of the experiment. The goal of this experiment is to calculate the cost of the amount of energy obtained, to compare it to the amount of energy that was put into the system. In doing so, a series of lab experiments were conducted to obtain a fuel that could potentially be a more sustainable energy source then current practices. Despite sustainability being a very desirable factor, economic feasibility is a more important aspect within the adoption process of a new fuel source. To calculate the cost to produce this algae oil fuel source at this

maximum rate of production for a comparison to current market fuel sources, the harvester’s energy usage was recorded using a kilowatt reader during the harvesting phase.

2.0 Methodology

In order to obtain notable results, the follow methods were performed during the length of this project. Each major area of focus is divided into subsections to explain in detail how this project was conducted.

2.1 Algae Cultivation

To create and retrieve algae oil, our project uses three distinct processes, the first being algae cultivation. Before the processing of algae cells can occur, the culture itself must be grown to the adequate size, then maintained and monitored. Nannochloropsis was chosen because it has a high oil yield compared to other strains and is a species of saltwater algae which can be grown offshore to cultivate large volumes. Nannochloropsis requires a consistent environment to grow in and the optimal environment being an aqueous solution with a salinity of approximately 37 parts per thousand [45]. To achieve optimal salinity, Instant Ocean Aquarium Salt was purchased and mixed with onsite water. A salinity curve was made by a previous algae capstone group which provided our team with an equation to gage how many grams of salt was needed for a desired amount of water. Once the measured salt is added to water, it needs to be thoroughly mixed to dissolve. The salinity needs to be measured after mixing to ensure that proper amounts of salt and water were mixed, this is done by using a refractometer which gauges the refractivity of a solution based on its salt content Show picture of refractometer After the desired salinity is reached, optimal temperature must be reached.

The temperature of the algae is another factor that needs to be controlled. Algae can experience a “shock” when there is a drastic change in the surrounding environment. Large fluctuations of salinity or temperature can shock the algae, causing large amounts of algae cells to die-off. To maintain our cultures temperature, four aquarium heaters set to 75 degrees Fahrenheit were added to the culture tub. This temperature mimics the oceans equatorial temperatures to reassure healthy rapid growth of the algae. An important factor for algae growth is the strength and duration of light exposure. Since our culture was grown indoors to maintain a consistent environment, plant grow lights were needed. The previous algae capstone team under the advisement of Dr. Christopher Bachmann obtained grow lights, one being a 600 Watt Metal Halide light and the other a 1000 Watt High Pressure Sodium light.

To control light duration, two electrical timers were purchased and set to turn on for a 12 hour period every 24 hours. This time frame was chosen to maximize cell growth by imitating the light exposure algae would receive near the equator. Algae also requires agitation to thoroughly mix the cells exposing them to necessary nutrient and carbon dioxide. Three aquarium pumps were purchased and placed inside our culture tub to properly mix the water. These pumps naturally collect algae cells over time and need to be cleaned once a month to reduce risk of becoming defective. After a controlled environment has been set up, algae cells can be introduced. To obtain the culture of Nannochloropsis algae, our team purchased several 100 milliliter vials from Carolina Biological.

Figure 2.1.1: A 600 watt metal halide and 1000 watt high pressure sodium light are used to grow our algae cells. They are controlled by timers to turn on for 12 hours daily.

Figure 2.1.2: Nannochloropsis algae is grown by Carolina Biological and available for purchase. Our team purchased several 100 milliliter vials to begin algae cultivation.

This amount alone will not be enough to extract a significant amount of oil. To harvest the algae, four gallons of algae culture is needed and to run a trial, twelve gallons is required. Once we received the algae it needed to be grown to a larger size which we deemed “size up”. Nannochloropsis cells double every 48 hours, which allows for a size up to be done every 48 hours. A size up consists of two factors, saltwater and nutrients. The saltwater is a medium for algae to grow in, allowing new cells to populate it, while the nutrient provides necessary resources for cells to grow and divide. To double the algae culture volume, equal parts salt water and algae culture must be mixed with appropriate amounts of nutrient. This size up process was done every other day until we cultured 120 gallons of saltwater algae. This amount allows our team to process several runs of algae a week and any algae removed for processing is regrown by sizing up.

To ensure the health of our 120 gallon culture, an automated top-off system was created. Fresh water constantly evaporates off the saltwater culture causing the salinity of the culture to increase. The top-off system we implemented uses a 55 gallon freshwater container connected to a buoy in our culture tub. When the desired water level drops the buoy drops and freshwater is released. The top-off system keeps the salinity of our culture consistent and reduces required maintenance.

Once a week the culture was fed Kent Marine Pro Culture Part A and Part B. This nutrient recommended that for every 20 gallons of algae, 10 milliliters of Part A and Part B were needed. For our 120 gallon culture, 60 milliliters of nutrient needed to be added once a week. To improve consistency and reduce risk

Figure 2.1.3: This top-off system provided freshwater to our culture to reduce risk of salinity shock. Algae nutrient was later added to the top-off to reduce nutrient shock. The left outlet valve leads directly to the culture and the center outlet valve was used to flush the system.

of culture shock, the nutrient was added to the top system. Every week 7 gallons of water would evaporate from the culture which the top-off system replenished. Since our top-off system could hold about 55 gallons of water, or 8 weeks of evaporated water, we added 8 weeks of nutrient supply.

A cell count was performed weekly to observe the algae’s cell concentration. Fifteen microliters of algae is pipetted onto hemocytometer slide under a light microscope and using 40x objective, the cells are counted using the slides grid. This gave our team an accurate representation of the cell concentration and overall health of the culture.

Figure: 2.1.4: Kent Pro-Culture part A and B was used to provide nutrients for our culture. The culture was fed 10 milliliters of both parts for every 20 gallons of algae. Part A contains minerals such as iron and Part B contains macro nutrients such as nitrate.

Figure 2.1.5: A cell count conducted on a hemocytometer slide is shown above. The 4 by 4 grid was used to determine cell density. The average of 5 different 4 by 4 grids was taken to more accurately determine cell density.

Once the culture was grown to proper volume and cell concentration, processing could occur. To safely operate the equipment and handle n-Hexane, Standard Operating Procedures were created. These procedures were created to provide a clear step-by-step process of how our team would retrieve algae oil. Familiarization with the SOP for this process is mandatory and ensures safety, accurate measurements, and reproducible results. The S.O.P. procedures were conducted for JMU’s safety marshal and received approval.

2.2 Harvesting

Previous biofuel teams under the advisement of Dr. Christopher Bachmann used flocculation harvesting to concentrate algae cells then dewater them for oil extraction. These methods as well as many other methods, are time consuming and energy intensive. This bottleneck has restricted algae oil from becoming a successful alternative thus far and before algae oil can be considered as the replacement for fossil fuels, the harvesting process must produce profitable energy, on a large scale, at a fast rate. Emulsification was considered as a possible alternative harvesting method that would avoid the need to concentrate cells and dewater them all together. The process of emulsification mixes two or more liquids that are normally unmixable or unbendable, such as oil and water. For our harvesting process we used emulsification to mix algae and hexane. Hexane is a powerful non-polar solvent that is commonly used by the industrial sector to extract plant oils. We hypothesized that emulsifying the algae in the presence of Hexane would effectively shear the cell walls, releasing the oil. Once released, the oil would bind to the hexane and separate itself from other by-products, such as salt water and soluble molecules.

We were able to obtain and emulsifier from Wholesome Energy and Nonox Ltd. With this device we were able to confirm our hypothesis and the emulsification of algae was a success because; Cells were being collected and harvested simultaneously, the algae cells were quickly sheared in the presence of hexane, this emulsification device could be incorporated into a flow-through process. The system was fully scalable, and most importantly the process did not require energy intensive harvesting steps.

We decided that for each harvest that 15 liters of algae would be emulsified with 1 liter of hexane at 50 psi. This pressure was necessary to properly shear cell walls but also maintain reasonable flow rates. After much trial and error we added a flow control valve to in order adjust the hexane input so we could maintain the 15:1 ratio previously mentioned.

To begin the harvesting process each participant must be properly equipped with adequate safety gear. A cell count is conducted on a hemocytometer slide to record the algae’s concentration of that day. The observed cell concentration must be recorded in lab notebook for future reference. The fume hood window needs to be opened and held it in place with the bungee cord located above. The fume hood timer needs to be activated and kept track of or it will shut off. Next, the desired amount of algae culture must be collected from the culture tub. The top-off system needs to be cut off by turning the lever below the top off system to be perpendicular to the tubing. This will restrict excess amounts of freshwater from entering the tub, potentially causing a salinity shock. The tub must be sized up and replenished before the top-off valve can be reopened.

Align the harvesting pump on the cart in front of the fume hood window and prepare the pump by wiping the tubes with a rag and plugging it into the power strip. Before running anything, it is imperative that all the tubes are correctly placed. The two upper valves on the pump are the inlet valves, the valve all the way to the left on the bottom is the outlet valve, and the valve on the right on the bottom right is not used in these procedures.

Figure 2.2.1: This emulsifier obtained from Wholesome Energy and Nonox Ltd was used to harvest our algae culture.

Of the two inlet valves, the valve in the middle with the flow rate dial is the hexane inlet, and the valve on the right is the algae inlet. One the tube on each of the used valves is for priming. The pump must have a substance to pull on or else the machinery will be damaged. To prevent damage and ensuring a constant flow in the system, the pump must be primed. We primed our the harvester using freshwater and a funnel. The primer tube was raised and freshwater primer was poured into the funnel connecting to the tube. Once the tube is filled with primer, the pump is turned on until the gear pump creates a vacuum and pulls water throughout the pump.

While the primer is running through the pump, the emulsifier chamber needed to be adjusted to a pressure of 50 psi. This is done by using a flat head screwdriver to adjust pressure up or down. As mentioned before, this pressure properly emulsifies cells at reasonable flow rates.

Figure 2.2.2: This schematic represents our emulsifier. Algae culture and n-Hexane are drawn in by the pump and emulsify in the emulsification chamber. The output is collected in a carboy that is placed inside our fume hood.

Next, the n-Hexane flow rate must be adjusted. A total flow rate of the harvesting system with two inputs was calculated using varying pressures. Table 2.2.4 below expresses the change in flow rates as the pressure of the pump increased and decreased.

As the pressure increased, the total flow rate of the harvester decreased as expected. From this table, a pressure of 50 psi was selected to continue working with throughout the duration of our project. A flow rate of the n-Hexane input needed to be established as well. From the flow rates obtained from 50 psi, the flow rate of the n-Hexane input and the total amount of time each run would take was calculated. Table 2.2.5 displays these results below.

Figure 2.2.3: To achieve emulsification, a pressure of 50 psi must be reached before the emulsification tube. The pressure was adjusted by turning a knob with screwdriver located on the tube, which either restricted or permitted flow.

Table 2.2.4: The established flow rates at each tested pressure is represented here. 50 psi was used for the continuation of this project because at this rate the flow rate was manageable, the energy costs were affordable, and the cell lysis was still effective.

After the flow rates were established with rigor, effective harvesting was ready to take action. Each full trial consisted of three runs simply to have a higher yield of algae crude oil for measuring purposes. Before the harvester is activated, a Kilo-Watt Energy Reader is attached to the plug to calculate the wattage the pump requires. Though only 8 effective runs were conducted with the harvester, we were able to gather 10 runs of data with the energy reader. Table 2.2.6 below shows the run data and the average energy consumed daily, per run, and the cost per run of the harvesting pump.

Table 2.2.5: The total run time as well as the flow rate of the n-Hexane input is displayed. Each 50 mL of n-Hexane should be consumed during the process should take approximately 19.43 seconds to maintain flow rates.

Table 2.2.6: Each of the energy readings for the runs conducted are displayed. The associated costs were calculated as well from results of the Kilowatt Energy Reader.

Once the flow rate is achieved and the energy reader is connected, the system is now ready to harvest. First, the n-Hexane needs to be safely transported to the fume hood using protective gear. A 1000 milliliter beaker is filled with the desired n-Hexane amount inside the fume hood, then the hexane container is returned to the flammable cabinet. Once the system is turned on, a timer should started, recording the total harvesting run time. Our harvesting process used 1000 milliliters of n-Hexane with 15,000 millimeters which required 6 minutes and 48 seconds to run.

We continuously checked the flow rate of the n-Hexane, and the pressure of the pump. The harvesting process ran its course until one or both of inputs depleted. If one input was depleted before the other, the system was turned off and the remaining volume was recorded. Stop the timer the moment the system is turned off and record the processing time. The KiloWatt reader energy data was then recorded.

The output tubes are then removed from the carboy and it was capped to reduce n-Hexane evaporation. The output mix inside the carboy needs sits for at least 24 hours, allowing the n-Hexane to separate from the intermediate and saltwater layer. Three of these runs are the equivalent to a single trial.

2.3 Separation of End Products

The objective of the centrifugation process is to separate n-Hexane and algae oil from the intermediate layer. Amphipathic molecules found inside the intermediate layer trap amounts of n-Hexane and algae oil. By using a centrifuge we can spin the intermediate layer down, separating the less dense n-Hexane from the denser amphipathic molecules. The more n-Hexane we obtain, the more we can recycle, improving the sustainability of the process. To begin centrifugation each team member must be properly equipped with adequate safety gear. The fume hood needs to be on and running before the carboy can be opened. The volume of each carboy is recorded before removing their contents. A large 3,500 milliliter beaker was used to remove the bottom layer. The bottom layer consisted of unprocessed cells and saltwater. This layer was removed from the carboy and safely disposed of. The intermediate layer and n-Hexane are then poured into 1000 milliliter beakers for more accurate measurement. To reduce n-Hexane evaporation, all 1000 milliliter beakers were covered with plastic wrap.

The centrifugation system is outside of the fume hood, so the intermediate being centrifuged must be poured into centrifugation tubes and capped. The centrifuge we used could only spin six, 15 milliliters tubes at a time. Once the six tubes were filled to the 15 milliliter mark and capped, they were then placed into the centrifuge and spun for four minutes. After four minutes, three distinct layers are present; the n-Hexane/oil mixture at the top, a small emulsified intermediate layer, and a green saltwater algae layer. We suspect that the intermediate layer contains minimal amounts of n-Hexane and oil which can never be fully separated due to the lack of power our centrifuge provides.

Figure 2.3.1: Left - This carboy contains the output of a harvest. The transparent top layer contains the emulsified n-Hexane and algae cells. The bottom green layer contains salt water, polar molecules, and unprocessed algae cells. Right - The bottom layer is poured off and disposed of while the top layer the emulsified top layer is collected into beakers for more accurate measurement. The right beaker contains only emulsified output while the right beaker contains some saltwater byproduct.

Figure 2.3.2: Left - This centrifuge was used to further separate the emulsified output to retrieve trapped n-Hexane and oil. Right - This test tube has been centrifuged, providing 3 distinct layers. The top layer is n-Hexane containing oil, the middle is amphipathic molecules which trap n-Hexane and the bottom layer is saltwater by-product.

The top layer was then poured off into a beaker in the fume hood and later used in our reclamation process. The intermediate layer had to be properly disposed of since it contained n-Hexane. This layer and the bottom layer were poured into beaker in the fume hood and left until all n-Hexane was evaporated off in. Once fully evaporated the remaining by-product could be safely disposed of. The final volume of n-Hexane/oil mix was recorded and now reclamation can be conducted.

2.4 Algae Oil Purification and Solvent Reclamation The idea of the reclamation system is to ultimately save costs, resources, and make the process as sustainable as possible. In this process, the n-Hexane/oil mix that has been collected from the harvesting and centrifuge process is poured into a distillery device. Our distillery device was created by Remy Biron, a former algae capstone member. The oil is dissolved into the n-Hexane solvent and the reclamation system separates the two. Water is heated to 90 degrees Celsius and pumped through tubes in the distillery. This temperature is above the boiling point of n-Hexane but below that of oil. This allows the n-Hexane to vaporize and rise to the condenser where it is collected back in its liquid phase. A more purified oil mixture falls to the bottom of the device where it is collected and rerun through the distillery to further purify it. All the reclaimed hexane can be recycled and reused for the harvesting process.

First, a 5 gallon bucket that contains a heating coil inside is filled with tap water and covered with a lid. A variable transformer is plugged into the wall outlet to pull more voltage, heating the water to our desired temperature. The temperature is measured with a glass thermometer that needs to be continually checked. Once 90 degrees Celsius is reached plastic pump tubes were placed inside the bucket and the pump was turned on. The condenser chamber requires ice water to be pumped through a copper coil. Roughly 2 gallons of water and 5 pounds of ice is mixed in bucket with a submersible pump. This pump is connected to the copper condenser coil with plastic tubes and the outlet is recirculated into the bucket.

Figure 2.4.1: This reclamation device was created by a previous JMU ISAT student, Remy Biron. It is used to distill n-Hexane off any retrieved algae oil. The device is pumped with hot water on the right side and pumped with cold water on the left. The n-Hexane harvester output is poured in, vaporizing any n-Hexane. The oil continues to fall downward as the n-Hexane rises to the condenser.

When the hot and cold water begins circulating through the system, the n-Hexane/oil mix can be poured in. A funnel is connected to tubing which leads to the miscella inlet. The funnel should be able to reach into the fume hood so any n-Hexane fumes can be removed by the fume hood. Both the condenser and evaporator valve should be fully closed when initially introducing the mix. A 500 milliliter beaker is placed under the evaporating valve outlet and a 1000 milliliter is used to collect condensed n-Hexane. The n-Hexane/oil mixture is slowly poured into the miscella funnel to allow the n-Hexane to fully vaporize. The system can “burp” the n-Hexane back up the inlet if too much is added quickly. Once the condenser valve is opened, recondensed n-Hexane will immediately start to drip out. This valve needed to remain open during the addition of n-Hexane/oil mix in order for n-Hexane fumes to pass by the condenser. When the condenser outlet stopped dripping, the valve was closed and collected n-Hexane was placed into the fume hood. With the 500 milliliter beaker sitting under the evaporator valve, the valve was opened and all contents were collected. Depending on the volume of collected purified oil, a second reclaiming may be needed. If we collected a volume larger than 100 milliliters, our team would re-reclaim this amount to further purify the oil and collect more n-Hexane.

All reclaimed n-Hexane was measured and recorded then combined and poured into a securable container to be stored in the flammable safety cabinet. The bottom residuals collected we left in the fume hood without a cover, so any remaining n-Hexane could evaporate off. Once n-Hexane is fully evaporated, oil will be the only remaining factor. The three runs are combined to create a trial and all the amount of algal oil is recorded.

2.5 Algae Oil Quantification

After all the procedures for harvesting, centrifuging and reclamation have been completed, the amount of algae oil can be quantified. By allowing the remnants of the n-Hexane in the small beaker evaporate off in the fume hood, algae oil is left as a result. To easily transfer this oil to a smaller tube for storage, the

Figure 2.4.2: Left - A beaker collects purified oil from the evaporator valve. Right - The reclaimed purified hexane is shown on the left and the purified oil on the right. This is the result from a single harvest run of 4 gallons.

beaker is heated on a hot plate slightly and once the oil is loose it can be scraped out. Meanwhile, the instruments and storage tube were tested for uncertainty with 1 mL of water. A 15 mL tube was used (smaller measureable tube is recommended but we did not have access to one), and an approximate measuring uncertainty was gauged by adding and removing water in a series of measurements (5 microliters, 50 microliters, 75 microliters, 100 microliters, etc.). Whichever amount of measurement was visibly different to the naked eye was used as the measurement uncertainty. For example, if a noticeable difference between adding 5 microliters to the contents of 1 mL was visible, that was the uncertainty. This was done for the yields and energy readings to understand that external factors like human error, instrument calibration, and other factors can highly affect the precision of results. After all the data was collected, the standard deviation was calculated for the results of the oil yields and for the associated cost per run obtained by the Kilowatt Reader, which connects to the harvester and reads the amount of energy the pump requires.

2.6 Determination of Energy Input vs. Output

As mentioned before, a Kilowatt Energy Reader is connected to the power outlet of the harvesting pump to assess its wattage and the amount of energy it requires to be powered on. This device produces wattage readings after the process is complete and that value is recorded. After each of the runs have obtained energy readings, the standard deviation is then calculated.

The combined uncertainty, displayed as Equation 1 below, can then be used to quantify the uncertainty of this measurement.

Combined Uncertainty=√[YieldRun

−1

∗σCost ]2

+[−( CostRun )(Yield

Run )−2

σ yield ]2

(1)

Figure 2.6.1. - A P3 International Kilowatt Energy Reader was used to determine our energy input for the harvesting process.

The uncertainty of a measurement can be for the better or worse for a scenario, but it is very important to include in error prone calculations. Once this was determined, it was applied to our oil yields to understand that it is difficult to have exact precision in a cheap lab setting.

Along with the calculation of the combined uncertainty, an energy ratio was calculated as well to understand if this process was efficient from an energy standpoint. It only makes sense to have a process that produces energy require a smaller amount of energy input and have a larger amount of energy output. This was done using Equation 2 displayed below.

Energy Ratio= Energy Readingof Pump(Joules)OilYield per run∗Algae oil energy density (Joules)

= Energy input (Joules)Energy output (Joules)

(2)

For this model, the amount of oil yield per run must be quantified and the energy input from the harvester. Since this is ratio, it is unit-less where the units on the top must match the units on the bottom to cancel out. From previous projects, a bomb calorimeter test was performed on the algae oil and determined that the energy density was 42 MJ/L. This value was then multiplied by the oil yields converted to liters to obtain an energy measurement. This was done for each of the trials performed and an energy ratio was presented in the final findings.

2.7 Mathematical Modeling of Theoretical Algae Oil Yields

The purpose of this report is to understand how algae biofuels can impact the decision to switch to a cleaner more sustainable fuel source while still being able to meet the current demand for liquid fuel. To understand the full potential algae microorganisms could have to contributing to the United States fuel supply, a comparison had to be made against conventional biofuels made from soybeans and corn. Data was gathered from reliable sources about the oil yielding per unit of area from both of the conventional biofuels used as a comparison. The data gathered during this experiment was used to determine the oil that can be yielded per unit area for this specific algae strain. Once the units correlated, a comparison is made to see which fuel is most efficient in terms of land use, area required, harvesting rate, and energy density per unit area.

2.8 Cost Analysis

The main component associated with the calculated costs was the amount of energy input the harvester pump is required. This was the only parameter that dictated the cost throughout this experiment. Despite there being other costs involved, in a real world application, it can be assumed that those costs would be mitigated which is why they were not included. Costs that fall into this category includes the grow lights, the centrifugation, the fume hood, and the reclaiming of n-Hexane. All of these would not be necessary in a large scale setting located in the ocean. Once the energy reading was obtained in kWh, the current cost of electricity in Harrisonburg, Virginia was used to obtain a total cost of usage for the pump. Then the rate at which oil was yielded to see how long the pump would have to run for in order to obtain a gallon, a

barrel or a total U.S. demand of algae oil. Now the cost can be compared to current fuels sources and other biodiesels.

3.0 Results

As the United States is in search for a cleaner fuel alternative to oil usage due to environmental concerns and energy security, this project was pursued with the objective to have an output of algae crude oil from our novel harvesting pump as energy efficient as possible. To further optimize our process included adjusting flow rates, adjusting the pressure of the system, understanding the energy costs as parameters change, and quantifying the energy density of our oil output. The first results obtained were the flow rates necessary to maintain the algae to hexane ratio of 15:1. This was already specified in a previous project and was not tampered with during our experimentation. After the flow rates were adjusted as shown in the Methodology sections, the energy readings were established and the process could then be applied in real time.

With the dictated energy results, a cost for the algae oil output was generated. But first, a n-Hexane loss was noted mainly from evaporation and throughout the system. Table 3.1 below shows an accumulation of the n-Hexane input and output for all the runs, showing that on average only 40% of the total n-Hexane put into the system is retrieved at the end of the oil extracting process.

Table 3.1: There was a noticeable n-Hexane loss throughout the system. The losses were documented and are presented in this table. The average loss of about 40% was used in further calculations.

The total findings from the 4 main trails are displayed below in Table 3.2, noting that each is only yielding on average 40% of its total capabilities. For comparison, for each trial is calculated with a theoretical return of n-Hexane of 100%. Trial 1 was obviously the best oil yielding trial that was conducted as seen from the table. If the process was capable of retrieving 100% n-Hexane back, this novel harvesting process would be energy efficient and borderline cost competitive.

The average oil yield per trial varied greatly depending on the health and concentration of the algae pool at the time of harvesting. As seen from the table above, the trials displayed had a range of oil yields because of the algae health. Figure 3.1 below depicts the oil yields per trial along with the standard deviation.

Table 3.2: The total run data for the duration of this project is presented above, as well as a projected 100% retrieval of n-Hexane. It is important to notice the data specifically for Trial 1 since this was the highest oil yielding trial. As it is 40% efficient in reclaiming n-Hexane, the cost per gallon is at its lowest and the energy ratio is almost less than 1. As it is 100% efficient in reclaiming n-Hexane, the cost per gallon substantially drops and the energy ratio is less than 1, meaning it is an energy efficient

Trial 1 was by far the best oil yield experienced in the short amount of lab time. Trials 2 and 3 suffered based on reasons to discuss later in this report, but ultimately because of the culture health. An average oil yield from all trials and the uncertainty of the measurement is displayed as Figure 3.2 below.

Figure 3.1: Above is a graph representing the algae oil yields for each trial conducted during the duration of this project. Each trial was a consolidation of 3 runs through the harvester. Trials 1 and 4 had a healthy algae concentration, while Trials 2 and 3 were the result of an algae die off. The error bars represent the standard deviation of the data set.

Figure 3.2: This graph represents the average oil yields for the sampled runs. The error bar represents the calculated combined uncertainty, which takes into account all the influence uncertain factors with measurements and some human error.

The average oil yield from all the trials was approximately 0.75 mL. The uncertainty of this has a rather large range due the range in yielding data from each of the trials. With the different oil yields, each trial experiences a difference in cost as well. Table 3.3 below is a sensitivity analysis of the amount of oil output by the system to the total cost per gallon of algae oil.

As the yields of oil increase, the price per gallon of oil decreases. This is displayed in Figure 3.3, where the graph steadily declines exponentially and plateaus. The cost to harvest for this data set was $0.24 per full day of harvest.

Applying the data to real time, Table 3.4 below indicates the amount of algae culture the United States would need if algae biofuels were to fuel the demand the year of 2015 experienced. For a comparison of data, corn based ethanol and soybean based biodiesel was calculated as well.

Table 3.3: The sensitivity analysis shows how a slight change in oil yields highly affects the cost per gallon. As the oil yields increase, the cost per gallon decreases. A standard energy cost of $0.24 per kWh was used.

Figure 3.3: The graph expresses the negative linear relationship that cost per gallon of algae crude oil has with the algae crude oil yields. As the amount of yields increases, the cost per gallon decreases.

The biggest takeaway from this table is that algae requires minimal area for cultivation compared to other alternative liquid fuels. The biggest contributing factor to this is the ability algae has to double every 48 hours. This creates an astounding advantage over crops like corn and soybeans that can only be harvested once a year. As shown by the data, algae as a fuel source shows a huge promise to the future of biodiesel. Other forms of biodiesel do not compare and this can largely be attributed to doubling rate algae experiences.

4.0 Discussion

As stated previously, the need for an alternative fuel for energy security and the betterment of the environment has experienced notable attention recently as global climate change becomes more prevalent. With this, algae biofuels have shown promise with oils yields but has chronic issues with cultivation and harvesting. The purpose of each of the experiments conducted was to optimize the novel harvesting method to yield the highest algae oil percentage as possible. In doing so, the first stage required an understanding of the harvester in terms of its harvesting speed at different pressures. As the pressure increases, the harvesting speed of the system decreases. In other words, as pressure increased, the flow rate of liquids circulating slows down. High pressure makes the pump work harder as well, which in turn increases the energy the system requires. As the pressure was adjusted as seen in Table 2.2.4 , a standardized pressure was needed to consistently run the harvester on. 50 psi of pressure was selected to continue working with because it allowed a slow enough flow rate to ensure emulsification, it was a manageable speed for lab members to work with, the pressure wasn’t too high that our energy input costs were expensive, the cell lysis was extremely effective and because the pump itself was not over stressed

Table 3.4: Application is a large aspect to any novel system. This table shows calculations of real world applications of algae oil harvesting and its dominance over other types of biofuels, where it requires less area of maintenance and has a continuous harvest. The main take away is that algae oil harvesting requires a lot less area to harvest a larger amount of oil.

at this level of pressure. These flow rates were established by using two inputs with a colored liquid (algae or dyed water); once the output showed color after the system was running, a timer was stopped. Now that an approximate flow rate was achieved for the algae input of our process, a n-Hexane flow rate needed to be established. An algae to n-Hexane ratio of 15:1 needs to be maintained throughout the process. This was a predetermined ratio established by previous experimental groups and was not tampered with during this project. A control flow rate valve was installed onto the pump on the n-Hexane input valve. For an effective 15:1 algae to n-Hexane ratio, with an algae speed at approximately 41.54 milliliters per second at 50 psi, the n-Hexane must flow at 2.57 milliliters per second. This was maintained by checking the speed of consuming each 50 milliliters of n-Hexane which should take approximately 19.43 seconds, giving the system a total run time of 6 minutes and 28 seconds shown in Table 2.2.5. With the flow rate data achieved, runs were harvested in sets of three. The three runs were consolidated and considered one full trial. This was done for measuring purposes since each run yielded a very small amount of algae crude oil, but when the volume was combined it was easier to denote a measurement. One of the main objectives of this project was to calculate the energy ratio. To make this process energy efficient, an energy ratio less than one is necessary, where a smaller amount of energy going into the system and a larger amount coming out of the system. To quantify the amount of energy required for our system, a Kilowatt energy reader was used which plugged into the harvester and read the total amount of wattage the harvester required. The average wattage each run, shown in Table 2.2.6, required was approximately 2810.2 watts, with a standard deviation of 140.9 watts and a calculated combined uncertainty of 314.367 watts. Despite the range of the data, the average was fairly close to the median, allowing the data to be interpreted normally. Using an energy density value of 42 MJ/L from a bomb calorimeter test conducted in a previous report, an energy ratio could now be calculated based on the amount of algae crude oil yielded. Equation 2 in the Methodology section exhibits how the ratio was obtained. As the team continued working with the harvester under the established parameters, a comparison between the input amount of n-Hexane and the output amount of n-Hexane was brought about. From this, a substantial n-Hexane loss was discovered. On average, our system loses about 60% of the total n-Hexane put into the system, where only 40% is obtained at the end of our process. Table 3.1 expresses the exact percentage of losses each run experienced along with the calculated standard deviation, and average value of the total losses. The losses of n-Hexane can largely be attributed to n-Hexane evaporation since its volatility is rather high and to the fatty lipid layer in our harvesting output, as well as some to getting caught up in the pump and reclamation system. In some procedures of this project, you can actually see the n-Hexane evaporate while working with the substance. This can be seen during harvesting, when the reclamation system drains the reclaimed n-Hexane, and when the output carboy is uncovered during the harvesting process. The fatty lipid layer of the harvesting output has strong indication that n-Hexane is still present in that layer. Previous projects before this one never experienced a fatty layer as large as seen during this project. Usually the n-Hexane flows to the top of the output beaker since its density is much lighter compared to algae, but throughout this project only a small portion of it floats to the top while the remaining amount is caught in the lipid layer. The amount of n-Hexane that is caught in this layer is unknown, but the smell of this layer suggests a large amount. To mitigate losses, centrifugation was

added to our procedures to gather as much n-Hexane as possible. The n-Hexane is important throughout this process because its job is to trap oil molecules and hang on to them. If there are substantial n-Hexane loss, there are also substantial oil losses, giving lower oil yields and a less efficient process. Despite the efforts to centrifuge with a very low powered surplus 15 mL tube centrifuge, the n-Hexane was still not fully retrieved. Even with the noted losses, the system always extracted algae oil. Table 3.2 represents all of the trials and the outputs associated. Trial 1 had an oil output of 1.75 mL, Trial 2 had 0.5 mL, Trial 3 had 0.5 mL, and Trial 4 had 1.0 mL. Each oil output has an energy input, energy ratio, associated cost, and combined uncertainty. It is important to note that this project did not really obtain a gallon let alone a barrel of algae oil. The combined uncertainty was calculated to understand the fluctuation within the presented data, knowing that not all of the measurements taken are not precise due to instrument calibration, human error, and given assumptions. The combined uncertainty was calculated using Equation 1 from the Methodology section. Trial 1 was by far the best trial, yielding the largest amount of oil in comparison to all the trials. This was the first three runs conducted, when the algae was its healthiest, most concentrated, and was provided all the necessary nutrients to prosper. With 1.75 mL of algae oil extracted from an input of roughly 45 liters of algae, an energy ratio was calculated to be around 1.72. Since this ratio is larger than one, the process at this yielding rate is not energy efficient because more energy is being put into the system than the system is providing in the end product. Basically, a larger amount of oil needs to be collected in order to lower the energy ratio. Assuming only 40% of the total oil was collected due to n-Hexane losses, a generated cost per gallon of $6.08 with a combined uncertainty of $0.28 was calculated for the oil yielding rate in Trial 1. With the current gasoline cost per gallon at $1.95 in Harrisonburg, VA, our process shows to not be economically feasible at this oil yielding rate. To compare a raw product to another raw product, a cost per barrel of algae crude oil was calculated at $255.31 per barrel with a combined uncertainty of $28.56. When compared to the current crude oil barrel cost at $40 per barrel, this method is still not economically feasible. Trials 2 and 3 can be grouped together because these were the result of the same issue and yielded the same amount of oil. These runs were conducted during a time period when the algae strain used was not health because of a bacterial infection, a low concentration of algae cells was present, and another algae strain had begun to thrive in the culture. This can mostly be attributed to an issue with the automated top off system that maintained the algae’s salinity when evaporation occurred. The top off system is controlled by a buoy, so when there is evaporation, the buoy stretches down and releases dechlorinated fresh water and nutrients. Fresh water is used in the top off system for the saltwater algae strain because as the water evaporates from the algae culture, the salt remains, keeping the salinity relatively consistent if freshwater is continuously added when needed. This top off system failed one day and released 15 gallons of fresh water into the saltwater system, shocking the culture and the result was a algae die off. The algae strain that thrived was one that could survive in lower salinities, but this was not a high oil yielding strain. It is thought that this contaminate algae strain was introduced to the culture by uncleaned glassware. With 0.5 mL of algae oil extracted from an input of roughly 45 liters of algae, an energy ratio was calculated to be around 6.02. Since this ratio is larger than one, the process at this yielding rate is not energy efficient because more energy is being put into the system than the system is providing in the end

product. Basically, a larger amount of oil needs to be collected in order to lower the energy ratio. Assuming only 40% of the total oil was collected due to n-Hexane losses, a generated cost per gallon of $21.28 with a combined uncertainty of $2.38 was calculated for the oil yielding rate in Trials 2 and 3. With the current gasoline cost per gallon at $1.95 in Harrisonburg, VA, our process shows to not be economically feasible at this oil yielding rate. To compare a raw product to another raw product, a cost per barrel of algae crude oil was calculated at $893.57 per barrel with a combined uncertainty of $99.96. When compared to the current crude oil barrel cost at $40 per barrel, this method is still not economically feasible. Trial 4 the algae culture began to replenish itself with extensive care and extra nutrients after a couple of months. The culture started to have a higher concentration of the algae strain with the higher oil yield and was slowly beating the contaminate algae strain. With 1.0 mL of algae oil extracted from an input of roughly 45 liters of algae, an energy ratio was calculated to be around 3.01. Since this ratio is larger than one, the process at this yielding rate is not energy efficient because more energy is being put into the system than the system is providing in the end product. Basically, a larger amount of oil needs to be collected in order to lower the energy ratio. Assuming only 40% of the total oil was collected due to n-Hexane losses, a generated cost per gallon of $10.64 with a combined uncertainty of $1.19 was calculated for the oil yielding rate in Trial 1. With the current gasoline cost per gallon at $1.95 in Harrisonburg, VA, our process shows to not be economically feasible at this oil yielding rate. To compare a raw product to another raw product, a cost per barrel of algae crude oil was calculated at $446.79 per barrel with a combined uncertainty of $49.98. When compared to the current crude oil barrel cost at $40 per barrel, this method is still not economically feasible. Knowing that all of the trials did not produce and energy efficient ratio and the price range was still high, the n-Hexane losses resurfaced in importance. 60% of the total n-Hexane put into the process is still missing. So, a 100% return of n-Hexane was calculated for each oil yielding rate for each trial. This was not actually obtained throughout this project’s time frame. Trial 1, yielding 1.75 mL of oil at only 40% return would have yielded 4.37 mL of oil if there was a 100% return of n-Hexane. Not only is 4.37 mL of oil a higher oil yield and much more intriguing, it also provides an energy ratio of 0.69, which is less than one meaning that more energy is being produced than is being used to produce the oil. The cost per gallon calculated was $2.43 with a combined uncertainty of $0.28, and a cost per barrel at $102.22 and a combined uncertainty of $11.80. If we were to achieve 100% return of the n-Hexane, the end product of oil would be a larger volume, provide an energy efficient process, and be almost cost competitive due to the fluctuations in the current oil market. Trials 2 and 3, yielding 0.5 mL of oil at only 40% return would have yielded 1.25 mL of oil if there was a 100% return of n-Hexane. Not only is 1.25 mL of oil a higher oil yield and much more intriguing, it also provides an energy ratio of 2.41, which is substantially less than the previous energy ratio of 6.02. The cost per gallon calculated was $8.52 with a combined uncertainty of $1.28, and a cost per barrel at $357.77 and a combined uncertainty of $53.68. If we were to achieve 100% return of the n-Hexane, the end product of oil would be a larger volume, but at this oil yielding rate the process would still not be energy efficient and not cost competitive.

Trial 4, yielding 1.0 mL of oil at only 40% return would have yielded 2.50 mL of oil if there was a 100% return of n-Hexane. Not only is 2.50 mL of oil a higher oil yield and much more intriguing, it also provides an energy ratio of 1.21, which is almost but not quite energy efficient, but it is less than the previous energy ratio of 3.01. The cost per gallon calculated was $4.26 with a combined uncertainty of $0.52, and a cost per barrel at $178.89 and a combined uncertainty of $21.92. If we were to achieve 100% return of the n-Hexane, the end product of oil would be a larger volume, but at this oil yielding rate the process would still not be energy efficient and not cost competitive.

From the presented data, energy input versus energy output is very depended on the oil yields from each of the trials. As the oil yields increases, the energy ratio decreases making the process energy efficient. The process needs to be efficient because if there is more energy going into the system, consumption increase as does cost. For this process to be feasible, the costs must be low to be somewhat competitive. The energy ratio is directly related to oil yields, and algae concentration highly impacts the amount of oil yielded. The more concentrated the cells, the more oil yielded; the more oil yielded, the more energy efficient and cost efficient the process is. Using the graphs displayed as Figure 3.1 and 3.2 in the Results section, a visual is provided for how much the oil yields differed and decreased from the salinity shock. This was a major setback our system experienced that will be explain in greater detail later on. The average oil yield suffered mainly due to Trials 2 and 3. But looking back at Table 3.2, notice how sensitive the cost and energy ratio is to just a milliliters of a difference in algae oil output. Table 3.3 is a brief sensitivity analysis of algae oil yields vs. cost per gallon of algae crude oil. Not only does this show how important each milliliter of oil output is, it also shows how important it is to understand the losses throughout this system. The system is highly effective, but there needs to be more efficient ways to obtain as much of the n-Hexane put into the system as possible. Either through a new and powerful centrifuge or testing new algae strains that would not produce a large fatty lipid layer. Figure 3.3 portrays the negative linear relationship the cost experiences as oil yields increases at a flat rate cost per harvest of $0.24 per day. Taking a step back from the fine details of the data, a statistic provided by the EIA showed that the United States consumed 19.4 million barrels of petroleum in the year 2015. Assuming that the algae crude oil could be substituted directly in the place of petroleum, a hypothetical scenario was calculated and is shown in Table 3.4. Using the best oil yielding rate from Trial 1, where 45 liters of algae harvests 1.75mL of algae crude oil, and the harvester’s actual 40% yielding efficiency, the algae culture size needed to produce 19.4 million barrels of crude oil is approximately 62 trillion gallons. This assumes a continuous flow process of harvesting and an algae double rate of approximately 48 hours. This volume required would require an area of space equal to approximately 98,000 square miles of land, which is equivalent to about two full states of Virginia. If the novel system was perfect and had a 100% yielding efficiency, an area less than the one state of Virginia of a culture size would be needed to sustain the petroleum demand in 2015. With that being said, it is important to note the area needed is equal to that of Virginia, but this process would prosper off-shore in portions of the ocean that have no life. In comparison, if another alternative biofuel source were to attempt to fulfill the demand of 2015, like corn based ethanol, an area equivalent to about 33 states of Virginia would be required. Likewise, if it were soybean based biofuels, an area equivalent to about 217 states of Virginia would be required. As stated, this data truly exhibits the potential and the promise algae has on future biofuel production. With its higher oil content and doubling

rate, algae could sustain the U.S. without disrupting the global food supply or causing a massive change the current infrastructure in place.

4.1 Unintended Consequences

In this project, setbacks were experienced due to a salinity shock leading to a massive die off of Nannochloropsis and a subsequent bacterial bloom. Due to the change in the salinity of the algae tub a contaminant algae strand was able to thrive under the new conditions. Once run through the harvester, the bacteria’s lipid membrane re-encapsulated the hexane and made it unrecoverable for the current technology we were working with. This resulted in a drop in a major drop in the oil yield. Contaminant algae strains and salinity changes would be some issues needing further investigation before real world implementation. For real world implementation of this project offshore cultivation would be the most economically and environmentally friendly. However, as with any new technology, there would need to be a further in depth environmental study on the possible effects of growing algae in dead zones. With the ocean currents and ocean storms, algae have the possibility of moving to an unplanned area. This could be particularly detrimental if the algae were to float over any reefs and block out the sunlight. Dead zones are also breeding grounds for whales and are intersected by Maritime trade routes so growing algae in these areas has the possibility of interfering with these practices. Algae also grows in blooms naturally so having the technology to harvest the algae oil could incentivize cleanup of harmful algal blooms but could pose the challenge of over harvesting algae blooms that are beneficial and necessary to the environment. Laws would need to be written on who was allowed to harvest algae and a deeper investigation would need to be conducted on environmental impacts but as far as other fuel sources algae has far more benefits than negative consequences.

5.0 Conclusion

The objective of this project was to provoke the extraction of algae biofuels and implement its use in the current fuel supply, ultimately because the environment is suffering from the pollution of current fuel sources. Through this project two main roadblocks mentioned by the DOE’s 2010 National Algal Biofuels Technology Roadmap were focused on to improve the harvesting method. As these roadblocks were noted, it was sought to overcome them. Throughout the extensive research and efforts during this project, it can be concluded that this novel process works, there is a consistent yield of algae oil, and the end algae oil product fits the current infrastructure. Algae biofuels in theory is the perfect fit to replace heavy polluting fuel sources like crude oil because there is no need for remodeling the current infrastructure. By making improvements to both the system and the culture, the oil yield can be increased to a point that makes it more competitive on the market. Despite the findings from this novel harvesting method, there are still challenges to overcome. Some include the unknowns involved with large scale cultivation and large scale harvesting. Algae as a biofuel contributes as a positive environmental impact giving it a promising future. When it becomes more of a reality is when people make a conscious effort of realizing the current situation people live in and take action to improve it.

6.0 Current and Future Direction of Research

A top priority for developed and developing nations has been to mitigate carbon dioxide emissions and attack other environmental concerns contributing to global climate change, along with securing the current energy crisis impairing sources of energy for future generations [44]. With the growing energy demand of the growing population, a quest for sustainable solutions at a low cost has brought about the idea to utilize biomass as a transportation fuel [44]. This option is readily available and does not require additional production costs [44]. Biomass includes naturally produced substances like algae, plants, and vegetables that are living or recently living, all of which are abundant in the United States. With thermal processing, or lipid transesterification to alkanes and biodiesel, blending the biomass byproduct with conventional diesel is at a low cost and tailored to the current infrastructure [44]. Not only would reductions occur in the total amount of crude oil use, but sustaining fuel supplies would be established until a more stable energy supply was achieved. Recent trials have even indicated that the energy quotient for catalytic alkane production from biomass is twice that of fermentation routes to ethanol [44]. Incorporating biodiesel into the transportation sector would create a more efficient energy use from its characteristics like its higher flash point, low sulfur concentrations and better lubrication efficiencies [45]. Carbon deposition is also decreased significantly when biodiesel is effectively mixed with petroleum diesel [45]. The process of transesterification has been the most commonly used method for the production of vegetable oil and algal oil tied with alcohol in the presence of an acid or base catalyst. Ultrasonic technology is thought to have a greater benefit to transesterification because the ultrasound inaudible to the human ear provides enough mechanical energy for mixing and the required activation energy for the total process, this in total increases the biodiesel net yield and shortens the reaction time of the process [45]. This process shows promise for biodiesel production with algae oil and vegetable oils, with algae exceeding the total production of vegetable oil because of its higher lipid content. As biodiesel production helps to mitigate carbon dioxide emissions, algae growth is also capable of producing fundamental lipids from carbon dioxide through its natural process of photosynthesis [45]. Algae is thirty times more proficient than plants in terms of its lipid production, showing propitious abilities to potentially replace the use of fossil fuels [45]. In a recent experiment conducted by Yeditepe University, biodiesel production from algae proved that about three fourths of work potential of algal biodiesel is used for its production and restoration of the environment, equating to a net gain of about one fourth of the work produced by the algal biodiesel [45]. This process can be easily manipulated to generate a better renewability indicator through the use of genetic engineering techniques. Genetic engineering not only can improve the process’s efficiency itself but it also can work to cultivate a species of algae that is suitable intimately for biodiesel production, and mitigating various environmental concerns. Research has shown favorable data from the Monoraphidium algae strain because it is cold-tolerant and bioremediated wastewater while producing lipids [44]. Cold-tolerance is a major subsidy this algae strain has to offer. Creating a warm environment for some algae species to live and thrive can be very energy intensive. Reducing this energy input can increase the total exergy of the algae biofuels production. An inferior component to biodiesel productions as a whole are the excess nitrogen emissions generated during its fuel use life cycle [44]. Working against the norm, Monoraphidium yields a lower nitrous oxide emissions during engine tests, exhibiting the greatest potential of the studied algae strains [44]. From a recent collaborative scientific article supported by the Department of Energy, scientists were

able to conclude that the alga is able grow with a large cell density, remove wastewater pollutants like nitrates and phosphates, grow in various freshwater climates with resilience, and perform at a comparable level as biodiesel with petrodiesel in terms of fuel efficiency [44]. Sustainable futuristic uses for algae biofuels are feasible in the current infrastructure and climate in accordance to the strain of algae cultivated, the process in which the lipids are extracted, and its ability to tackle the overall global climate change concern by reduce pollution emissions in the production and usage. Future research and improvement could be a valuable asset to this program. One big improvement to this project would be getting a centrifuge that has over 10,000 revolutions per minute (RPM) and could handle the large volume amount after the harvesting process. The higher the RPMs, the more the hydrophobic lipid layer is broken down, allowing more of the oil that is dissolved in the n-hexane to be released. Current technology is not very efficient and is extremely time consuming. The system as whole is a proven process that works and produces a crude biodiesel every time the system is run. To optimize the process to a higher level, future researchers can explore the n-hexane level inputs into the algae emulsifier. Currently the process inputs 1000 mL into the algae emulsifier. Theses amount can be adjusted to a smaller volume to find the optimal n-Hexane to algae saltwater ratio. Due to n-Hexanes effectiveness, we hypothesized that there should be a smaller volume put into the system. By reducing the amount of n-Hexane into the system, the amount of n-Hexane loss through evaporation, poor techniques, and unknown losses would be minimized. Once the optimal level of n-hexane is discovered, the focus should change to different algae strains and their oil yield amounts. Nannochloropsis was on the Department of Energy's list of high-lipid content algae strains and thus why it was chosen for this research. It also has the ability to survive extreme conditions regarding the temperature, pH, and salinity of the saltwater. Future research should address the different algal strains and determine which one, under the same growing conditions of each strain, has the highest oil yields. The high oil yield is a critical part of the algal strain selection process, but the doubling rate of the algal strain is just as important. The end goal of this project would be to have a continuous closed loop system that could constantly harvest algae and reclaim the oil. This would make the fast doubling rate a critical trait that the algae must have in order to be processed continuously. The United States Navy is a major contributor to the research and application of renewable biofuels. The Navy is using its authority under the Defense Production Act, which allows the Navy, in partnership with the U.S. Department of Energy (DOE) and the U.S. Department of Agriculture (USDA) to invest in industries that are determined critical to national security; in this case, biofuels. They are continuing to use algae biodiesel as a fuel source option in the future. The Navy has already used 20,000 gallons of algae biofuels to power a decommissioned destroyer. They have called for shipping out an entire fleet running on alternative fuels by 2016. Dennis McGinn, the U.S. Navy Assistant Secretary for Energy, Installations, and Environment, said, “The Navy wants to buy anywhere between 10 and 50 percent biofuel blends for our ships. We want it to be a cost-competitive program. We are working specifically with the USDA to bring down biofuel costs to $3.50 a gallon or less at the commercial scale of 170 million gallons a year by 2016.” Algae based biofuels biggest success story is through the betterment it has to offer the environment. Algae based biofuels as a sustainable and scalable fuel source give it a promising future. As societal

norms change to a more ‘green’ perspective, algae based biofuels could fill the shoes of a green means to a fuel source for energy use. This process to emulsify algae is not set in stone. Other techniques may be utilized to obtain higher yields of oil, minimize energy input, or even using different reclamation chemicals other than n-Hexane. In this particular experiment, a solid separator might help to enhance the concentration of algae before it is transferred into the harvesting pump. Increasing the concentration of the algae and therefore the cell density could lead to a higher oil output in the end. Other techniques to improve upon the process could include making the process a one pass system instead of recirculation through the harvester.

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