Integration of waste processing and biomass production systems as part of the KSC Breadboard project.

After initial emphasis on large-scale baseline crop tests, the Kennedy Space Center (KSC) Breadboard project has begun to evaluate long-term operation of the biomass production system with increasing material closure. Our goal is to define the minimum biological processing necessary to make waste streams compatible with plant growth in hydroponic systems, thereby recycling nutrients into plant biomass and recovering water via atmospheric condensate. Initial small and intermediate-scale studies focused on the recycling of nutrients contained in inedible plant biomass. Studies conducted between 1989-1992 indicated that the majority of nutrients could be rapidly solubilized in water, but the direct use of this crop "leachate" was deleterious to plant growth due to the presence of soluble organic compounds. Subsequent studies at both the intermediate scale and in the large-scale Biomass Production Chamber (BPC) have indicated that aerobic microbiological processing of crop residue prior to incorporation into recirculating hydroponic solutions eliminated any phytotoxic effect, even when the majority of the plant nutrient demand was provided from recycled biomass during long term studies (i.e. up to 418 days). Current and future studies are focused on optimizing biological processing of both plant and human waste streams.     

Dynamics of microorganism populations in recirculating nutrient solutions.

This overview covers the basic microbial ecology of recirculating hydroponic solutions. Examples from NASA and Soviet CELSS tests and the commercial hydroponic industry will be used. The sources of microorganisms in nutrient solutions include air, water, seeds, plant containers and plumbing, biological vectors, and personnel. Microbial fates include growth, death, and emigration. Important microbial habitats within nutrient delivery systems are root surfaces, hardware surfaces (biofilms), and solution suspension. Numbers of bacteria on root surfaces usually exceed those from the other habitats by several orders of magnitude. Gram negative bacteria dominate the microflora with fungal counts usually much lower. Trends typically show a decrease in counts with increasing time unless stressed plants increase root exudates. Important microbial activities include carbon mineralization and nitrogen transformations. Important detrimental interactions include competition with plants, and human and plant pathogenesis.     

Comparison of aerobically-treated and untreated crop residue as a source of recycled nutrients in a recirculating hydroponic system.

This study compared the growth of potato plants on nutrients recycled from inedible potato biomass. Plants were grown for 105 days in recirculating, thin-film hydroponic systems containing four separate nutrient solution treatments: (1) modified half-strength Hoagland's (control), 2) liquid effluent from a bioreactor containing inedible potato biomass, 3) filtered (0.2 micrometer) effluent, and 4) the water soluble fraction of inedible potato biomass (leachate). Approximately 50% of the total nutrient requirement in treatments 2-4 were provided (recycled) from the potato biomass. Leachate had an inhibitory effect on leaf conductance, photosynthetic rate, and growth (50% reduction in plant height and 60% reduction in tuber yield). Plants grown on bioreactor effluent (filtered or unfiltered) were similar to the control plants. These results indicated that rapidly degraded, water soluble organic material contained in the inedible biomass, i.e., material in leachate, brought about phytotoxicity in the hydroponic culture of potato. Recalcitrant, water soluble organic material accumulated in all nutrient recycling treatments (650% increase after 105 days), but no increase in rhizosphere microbial numbers was observed.     

Hydroponic potato production on nutrients derived from anaerobically-processed potato plant residues.

Bioregenerative methods are being developed for recycling plant minerals from harvested inedible biomass as part of NASA's Advanced Life Support (ALS) research. Anaerobic processing produces secondary metabolites, a food source for yeast production, while providing a source of water soluble nutrients for plant growth. Since NH4-N is the nitrogen product, processing the effluent through a nitrification reactor was used to convert this to NO3-N, a more acceptable form for plants. Potato (Solanum tuberosum L.) cv. Norland plants were used to test the effects of anaerobically-produced effluent after processing through a yeast reactor or nitrification reactor. These treatments were compared to a mixed-N treatment (75:25, NO3:NH4) or a NO3-N control, both containing only reagent-grade salts. Plant growth and tuber yields were greatest in the NO3-N control and yeast reactor effluent treatments, which is noteworthy, considering the yeast reactor treatment had high organic loading in the nutrient solution and concomitant microbial activity.     

Basic design concept of Closed Ecology Experiment Facilities.

In order to study the relationship between the physiological metabolism of living things and the environmental factors such as the atmospheric contents and so on within the closed ecosystem, Closed Ecology Experiment Facilities (CEEF) were designed and now under construction based on the following concepts: (1) Individual sealed chambers (called modules) for the plant cultivation, animal breeding, human habitation and microbial waste treatment are to be constructed independently to be able to study the metabolic effects of each living thing on the environmental factors within closed ecosystem. (2) A chamber for the microbial waste treatment are to be replaced with two systems; wet oxidation reactors and chemical nitrogen fixation reactors. (3) Atmospheric control systems are to be independently attached to each module for stabilizing atmospheric contents in each module. (4) Any construction materials having the possibility to absorb oxygen and carbon dioxide are to be prohibited to use in each module for sustaining material balance. (5) Facilities have to be developed so that the closed plant and animal experiments can be done independently, as well as integrated experiments with plants and animals through exchanging foods, carbon dioxide, oxygen, condensed water and nutrient solution.     

Significance of rhizosphere microorganisms in reclaiming water in a CELSS.

Plant-microbe interactions, such as those of the rhizosphere, may be ideally suited for recycling water in a Controlled Ecological Life Support System (CELSS). The primary contaminant of waste hygiene water will be surfactants or soaps. We identified changes in the microbial ecology in the rhizosphere of hydroponical1y grown lettuce during exposure to surfactant. Six week old lettuce plants were transferred into a chamber with a recirculating hydroponic system. Microbial density and population composition were determined for the nutrient solution prior to introduction of plants and then again with plants prior to surfactant addition. The surfactant Igepon was added to the recirculating nutrient solution to a final concentration of 1.0 g L-1. Bacteria density and species diversity of the solution were monitored over a 72-h period following introduction of Igepon. Nine distinct bacterial types were identified in the rhisosphere; three species accounted for 87% of the normal rhizosphere population. Microbial cell number increased in the presence of Igepon, however species diversity declined. At the point when Igepon was degraded from solution, diversity was reduced to only two species. Igepon was found to be degraded directly by only one species found in the rhizosphere. Since surfactants are degraded from the waste hygiene water within 24 h, the potential for using rhizosphere bacteria as a waste processor in a CELSS is promising.     

Advantages of using subsurface flow constructed wetlands for wastewater treatment in space applications: ground-based Mars Base prototype.

Research and design of subsurface flow wetland wastewater treatment systems for a ground-based experimental prototype Mars Base facility has been carried out, using a subsurface flow approach. These systems have distinct advantages in planetary exploration scenarios: they are odorless, relatively low-labor and low-energy, assist in purification of water and recycling of atmospheric CO2, and will support some food crops. An area of 6-8 m2 may be sufficient for integration of wetland wastewater treatment with a prototype Mars Base supporting 4-5 people. Discharge water from the wetland system will be used as irrigation water for the agricultural crop area, thus ensuring complete recycling and utilization of nutrients. Since the primary requirements for wetland treatment systems are warm temperatures and lighting, such bioregenerative systems may be integrated into early Mars base habitats, since waste heat from the lights may be used for temperature maintenance in the human living environment. "Wastewater gardens (TM)" can be modified for space habitats to lower space and mass requirements. Many of its construction requirements can eventually be met with use of in-situ materials, such as gravel from the Mars surface. Because the technology requires little machinery and no chemicals, and relies more on natural ecological mechanisms (microbial and plant metabolism), maintenance requirements are minimized, and systems can be expected to have long operating lifetimes. Research needs include suitability of Martian soil and gravel for wetland systems, system sealing and liner options in a Mars Base, and wetland water quality efficiency under varying temperature and light regimes.     

The structure and function of microbial communities in recirculating hydroponic systems.

Strategies to control the microbial community associated with plant growth systems need to be based on a fundamental understanding of the factors which structure and regulate the community. Spatial and temporal patterns in the abundance and production rate of microorganisms in hydroponic systems containing wheat were examined to evaluate how root-derived carbon is processed. The relevance of results to monitoring and control strategies is discussed.     

Production characteristics of the “higher plants–soil-like substrate” system as an element of the bioregenerative life support system

The study addresses the possibility of long-duration operation of a higher plant conveyor, using a soil-like substrate (SLS) as the root zone. Chufa (Cyperus esculentus L.), radish (Raphanus sativus L.), and lettuce (Lactuca sativa L.) were used as study material. A chufa community consisting of 4 age groups and radish and lettuce communities consisting of 2 age groups were irrigated with a nutrient solution, which contained mineral elements extracted from the SLS. After each harvest, inedible biomass of the harvested plants and inedible biomasses of wheat and saltwort were added to the SLS. The amounts of the inedible biomasses of wheat and saltwort to be added to the SLS were determined based on the nitrogen content of the edible mass of harvested plants. CO₂ concentration in the growth chamber was maintained within the range of 1100–1700 ppm. The results of the study show that higher plants can be grown quite successfully using the proposed process of plant waste utilization in the SLS. The addition of chufa inedible biomass to the SLS resulted in species-specific inhibition of growth of both cultivated crops and microorganisms in the “higher plants – SLS” system. There were certain differences between the amounts of some mineral elements removed from the SLS with the harvested edible biomass and those added to it with the inedible biomasses of wheat and saltwort.     

Use of biologically reclaimed minerals for continuous hydroponic potato production in a CELSS.

Plant-derived nutrients were successfully recycled in a Controlled Ecological Life Support System (CELSS) using biological methods. The majority of the essential nutrients were recovered by microbiologically treating the plant biomass in an aerobic bioreactor. Liquid effluent containing the nutrients was then returned to the biomass production component via a recirculating hydroponic system. Potato (Solanum tuberosum L.) cv. Norland plants were grown on those nutrients in either a batch production mode (same age plants on a nutrient solution) or a staggered production mode (4 different ages of plants on a nutrient solution). The study continued over a period of 418 days, within NASA Breadboard Project's Biomass Production Chamber at the Kennedy Space Center. During this period, four consecutive batch cycles (104-day harvests) and 13 consecutive staggered cycles (26-day harvests) were completed using reclaimed minerals and compared to plants grown with standard nutrient solutions. All nutrient solutions were continually recirculated during the entire 418 day study. In general, tuber yields with reclaimed minerals were within 10% of control solutions. Contaminants, such as sodium and recalcitrant organics tended to increase over time in solutions containing reclaimed minerals, however tuber composition was comparable to tubers grown in the control solutions.     

Recycling of inorganic nutrients for hydroponic crop production following incineration of inedible biomass.

The goal of resource recovery in a regenerative life support system is maintenance of product quality to sure support of reliable and predictable levels of life support function performance by the crop plant component. Further, these systems must be maintained over extended periods of time, requiring maintenance of nutrient solutions to avoid toxicity and deficiencies. The focus of this study was to determine the suitability of the ash product following incineration of inedible biomass as a source of inorganic nutrients for hydroponic crop production. Inedible wheat biomass was incinerated and ash quality characterized. The incinerator ash was dissolved in adequate nitric acid to establish a consistent nitrogen concentration is all nutrient solution treatments. Four experimental nutrient treatments were included: control, ash only, ash supplemented to match the control treatment, and ash only quality formulated with reagent grade chemicals. When nutrient solutions were formulated using only ash following incineration of inedible biomass, a balance in solution is established representing elemental retention following incineration and nutrient proportions present in the original biomass. The resulting solution is not identical to the control. This imbalance resulted in a suppression of crop growth. When the ash is supplemented with reagent grade chemicals to establish the same balance as in the control--growth is identical to the control. The ash appears to carry no phytotoxic materials. Growth in solution formulated with reagent grade chemicals but matching the quality of the ash only treatment resulted in similar growth to that of the ash only treatment. The ash product resulting from incineration of inedible biomass appears to be a suitable form for recycle of inorganic nutrients to crop production.     

Accumulation and effect of volatile organic compounds in closed life support systems.

Bioregenerative life support systems (BLSS) being considered for long duration space missions will operate with limited resupply and utilize biological systems to revitalize the atmosphere, purify water, and produce food. The presence of man-made materials, plant and microbial communities, and human activities will result in the production of volatile organic compounds (VOCs). A database of VOC production from potential BLSS crops is being developed by the Breadboard Project at Kennedy Space Center. Most research to date has focused on the development of air revitalization systems that minimize the concentration of atmospheric contaminants in a closed environment. Similar approaches are being pursued in the design of atmospheric revitalization systems in bioregenerative life support systems. in a BLSS one must consider the effect of VOC concentration on the performance of plants being used for water and atmospheric purification processes. In addition to phytotoxic responses, the impact of removing biogenic compounds from the atmosphere on BLSS function needs to be assessed. This paper provides a synopsis of criteria for setting exposure limits, gives an overview of existing information, and discusses production of biogenic compounds from plants grown in the Biomass Production Chamber at Kennedy Space Center.     

Use of halophytic plants for recycling NaCl in human liquid waste in a bioregenerative life support system

The purpose of this work was to develop technology for recycling NaCl containing in human liquid waste as intrasystem matter in a bioregenerative life support system (BLSS). The circulation of Na⁺ and Cl⁻ excreted in urine is achieved by inclusion of halophytes, i.e. plants that naturally inhabit salt-rich soils and accumulate NaCl in their organs. A model of Na⁺ and Cl⁻ recycling in a BLSS was designed, based on the NaCl turnover in the human–urine–nutrient solution–halophytic plant–human cycle. The study consisted of (i) selecting a halophyte suitable for inclusion in a BLSS, and (ii) determining growth conditions supporting maximal Na⁺ and Cl⁻ accumulation in the shoots of the halophyte growing in a nutrient solution simulating mineralized urine. For the selected halophytic plant, Salicornia europaea, growth rate under optimal conditions, biomass production and quantities of Na⁺ and Cl⁻ absorbed were determined. Characteristics of a plant production conveyor consisting of S.europaea at various ages, and allowing continuity of Na⁺ and Cl⁻ turnover, were estimated. It was shown that closure of the NaCl cycle in a BLSS can be attained if the daily ration of fresh Salicornia biomass for a BLSS inhabitant is approximately 360 g.     

Current concepts and future directions of CELSS.

Studies of bioregenerative life support systems for use in space indicate that they are scientifically feasible. Preliminary data suggest that they would provide cost- and weight-saving benefits for low Earth orbit, long duration space platforms. Concepts of such systems include the use of higher plants and/or micro-algae as sources of food, potable water and oxygen, and as sinks for carbon dioxide and metabolic wastes. Recycling of materials within the system will require processing of food organism and crew wastes using microbiological and/or physical chemical techniques. The dynamics of material flow within the system will require monitoring, control, stabilization and maintenance imposed by computers. Future phases of study will continue investigations of higher plant and algal physiology, environmental responses, and control; flight experiments for testing responses of organisms to weightlessness and increased radiation levels; and development of ground-based facilities for the study of recycling within a bioregenerative life support system.     

Canadian advanced life support capacities and future directions

Canada began research on space-relevant biological life support systems in the early 1990s. Since that time Canadian capabilities have grown tremendously, placing Canada among the emerging leaders in biological life support systems. The rapid growth of Canadian expertise has been the result of several factors including a large and technically sophisticated greenhouse sector which successfully operates under challenging climatic conditions, well planned technology transfer strategies between the academic and industrial sectors, and a strong emphasis on international research collaborations. Recent activities such as Canada’s contribution of the Higher Plant Compartment of the European Space Agency’s MELiSSA Pilot Plant and the remote operation of the Arthur Clarke Mars Greenhouse in the Canadian High Arctic continue to demonstrate Canadian capabilities with direct applicability to advanced life support systems. There is also a significant latent potential within Canadian institutions and organizations with respect to directly applicable advanced life support technologies. These directly applicable research interests include such areas as horticultural management strategies (for candidate crops), growth media, food processing, water management, atmosphere management, energy management, waste management, imaging, environment sensors, thermal control, lighting systems, robotics, command and data handling, communications systems, structures, in-situ resource utilization, space analogues and mission operations. With this background and in collaboration with the Canadian aerospace industry sector, a roadmap for future life support contributions is presented here. This roadmap targets an objective of at least 50% food closure by 2050 (providing greater closure in oxygen, water recycling and carbon dioxide uptake). The Canadian advanced life support community has chosen to focus on lunar surface infrastructure and not low Earth orbit or transit systems (i.e. microgravity applications). To advance the technical readiness for the proposed lunar missions, including a lunar plant growth lander, lunar “salad machine” (i.e. small scale plant production unit) and a full scale lunar plant production system, a suite of terrestrial developments and analogue systems are proposed. As has been successfully demonstrated by past Canadian advanced life support activities, terrestrial technology transfer and the development of highly qualified personnel will serve as key outputs for Canadian advanced life support system research programs. This approach is designed to serve the Canadian greenhouse industry by developing compliance measures for mitigating environmental impact, reducing labour and energy costs as well as improving Canadian food security, safety and benefit northern/remote communities.     

Changes in hyperspectral reflectance signatures of lettuce leaves in response to macronutrient deficiencies

The adaptation of specific remote sensing and hyperspectral analysis techniques for the determination of incipient nutrient stress in plants could allow early detection and precision supplementation for remediation, important considerations for minimizing mass of advanced life support systems on space station and long term missions. This experiment was conducted to determine if hyperspectral reflectance could be used to detect nutrient stress in Lactuca sativa L. cv. Black Seeded Simpson. Lettuce seedlings were grown for 90 days in a greenhouse or growth chamber in vermiculite containing modified Hoagland’s nutrient solution with key macronutrient elements removed in order to induce a range of nutrient stresses, including nitrogen, phosphorus, potassium, calcium, and magnesium. Leaf tissue nutrient concentrations were compared with corresponding spectral reflectances taken at the end of 90 days. Spectral reflectances varied with growing location, position on the leaf, and nutrient deficiency treatment. Spectral responses of lettuce leaves under macronutrient deficiency conditions showed an increase in reflectance in the red, near red, and infrared wavelength ranges. The data obtained suggest that spectral reflectance shows the potential as a diagnostic tool in predicting nutrient deficiencies in general. Overlapping of spectral signatures makes the use of wavelengths of narrow bandwidths or individual bands for the discrimination of specific nutrient stresses difficult without further data processing.     

Progress in ultrasonic bioreactors for CELSS applications.

An important issue in Controlled Ecological Life Support Systems (CELSS) is the recycling of inedible crop residues to recover inorganic plant nutrients such as nitrates, phosphates, potassium and other macro- and micro-nutrients. In a closed system in space, such regeneration is vital to the long term viability of plant growth necessary for the food production and waste handling process. Chemical approaches to recycling such as incineration and wet oxidation are not compatible with low energy and environmentally friendly regeneration of such nutrients. Biological regeneration is more acceptable environmentally, but it is a very slow process and does not typically result in complete recovery of inorganic and organic nutrients. A new approach to biological regeneration is described here involving the combined use of special enzymatic catalysts and ultrasonic energy in a bioreactor system. This new system has the potential for rapid, efficient, environmentally friendly and complete conversion of crop wastes to inorganic plant nutrients and food recovery from cellulose materials. A series of experimental tests were carried out with a soybean crop residue meal substrate. Biochemical conversion rates were significantly expedited with the addition of enzymes and further enhanced through ultrasonic stimulation of these enzymes. The difference in conversion rates was particularly increased after the initial period of soluble organics conversion. The remaining cellulose substrate is much more difficult to biodegrade, and the ultrasonically-enhanced reaction was able to demonstrate a much higher rate of substrate conversion.     

Management and control of microbial populations' development in LSS of missions of different durations.

The problem of interaction between man and microorganisms in closed habitats is an inextricable part of the whole problem of co-existence between macro- and microorganisms. Concerning the support of human life in closed habitat, we can, conventionally, divide microorganisms, acting in life support system (LSS) into three groups: useful, neutral and harmful. The tasks, for human beings for optimal coexistence with microhabitants seem to be trivial: (1) to increase the activity of useful forms, (2) decrease the activity harmful forms, (3) not allow the neutral forms to become the harmful ones and even to help them to gain useful activity. The task of efficient management and control of microbial population's development in LSS highly depends on mission duration. As for short-term missions without recycling, the proper hygienic procedures are developed. For longer missions, the probability of transformation of the neutral forms into the harmful ones is becoming more dangerous. The LSS for long-term missions are to use cycling-recycling systems, including system with biological recycling. In these systems, microbial populations as regenerative link should be useful and active agents. Some problems of microbial populations control and management are discussed in the paper.     

Soybean cultivation for Bioregenerative Life Support Systems (BLSSs): The effect of hydroponic system and nitrogen source

Soybean [Glycine max (L.) Merr.] is one of the plant species selected within the European Space Agency (ESA) Micro-Ecological Life Support System Alternative (MELiSSA) project for hydroponic cultivation in Biological Life Support Systems (BLSSs), because of the high nutritional value of seeds. Root symbiosis of soybean with Bradirhizobium japonicum contributes to plant nutrition in soil, providing ammonium through the bacterial fixation of atmospheric nitrogen. The aim of this study was to evaluate the effects of two hydroponic systems, Nutrient Film Technique (NFT) and cultivation on rockwool, and two nitrogen sources in the nutrient solution, nitrate (as Ca(NO₃)₂ and KNO₃) and urea (CO(NH₂)₂), on root symbiosis, plant growth and seeds production of soybean. Plants of cultivar ‘OT8914’, inoculated with B. japonicum strain BUS-2, were grown in a growth chamber, under controlled environmental conditions.