Final Summary for the project “A prototype bioreactor for efficient alga production to provide food and feed for future colonies on Mars” Mate Ravasz, Daniel Budinov, Stephan Matthiesen, and Jiri Jirout

1. Executive summary In a new era of space exploration we need to develop strategies to enable long-term human settlements on distant worlds. Our project, made possible by the MBR Space Settlement Challenge, proposes an innovative alga growth system that is capable of producing fresh food while using several local resources available on Mars. The principle applied can also be used in any situation where humans can produce energy, but lack water or soil for traditional agriculture. Using our proof-of-concept design we developed in the last six months, we are now establishing a business that we hope will provide a boost to the emerging alga biotech industry. Manned exploration of space is again in focus. China, the USA, Europe, India, Japan and the UAE have all announced future manned missions off world and are investing heavily in life support systems. Without exception these missions are part of roadmaps to establish permanent bases outside Earth. One of the main unsolved challenges for long-term manned space missions is providing life support for the crew in a renewable manner. Current plans are based on carrying enough food, water and oxygen for the whole mission or conducting regular resupply runs, both of which result in prohibitively high launch costs while condemning the crew to consume canned food during the whole operation. Our project received seed funding from the Dubai Future Foundation through the Guaana.com open research platform. It proposes a radical, innovative solution to provide fresh food for long term space missions: we are designing an automated algae growth platform that can be operated remotely to constantly produce fresh food in any environment.

1.1 How the reactor works In essence, our machine is a combination of an automated greenhouse and an incubator, that is fully self contained and remote controlled. The current prototype is

designed for a Martian setting to use as many local resources on the red planet as possible, thereby minimizing reliance on terrestrial materials to produce food. The system is based around the concept of a photobioreactor. Algae are grown in a chamber filled with liquid growth medium while being surrounded by an array of sensors that monitor culture parameters. The collected data is sent to a remote computer where either a human operator or a predefined growth program issues commands in response. The reactor responds to such commands by altering culture parameters thus controlling algal growth. The grown algae can be harvested and consumed or used for other applications as needed. We optimized our current design for a Martian setting. The culture vessel here is an airtight container connected to a gas box. The gas box contains sensors for CO2, O2, humidity, and temperature. It is also equipped with an automated pressure valve to release excess oxygen produced by the algae, and an electronic air inlet that is connected to a canister of CO2. The latter models the Martian atmosphere which is made of 96% CO2, and could provide an excellent carbon source for the algae if heated and pressurized appropriately. The culture vessel that contains the algae is also packed with sensors. A temperature sensor is placed within the liquid, while a colour sensor and a turbidity sensor is located on its outside to constantly monitor alga health and growth respectively. Light for the algae is supplied in the form of an adjustable LED panel that can be regulated to control a day-night cycle. It is specifically designed to only emit wavelengths of light that are taken up by the algae thereby optimizing energy efficiency and providing better lighting conditions for the algae than the Sun on Mars would. The LEDs also allow operation during extended dust storms on Mars which can last for weeks and significantly obscure natural sunlight for the duration. The culture vessel is also connected to a heating-cooling system to regulate temperature, and a bubbler that mixes the culture itself, and also allows efficient exchange between the liquid and gas phases. Sensor information is collected by a microcontroller that transmits it to a remote computer which runs our own control software that stores and visualizes incoming data, while providing an interface to give instructions to the reactor. Commands

Figure 1: 3D render of the photo-bioreactor prototype

issued to the reactor control heaters, gas valves, the cooler, and the light source as required.

1.2 Algae: the future of food We have also carefully chosen what to grow in the reactor. Higher order plants were quickly discarded as they are inefficient at converting light to chemical energy, produce plant material that is largely inedible like branches, stems or roots, grow slowly, waste water, and need too much space. In contrast, algae are the most efficient producers of biomass from inorganics known to man. They are rich in protein and also contain a range of useful lipids, carbohydrates, and other metabolites. They can grow in liquid which is much easier to manipulate by machinery than a tree or bush rooted in the ground, they produce no inedible biomass, use water effectively, and harvest light with incredible efficiency. Out of a plethora of species, we chose organisms belonging to the family of Nostocs. These cyanobacteria - also called blue-green algae - are known for their rapid division, efficient use of sunlight, and their special ability to take up nitrogen from the air. The latter quality is extremely useful for a Martian setting where nitrogen is mostly available from the atmosphere. On Earth, soluble nitrates are also common, but they still form one of the main bottlenecks for agriculture, which is why soils are regularly topped up by nitrogen-rich fertilizers all over the globe.

1.3 Growing algae for terrestrial applications The system is not only useful for Mars. The principle of a closed, automated food production unit can be highly useful for any human settlement where traditional agriculture is not possible. A container churning out fresh algae can be a significant quality-of-life improvement for polar research outposts, oil rigs, cruise ships, submarines, remote military outposts, and any settlement where electricity can be produced, but supplying fresh food is not straightforward. The technology can also provide the basis for future large-scale operations to provide staple food supplies in areas where it is mostly imported, notably northern Africa and parts of the Middle-East. Algae also have potential applications outside the food industry. As they contain more protein than most other plants, they can provide an excellent protein-rich feedstock for industrial livestock production, and they are the main food source for several commercially produced fish species and crustaceans.

They are also increasingly part of waste management. Academic institutions in the UK successfully tested algae to remediate communal wastewater, runoff from dairy farms, and even toxic sludge from mining. The most lucrative application however is in the form of producing high-value products. Algae can also be used to produce a range of important chemicals from food additives, colorants, beauty products and drug components.

1.4 Our business model If algae outperform plants in most applications why are they not more readily produced? The answer is that algae are much less well researched than most crops. Companies that do try to grow algae most often start by adjusting parameters by hand, using alga strains randomly collected, and relying on outdated, scarce information for growth conditions. This leads to poor yields, slow progression and expensive setup costs. We aim to change this. The funding from the MBR space challenge enabled us to not only design our prototype, but to lay the foundations for a future business based around it. Our plan is to to create an alga strain development service. Our prototype alga growth reactor is fully automated, cheap to build, works on a small scale, and can control a range of parameters on the fly, making it a unique product on the market. By building an array of such reactors, we can quickly test hundreds of growth conditions for an alga strain of choice, or test several alga strains side by side and so can rapidly analyze culture conditions for companies wanting to optimize their alga production systems. This is currently done by hand or not done at all due to its prohibitive cost and lack of equipment, but our first market research revealed there would be a great demand for such a service. We believe that this alga strain development service would enable companies to successfully grow algae for a range of purposes. Most large scale alga production companies operate in environments where land is readily obtainable, sunlight is abundant and daytime temperatures are high. This puts the North Africa, the Gulf states, and the southern US in ideal location for this new industry. The latter is already home to the largest producers, but the alga biotech market is rapidly growing as this exciting new field develops. We are hoping to take part in shaping this new field as algae become important agricultural and industrial products on Earth and beyond.

2. Main results and outputs

2.1 photobioreactor design

Figure 2: Components of the photobioreactor. A 3D render of the photobioreactor was created to showcase the designed concept. Notable components are indicated by the arrows. The gas box is largely hidden from the front, and contains CO2, and O2 sensors, along with an additional heater and the inlets from the CO2 canister the gas from the culture vessel, and an outlet to either the pump, or to outside via a pressure release valve. The main deliverable of our project is the design of a novel, fully automated photobioreactor to grow algae in a closed environment. The reactor is designed to allow for algae to be grown in the smallest possible volume, while fully automating the culture procedure and measuring an unprecedented amount of parameters. For a short introduction, please refer to chapter 1.1. The reactor is based around the specifications of the 50ml Falcon tube as reaction vessel, which is a cylindrical container ubiquitously used in biomedical research. Since the tube is transparent, autoclavable, airtight, cheap and readily available in large quantities in any laboratory, it will instantly be familiar in any setting. The reactor features a 3D printed cap that attaches seamlessly to any standard Falcon tube and has airtight inlets for gas in, gas out, and the temperature probe. This allows any alga culture to be placed into - or removed from - the reactor within seconds and with minimal effort. The container holds maximum 50ml culture volume, making the reactor volume smaller than any design on the market. This is a clear

advantage as experimental alga cultures are often made on a small scale and growing them up the standard 1L volume often takes weeks or months before any analysis can begin. With a 20x small culture volume, the initial waiting time is significantly less and results can also be collected more rapidly. The cylindrical nature of the vessel also makes sure no corners of edges are present within the culture vessel which would lead to algae settling in corners or being subjected to uneven conditions inside the reactor vessel depending on exact position. By mimicking the cylindrical nature of larger alga bioreactors, the Falcon tube design provides an excellent starting point for upscaling to large scale production. The design is also made with parallelization in mind. Reactors can be chained together and can report to the same remote computer simultaneously. This enables to potentially run dozens of reactors in parallel, even with slightly different culture conditions. The remote operator can then establish which culture condition was the most advantageous or even different alga strains can be compared to compare their performance under the same culture conditions.

2.2 Sensors The well-being of the culture is estimated from multiple parameters, namely the turbidity sensor, the colour sensor, O2 production, and CO2 consumption. Additional sensors measure temperature in the growth chamber and the gas box separately to allow fine temperature control, and humidity of the gas phase to help calibrate all the light-based sensors in the gas phase. All sensors report data to the remote computer where it is stored and graphed for operators, and evaluated to activate feedback mechanisms if needed. The optical density (OD) of any microbial culture is commonly used for estimating culture density, and measuring it over time allows to gather information on cell division cycle time. This is considered a crucial parameter as a culture with dividing cells is a healthy culture, and increase of optical density is often considered the main parameter to optimize growth conditions for. For bacteria, light at 600nm is generally used to estimate optical density (OD600). For algae however this wavelength is suboptimal as chlorophyll also absorbs light at this wavelength. This can lead to an unwanted side effect of increasing OD600 numbers as the culture ages, as more and more chlorophyll is produced by the same number of cells, therefore the correlation between OD600 and cell numbers becomes weaker. A near infrared light at 750nm is much more suited to measure OD of algae as photosynthetic pigments prefer the visible spectrum, and any absorbance at 750nm is closely associated with the accumulation of biomass, rather than just pigments. For this reason, our turbidity sensor employs a 750nm sensor which should generally be appropriate for a wide range of algal species. The turbidity sensor itself is located on the outside of the culture chamber, and measures a beam of light travelling through the culture liquid from one side of the culture vessel to the other.

The second sensor to measure culture health is the colour sensor. While the previously described turbidity sensor estimates culture density over time, information also needs to be gathered about the health of the biomass that make up the culture. The colour sensor is able to detect red, green, blue, and white light with high accuracy. This enables the controller to get information on whether pigments are still being produced by the culture, which is indicative of nominal growth. The colour sensor itself reports back red, green, blue, and white light intensities as analogue frequency. It also filters out infrared light, reducing any background from the neighbouring turbidity sensor. It is able to use a built-in single LED to make measurements, or can make use of the LED panel that illuminates the reactor itself. Only a single sensor is located submerged into the alga culture itself: a thermistor. This measures the temperature of the liquid. We have considered to use either a thermocouple or a thermistor for this purpose and did a thorough comparison of both. The thermistor measures temperature change by changing resistance, while the thermocouple produces voltage. The former therefore requires power, while the latter does not. The power requirements for a thermistor are however minuscule, around 0.05mW, so both temperature sensor types are able to operate in our design. In terms of physical parameters, the thermistor is slightly bulkier than the thermocouple (which is essentially a wire) but thermistors are also readily produced in waterproof cases in a size that easily fits into our reactor without obstructing water flow or light. The additional electronics requirements are also slightly different between the two sensor types, but these are also not relevant for our purposes. The real difference between them comes in range and accuracy. Thermocouples tend to have a broad temperature range generally spanning hundreds of degrees C, while thermistors operate in the more modest -50C to +150C range. Since our reactor generally operates between +10C to +30C, the range of both sensors are equally acceptable. However, when it comes to accuracy, thermocouples are rarely more accurate then 1.0C, while thermistors can reach accuracies of 0.1C or below. This became the defining feature to influence our decision as in order to grow algae at accurately controlled temperatures, we wanted go below 1.0C in measurement accuracy, therefore we incorporated a thermistor into our design. This is yet another feature that is a direct upgrade to competing designs that we have seen to date which either employ thermistors or actual thermometers. Additional data is gathered from the gas phase. We focused on measuring gas components as this allows us to gather accurate data about culture performance while not needing to place actual sensors into the culture medium at the same time. Both the CO2 and O2 sensors in the gas box operate by using a small infrared sensors within the the sensor housing. This means measurements need to be corrected based on temperature and humidity, therefore we also installed additional sensors to measure these parameters, thereby allowing continuous corrections.

2.3 Attenuators The main parameters the reactor can control are light, temperature, airflow, and CO2 concentration. Light is a defining requirement for autotrophic growth, but algae species will have different different light requirements. Therefore we chose a panel of LEDs that emits blue, red, green and white light, with each colour intensity being adjustable separately. This allows a broad range of alga species to be grown in the reactor, while also allowing cutting down on energy costs for setups where only certain lights are required. The chosen LEDs are also specific: a combination of blue and red light are optimal for most land plants, however it has also been shown that adding additional green light is beneficial for growth, despite green light itself not being used by the photosystems themselves. The additional white light provides a broad spectrum alternative in cases where narrow wavelengths are not required, but instead high intensity full spectrum light is preferred. This is the only part of the prototype that we were not able to finish physically building in the given 6 month timeframe for this project. Temperature is controlled by a peltier-based heater-cooler located under the reaction chamber. Depending on the direction of the current, the same unit can perform both heating and cooling, thereby being able to effectively regulate temperature based on the thermistor readings inside the culture vessel. Heat (or cold) from the peltier reaches the culture tube via a silicone form specifically moulded to the shape of the tip of a 50ml Falcon tube. At the bottom of the heating unit is a heatsink to aid effective heat transfer between the heating unit and the environment. This is required for efficient heat transfer to the culture vessel itself. A second heater pad is located inside the gas chamber. The role of this is to keep the temperature of the gas inside the gas box always slightly above the temperature in the culture vessel. This is to prevent condensation within the gas box which would otherwise be detrimental to the electronics and the sensors inside the box. The air pump is required to provide the means to circulate gas between the gas box and the reactor vessel. Also, the air pump ensures that gas is not only pumped into the reactor vessel, but is also pumped to the bottom of the alga culture. This leads to the constant bubbling of gas through the culture medium which is required for efficient gas exchange between the liquid and gas phases, and also serves to agitate the culture thereby allowing continuous mixing. Bubble columns are widely used in even large scale bioreactors as they provide an efficient but gentle means of ensuring algae do not settle to the bottom of the container and nutrients are distributed equally inside the medium. The continuous flow of algae also makes sure they receive appropriate lighting by not being constantly near the wall where the light enters the reactor chamber and where it may be higher than optimal. Also, algae shield one another, therefore in a resting culture, algae that are not in the top layer

facing the sunlight get gradually less light inside the culture medium, but the continuous mixing ensures individual cells are not stuck in a low light environment either. A special feature of this reactor design is the inclusion of a CO2 source. This system is in place to model the atmosphere of Mars, which is 96% CO2. Essentially when gas is pumped from the canister into the culture, this is modelling what would happen if gas from the Martian atmosphere was compressed, heated, and added into the culture vessel to provide an in-situ carbon source for the culture. This is implemented by a CO2 canister that is installed into the reactor that can deliver gaseous CO2 into the gas box through a gas valve controlled by a stepper motor. The controlled inlet allows changing CO2 concentration either at will, or according to any preset program. Since the amount of released CO2 is always known and is instantly measured upon entering the gas chamber, CO2 production by the algae can be measured by the same CO2 sensor accurately, and can instantly be corrected by the amount coming in from the canister. It is well documented that plants and algae are able to grow much more rapidly in an environment where CO2 concentrations are higher than in Earth’s atmosphere, and this system allows testing this feature extensively. It also ensures that future, upscaled versions of the reactor can deliver yields significantly higher than similarly sized open pond systems that rely on CO2 uptake from the atmosphere.

2.4 The case for growing life on Mars Humanity currently eyes Mars in its attempts to establish the first permanent base outside Earth. To achieve this, in situ resource utilization (ISRU) is one of the defining requirements. If a future Mars outpost has to be regularly refuelled from Earth, or the initial mission has to transfer enough food and water for years of operation, then the cost of logistics rapidly becomes prohibitive. Therefore, a method to grow food on Mars using local resources would be a game changer in space colonization.

Figure 3: relevant elements on Mars. Martian atmosphere is composed mostly of CO2 with 2-2% of either nitrogen and argon, while the regolith is abundant in several elements, but it notably contains water (ice, and in minerals) a high amount of Sulphur, and some amount of phosphorous.

In order to grow plants, the basic building blocks of life: C,H,N,O,P and S have to be sourced and made available for a proposed Martian farm. These are also often referred to as macro elements for agriculture. Meso elements are Ca and Mg which are also crucial for enzymatic reactions but are required in lesser amounts, while microelements add up to a much longer list, but are needed in such small quantities even for large scale agricultural operations that they are easily transported from Earth as part of the mission cargo. Macro and meso elements however need to be sourced locally if possible. A quick look at the composition of Mars reveals it as a diverse world with a wide array of resources. Water ice has been discovered in recent years, and there is even evidence of large, frozen underground lakes on the red planet. This could provide a source of the much needed H and O in a ready-to-use molecular form of water. Carbon is available from the atmosphere: 96% of the Martian atmosphere is CO2, it merely needs to be compressed and heated to be used, making it the most easily available element for plant (or algal) growth. Sulphur is available in surprising abundance on Mars, it is even more common in the Martian regolith than on Earth. In fact, it is such a common component that it has been proposed to be used as the major building block in Martian concrete. Nitrogen is less readily available on Mars than on Earth. While it is present, its relative abundance is low. In the regolith, no notable nitrogen deposits have been found to date, leaving the only source of N2 the atmosphere. It has approximately 2% N2 content, compared to the 78% on Earth. However this is still sufficient as atmospheric gas is easily harvestable, and the other major component - CO2 - can be readily sequestered by a simple, easily regenerating Calcium based redox system. Such a system would allow rapid generation of almost pure N2 in near infinite amounts. The only consideration in this respect is that N2 is not readily usable as a nitrogen source to most plants as most autotrophs can only metabolize nitrates and amines, but not the nitrogen molecule itself. Cyanobacteria are one of the very few exceptions to this as via heterocysts the are able to capture nitrogen by breaking up the triple bond holding the N2 molecule together. This makes them superior to any plant and most other organisms and ideally suited to be the pioneer species to cultivate on Mars, that allows the creation of nitrates for subsequent organisms. The final required building block is phosphorus. This element has been detected in the Martian regolith, but it would need to be mined and the efficiency of such a process is currently questionable as no extraction method has been proposed to date. Bringing some amount of phosphorous from Earth in the form of fertilizer may therefore be a requirement of the first settlement on Mars. The alternative is to recycle human waste. This is abundant in phosphorus - similar to most other types of manure - which is why it is primarily employed as soil additive on Earth as well. The continuous recycling of human waste as fertilizer brings additional challenges for a mission to Mars, which is beyond the scope of this document, however, complete waste recycling is likely to be a requirement for such a mission anyway in order to conserve water, heat, and organics in general.

Meso elements Ca and Mg are in contrast abundantly available in regolith and can be readily mined if required in large quantities. Apart from the main building blocks, the list of microelements is much longer, and some of them have not been extensively studied in the Martian regolith yet. Whether they are present at all is however of lesser concern as the tiny quantities can be supplied from terrestrial sources. In summary, all main building blocks for life are present on Mars and are easily accessible. Of note, nitrogen is mostly available in molecular form and is therefore primarily available to cyanobacteria, and phosphorous supplies are scarce, but obtainable.

2.5 Algae: the best candidate for cultivation Having all material available for life does not mean any organism can flourish in a Martian setting. Conditions on the red planet are notably different from Earth, and the significantly colder temperatures, lack of liquid water, low pressure, comparably high radiation, and the abundance of perchlorates in regolith in combination mean that no life can survive and develop on Mars for prolonged periods of time. However, in a protected container where environmental conditions are controlled, suddenly a plethora of organisms are available to grow. The possible candidates to grow on Mars can quickly be reduced to autotrophs: heterotrophs would require pre existing organics in large abundance, which are not yet present on Mars. Chemoautotrophs can also be ruled out as they rarely have direct applications in agriculture on Earth. This is unfortunate as otherwise they would be interesting candidates with rapid growth cycles, only inorganic inputs, and generally an extremophile nature which enables them to live hostile environments. However, due to the lack of human use for these organisms, they are not part of the present document. This leaves photoautotrophs - plants and algae - as the main candidates. Out of the two groups, algae have a number of notable advantages. When comparing regular crop plants like rice, wheat, maize or Mark Watney's famous potatoes to either leafy greens or algae, the first notable difference is that the biomass produced by crop plants is largely inedible. Stems, roots and leaves on regular crops are not readily consumed by humans and so the energy that was invested into making them is lost after harvest. The ideal plant should be almost fully edible. While leafy greens excel in this respect as lettuce would have all of its leaves consumed, they still produce roots and therefore about 10% of their biomass cannot be consumed. Algae are almost 100% edible on the other hand as the entire algal cell is part of human diet, which makes them a significantly more efficient organism to grow than any plant.

Figure 4: comparison of organisms to grow on Mars. These selected criteria were chosen to evaluate performance of agriculturally relevant organism groups for cultivation in a controlled, resource limited environment The second number to take into consideration is land use. While land use is based on terrestrial measurements generally in open field cultivation and is therefore not directly applicable to Mars, it still serves as a good indicator of biomass yield and culture density. Crop plants are carefully adapted to monoculture growth and therefore use land fairly effectively, that is, to deliver 317GJ of energy, they only need 1.3 hectares of land, much less than leafy greens would. Algae however grow into such dense cultures that that they are even able to surpass crops in this respect and can generate biomass with the minimal footprint. A key advantage on a resource-limited location like Mars, where constructing growth chambers is an inefficient and resource intensive procedure. Producing edible biomass is not the only concern when growing crops, the quality of the biomass is also important. Plants tend to store energy in the form of carbohydrates, generally either sugar or starch leading to a carbohydrate-heavy diet. While this is not necessarily disadvantageous, a balanced diet featuring ample protein is generally preferred, where again algae are provide more options. Depending on the species, up to 40% of the produced biomass can be protein with the remainder being lipids and carbohydrates, providing a more varied food and feedstock than a given crop plant. The final characteristic we considered important when choosing what to grow at a future Martian settlement was light energy harvesting efficiency. Sunlight on Mars is about 40% weaker than on Earth. While that is still sufficient for most autotrophs even light intensity on Earth can be below optimal ideal growth of a chosen

organism. Therefore artificial light is preferred which adds a significant energy burden to the Martian colony. Depending on how energy is produced, this may be a significant limit along with heating to determine culture chamber size. Therefore anything that will be grown has to be able to use the light with the highest efficiency possible. Here again algae come out on top. Not only because individual algal cells are more efficient in converting electromagnetic energy to chemical energy, but also because the liquid alga culture can cover 100% of the culture chamber floor, whereas a leafy plant will always have dead areas where the light shines between the leaves and hits the floor without any particular use. The microscopic nature of algae and their ability to grow in liquid medium is further advantageous for automation purposes. Planting, watering, harvesting and processing plants requires complex machinery that is able to accommodate the unique geometry of plants. Algae on the other hand can be handled as liquid or as paste leading to simpler mechanics and less chance of failure. In summary, we have performed an extensive literature search and consulted with leading experts in the field to be able to assess which organism is best suited for cultivation on Mars. Our results indicate that algae are superior in terms of efficiency for culturing on a future Martian base. Mars itself harbours all the required building blocks needed for growth, most of which are abundant, and easily obtainable. Apart from the required elements, growing terrestrial organisms requires terrestrial conditions. Life on Earth is mostly suited to the specific temperature range, pressure, radiation, and lighting conditions that are familiar on the blue planet. Therefore any agricultural operation elsewhere has to recreate these conditions. Our bioreactor aims to provide appropriate temperature, pressure, and light and therefore demonstrates that if placed in an appropriate radiation shield and supplied with energy, in theory it can harbor life on Mars. And as all crucial elements for life are also available locally together they provide a strong case for growing algae on Mars. This is our main conclusion from this project.

3. obstacles or changes in direction during the project

3.1 finding the appropriate culture vessel After the first iteration, our photobioreactor featured a a cell culture flask as a culture vessel instead of the falcon tube as seen in the current design. This was originally chosen as cell culture flasks are also readily used to grow algae and our engineers also favoured flat surfaces on all sides which enabled the convenient positioning of sensors and the heater. However, the cell culture flasks also came with a number of drawbacks. First, we were unable to obtain an accurate drawing of such a vessel,

which led to us measuring parameters by hand. It was then that we realized that cell culture flasks by different suppliers are not the same, and therefore our design would need to be used with one specific supplier to be able to fit into the reactor seamlessly. Since we did not want to commit to a specific brand at this stage, we instead had to find another container that is supplier-independent. The flat surfaces also lead to the appearance of edges and corners in the reaction vessel, which would have lead to building up cells in areas the bubbler would not easily reach. Finally, when consulting with potential customers, they

unanimously preferred a smaller culture volume and round containers, or wanted to have a shaker mechanism for the whole vessel to allow continuous agitation of all parts of the culture. Therefore, after having these discussions we changed the culture vessel to the falcon tube, which proved much more suited to our application.

3.2 alga species considerations When designing this project, we originally wanted to evaluate the feasibility to grow either Chlorella vulgaris or Arthrospira platensis. These two species were chosen as they were employed by the Russian and the US space agency respectively as the choice of algae to grow in space. They are also both known, widely cultivated and edible species, therefore ideal candidates for further study. However, upon completing the literature review, we were confronted with the fact that nitrogen availability on Mars is limited if we are unable to harvest atmospheric N2. We therefore also added members of the Nostocaceae family to the list of possible algae to grow as they are readily capable of using atmospheric N2, can grow rapidly, and are photoautotrophic cyanobacteria. Nostocs are however notable for producing alga toxins which lead to mass fish poisonings and are potentially harmful for humans as well. They are also yet untested as food source for humans. We have therefore decided to approach the Scottish Culture Collection of Algae where they tested toxin production in several strains and have identified some toxin free cultivars. The exact strain numbers however have not been chosen by the time of writing, therefore finding the perfect strain is still underway and will likely only to be finalized after being tested in the prototype reactor.

Figure 5: Render of the first photo-bioreactor prototype designed for this project. This preliminary featured a different sensor layout and a rectangular culture flask. It was discontinued in favour of the Falcon-based design

4) potential impact and opportunities for implementation of the results Our study demonstrates that Mars does contain all the resources needed for automated agriculture and we propose algae as the ideal candidate to grow. We also designed an automated photobioreactor to demonstrate the principle of how cultivation could operate. We demonstrated our results at the PHYCONET annual conference, the largest gathering of alga researchers in the UK, with great success. Our results altogether add important detail to plans of space agencies that aim to establish human presence on Mars. Building on these results can pave the way for providing food and feed permanently on the red planet, which would be a historical event. If we managed to add even the tiniest detail to that achievement, then we would consider our study to be of very high impact. In addition, our automated photobioreactor can also be used in terrestrial applications. As it can essentially measure alga culture parameters with an unprecedented accuracy, in a small volume, and with high speed, it provides a testbed for a wide array of algal strains that can be evaluated for a plethora of biotech applications. We consider this aspect of our reactor so important that we aim to create a company for this purpose. Therefore, a potential application of our design would be to use it as a basis for a biotech start-up. With it we aim to test algae for novel applications that can deliver new medicines, more efficient industrial processes, recycle waste, or bring food to areas where traditional agriculture is not possible.

5) conclusion and next steps This six-month project saw the development of an idea into a four-man team, a novel, innovative photobioreactor design, and a complete business plan. We have performed an extensive literature review and consulted with other professions to elucidate a feasible approach to grow algae on Mars. Then we designed a new, automated photobioreactor to grow algae from scratch, which can serve as a proof of concept prototype to demonstrate the principles of growing algae on Mars. The support given by the MBR Space Challenge has enabled us to lay the foundations for a new biotech company. Next month we are beginning to pitch our business plan to investors and plan to attract additional funding to build our first functioning prototypes. With additional funding in hand by the end of the year we will be in position to create a minimum viable product and step on the market in 2020. We aim to be on the forefront of the rapidly developing alga biotech sector and hope to take part in the establishment of novel alga cultivation pipelines on Earth and Mars. Do not hesitate to get in touch with us if you share our vision.