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All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Editorial: Phenylpropanoid Systems Biology and Biotechnology

Phenylpropanoids are specialized metabolites involved in several aspects of plant growth and development and in the responses of plants to environmental stimuli. These compounds are synthesized from key intermediates of the shikimate pathway, which are structurally modified by the combined activities of lyases, transferases, ligases, reductases and oxygenases, resulting in the organ- and developmental-specific synthesis and accumulation of diverse metabolites (Vogt, 2010). The phenylpropanoid pathway provides the building blocks for lignin, suberin, and condensed tannins that play a role in structural support and mechanical strength. Lignin is a major contributor to feedstock recalcitrance and negatively affects the conversion of plant biomass into downstream products in biorefineries (Liu et al., 2021). Further, this pathway is key for the production of anthocyanins for organ pigmentation, flavonols and flavones for UV protection, various flavonoids and isoflavonoids for plant-microbe interactions, and antimicrobial phytoalexins for protection against pathogens (Deng and Lu, 2017). In addition to their biological functions in planta, phenylpropanoids are economically important metabolites. They constitute important components in the human diet, acting as nutraceutical compounds with antioxidant, chemopreventive, antimitotic, neuroprotective, cardioprotective, and anti-inflammatory activities. Several phenylpropanoids are considered high-value biochemicals employed in the production of fragrances, pharmaceuticals and biopolymers (Lin and Eudes, 2020).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Innovation and Winemaking By-Product Valorization: An Ohmic Heating Approach

The by-products of the winemaking process can represent chances for the development of new products. This study focused on the “zero waste” strategy development for by-products generated within winemaking from white and red grape varieties cultivated in the north of Portugal. The phytochemical properties of by-products were identified and characterized. Ohmic heating (OH) as a green extraction method was also applied to grape pomace due to their unknown effects on centesimal and phytochemical compositions. Both protein and carbohydrates were shown to be higher in grape bagasse than in stems. Additionally, red bagasse is richer in bioactive compounds (BC) than white bagasse. The sugar content was 21.91 and 11.01 g/100 g of DW in red and white grape bagasse, respectively. The amount of protein was 12.46 g/100 g of DW for red grape bagasse and 13.18 g/100 g of DW for white. Regarding the extraction methods, two fractions were obtained, a liquid fraction and solid (the remainder after the methodology application). OH presented a higher antioxidant capacity than a conventional (CONV) method. In addition, both extracts presented similar contents of anthocyanins, e.g., delphinidin-3-O-glucoside, petunidin-3-O-glucoside, and peonidin-3-O-glucoside. The solid fraction presented higher amounts of protein and phenols bound to fiber than CONV, which allows its use as a functional ingredient. In conclusion, OH can be an alternative extraction method compared with CONV methods, avoiding non-food grade solvents, thus contributing to circular economy implementation.

Coelho, Marta C. (ORCID:000000026542175X)↗
NASA NTRS1998

Isolation and characterization of Arabidopsis mutants defective in the induction of ethylene biosynthesis by cytokinin

Cytokinins elevate ethylene biosynthesis in etiolated Arabidopsis seedlings via a post-transcriptional modification of one isoform of the key biosynthetic enzyme ACC synthase. In order to begin to dissect the signaling events leading from cytokinin perception to this modification, we have isolated a series of mutants that lack the ethylene-mediated triple response in the presence of cytokinin due to their failure to increase ethylene biosynthesis. Analysis of genetic complementation and mapping revealed that these Cin mutants (cytokinin-insensitive) represent four distinct complementation groups, one of which, cin4, is allelic to the constitutive photomorphogenic mutant fus9/cop10. The Cin mutants have subtle effects on the morphology of adult plants. We further characterized the Cin mutants by analyzing ethylene biosynthesis in response to various other inducers and in adult tissues, as well as by assaying additional cytokinin responses. The cin3 mutant did not disrupt ethylene biosynthesis under any other conditions, nor did it disrupt any other cytokinin responses. Only cin2 disrupted ethylene biosynthesis in multiple circumstances. cin1 and cin2 made less anthocyanin in response to cytokinin. cin1 also displayed reduced shoot initiation in tissue culture in response to cytokinin, suggesting that it affects a cytokinin signaling element.

Non-NASA Center↗
NASA NTRS2015

Selection of Leafy Green Vegetable Varieties for a Pick-and-Eat Diet Supplement on ISS

Several varieties of leafy vegetables were evaluated with the goal of selecting those with the best growth, nutrition, and organoleptic acceptability for ISS. Candidate species were narrowed to commercially available cultivars with desirable growth attributes for space (e.g., short stature and rapid growth). Seeds were germinated in controlled environment chambers under conditions similar to what might be found in the Veggie plant growth chamber on ISS. Eight varieties of leafy greens were grown: 'Tyee' spinach, 'Flamingo' spinach, 'Outredgeous' Red Romaine lettuce, 'Waldmann's Dark Green' leaf lettuce, 'Bull's Blood' beet, 'Rhubarb' Swiss chard, 'Tokyo Bekana' Chinese cabbage, and Mizuna. Plants were harvested at maturity and biometric data on plant height, diameter, chlorophyll content, and fresh mass were obtained. Tissue was ground and extractions were performed to determine the tissue elemental content of Potassium (K), Magnesium (Mg), Calcium (Ca) and Iron (Fe). Following the biometric/elemental evaluation, four of the eight varieties were tested further for levels of anthocyanins, antioxidant (ORAC-fluorescein) capacity, lutein, zeaxanthin, and Vitamin K. For sensory evaluation, 'Outredgeous' lettuce, Swiss chard, Chinese cabbage, and Mizuna plants were grown, harvested when mature, packaged under refrigerated conditions, and sent to the JSC Space Food Systems Laboratory. Tasters evaluated overall acceptability, appearance, color intensity, bitterness, flavor, texture, crispness and tenderness. All varieties received acceptable scores with overall ratings greater than 6 on a 9-point hedonic scale. Chinese cabbage was the highest rated, followed by Mizuna, 'Outredgeous' lettuce, and Swiss chard. Based on our results, the selected varieties of Chinese cabbage, lettuce, Swiss chard and Mizuna seem suitable for a pick-and-eat scenario on ISS with a ranking based on all factors analyzed to help establish priority.

Nutrition↗
NASA NTRS2016

Pick and Eat Crop Testing: Dwarf Tomato and Pepper as Candidate Space Crops

Several dwarf tomato and pepper varieties were evaluated under International Space Station (ISS)-simulated growth conditions (22 degrees Centigrade, 50 percent relative humidity, 1500 parts per million CO2, and 300 micromoles per square meter per second of light for 16 hours per day) with the goal of selecting those with the best growth, nutrition, and organoleptic potential for use in a pick and eat salad crop system on ISS and future exploration flights. Testing included six cultivars of tomato (Red Robin, Scarlet Sweet 'N' Neat, Tiny Tim, Mohamed, Patio Princess, and Tumbler) and six cultivars of pepper (Red Skin, Fruit Basket, Cajun Belle, Chablis, Sweet Pickle, and Pompeii). Plants were grown to an age sufficient to produce fruit (up to 106 days for tomato and 109 days for pepper) using Turface (arcillite) potting media with 18-6-8 control-release fertilizer and supplemental nutrient solution beginning around 60-days-age. Tomato fruits were harvested when they showed full red color, beginning around 70-days age and then at weekly intervals thereafter, while peppers were grown until fruits showed color and were harvested twice (first test) and just once at the end of the second test, with the final harvests including colored and green fruit. Plant sizes, yields, and nutritional attributes were measured and used to down-select to three cultivars for each species. In particular, we were interested in cultivars that were short (dwarf) but still produced high yields. Nutritional data included elemental (Ca, Mg, Fe, and K) content, vitamin K, phenolics, lycopene (for tomato), anthocyanin, lutein, and zeaxanthin. The three down-selected cultivars for each species were grown again and the harvested fruit sent to NASA's Johnson Space Center for sensory evaluation, which included overall acceptability, appearance, color intensity, aroma, flavor and texture. The combined data were compared and given weighting factors to rank the cultivars as candidates for testing in space. Weightings gave maximum importance to plant size (smaller being good) and fruit yield (greater yields being good). For tomato, the ranking was 1) cultivar Mohamed and cultivar Red Robin (tied), and 3) cultivar Sweet N' Neat. For pepper, the ranking was 1) cultivar Pompeii, 2) cultivar Red Skin, and 3) cultivar Fruit Basket. These rankings are somewhat subjective but provide a starting point for conducting higher fidelity testing with these crops (e.g., testing with light emitting diode lighting similar to the Veggie plant unit on ISS), and ultimately conducting a flight experiment.

Food↗
NASA NTRS2016

Development and Provision of Functional Foods to Promote Health on Long-Duration Space Missions

During long-duration NASA space missions, such as proposed missions to Mars, astronauts may experience negative physiological effects such as bone loss. Functional foods such as high-lycopene, high-flavonoids and high-omega-3 products and fruits and vegetables may mitigate the negative effects of spaceflight on physiological factors including the bone health of crewmembers. Previous studies showed that current ISS provisions provide high-lycopene and high-omega-3 food items but the variety is limited, which could promote menu fatigue. Bioactive compounds can degrade like other chemical compounds and lose functionality. The native concentrations and stability of bioactive compounds have never been determined in spaceflight foods, and adequate information is not available for commercial products for the storage durations required for space exploration (5 years). The purpose of this task is to develop new spaceflight foods that are high in omega-3 fatty acids, lycopene, or flavonoids, identify commercial products with these bioactive compounds that meet spaceflight requirements, and define the stability of these nutrients in storage to enable purposeful functional food incorporation into the space food system. The impact of storage temperature on the stability of lutein, lycopene, beta-carotene, omega-3 fatty acids, phenolics, anthocyanins and sterols is being studied in 12 ISS menu items stored at three different temperatures (4, 21, 35 degree C) over 2 years. Additionally, nutrient and quality stability are being assessed on a larger food set stored at 21 degree C over 2 years that contains twelve newly developed foods, 10 commercial products repackaged to spaceflight requirements, and another 5 current ISS menu items expected to be good sources of omega-3 fatty acids, lycopene, or flavonoids. All items were shipped overnight to the Linus Pauling Institute at Oregon State University (Corvalis, OR) after processing and 1-year of storage and analyzed for bioactive compound concentrations. Sensory evaluation was conducted on the newly developed functional foods and commercial products with untrained panelists (n is greater than or equal to 25) using a 9-point Hedonic scale to test sensory attributes and overall acceptability after processing and 1-year of storage (21 degree C). Repeat nutritional and sensory analyses will be conducted in the same foods after the 2-year storage period is completed. The stability of bioactive compounds in the selected foods was dependent on storage temperature and food matrix. Omega-3 showed excellent stability in the analyzed products after 1-year of storage, regardless of the storage temperature; phenolic compounds also showed good stability. Lycopene was more stable in oil-based products rather than water-based products because of the protection that lipids offer to lycopene molecules. Also, lycopene was more stable in freeze-dried products than in high moisture foods. The 12 newly developed functional foods showed good overall acceptability in sensory attributes after processing (average score 7.2 out of 9.0) and maintained sensory quality through 1-year (21 degree C); the overall acceptability was on average 7.1 after storage. Similar behavior was observed for the 10 commercial products after 1 year. The developed products are good sources of omega-3 (both plant and marine), vegetables (7 vegetable-based products), and good sources of carotenoids, such as the Curry Pumpkin Soup and the Sweet and Savory Kale. Nine of the new products, such as Mango Salad, Pickled Beets, and Braised Red Cabbage, are rich in phenolic compounds. Stability of most of the studied nutrients seems to be adequate after 1-year of storage in most of the tested foods. However, storage temperature of the food must be considered during long-duration space missions to achieve stability of all nutrients. Likewise, more information is needed regarding nutrient retention after 2-years of storage to identify nutritional gaps that may be expected over the 5-year shelf life required for a Mars mission. New developed products will be filling a gap in the current space food system to minimize menu fatigue, provide specific nutrients to reduce the negative effects of long-duration space missions and maintain crew members' health. Information about bioactive compounds in developed products after 1-year and 2-year of storage will provide the knowledge base for further product development.

Bermudez-Aguirre, D.↗
NASA NTRS2016

Veggie: Space Vegetables for the International Space Station and Beyond

The Veggie vegetable production system was launched to the International Space Station (ISS) in 2014. Veggie was designed by ORBITEC to be a compact, low mass, low power vegetable production system for astronaut crews. Veggie consists of a light cap containing red, blue, and green LEDs, an extensible transparent bellows, and a baseplate with a root mat reservoir. Seeds are planted in plant pillows, small growing bags that interface with the reservoir. The Veggie technology validation test, VEG-01, was initiated with the first test crop of 'Outredgeous' red romaine lettuce. Prior to flight, lettuce seeds were sanitized and planted in a substrate of arcillite (baked ceramic) mixed with controlled release fertilizer. Upon initiation, astronauts open the packaged plant pillows, install them in the Veggie hardware, and prime the system with water. Operations include plant thinning, watering, and photography. Plants were grown on the ISS for 33 days, harvested, and returned frozen to Earth for analysis. Ground controls were conducted at Kennedy Space Center in controlled environment chambers reproducing ISS conditions of temperature, relative humidity, and CO2. Returned plant samples were analyzed for microbial food safety and chemistry including elements, antioxidants, anthocyanins and phenolics. In addition the entire plant microbiome was sequenced, and returned plant pillows were analyzed via x-ray tomography. Food safety analyses allowed us to gain approvals for future consumption of lettuce by the flight surgeons and the payload safety office. A second crop of lettuce was grown in 2015, and the crew consumed half the produce, with the remainder frozen for later analysis. This growth test was followed by testing of a new crop in Veggie, zinnias. Zinnias were grown to test a longer duration flowering crop in preparation for tests of tomatoes and other fruiting crops in the future. Zinnias were harvested in February. Samples from the second harvest of lettuce and the zinnia harvest are frozen on the ISS and will return with the next cargo return flight. Some challenges occurred in all tests, especially in the area of watering, with plants receiving insufficient or excess water leading to stressed growth. Zinnia plants were also challenged with fungal growth. Initial tests with Veggie have given us great insight into future crop production scenarios as we work to develop regular supplemental salad crop production on ISS and larger food production systems for our journey to Mars. Funding for this research comes from NASA's Space Biology Program.

Food Safety↗
NASA NTRS2017

The Stability of Bioactive Compounds in Spaceflight Foods

The status and stability of bioactive compounds in the processed and shelf-stable spaceflight food system have not previously been investigated though the presence of such compounds in aged space foods could have health significance for crews on long duration exploration missions. Over forty foods - either existing International Space Station (ISS) food provisioning items, newly developed foods for spaceflight, or commercially-available ready-to-eat foods - that were predicted to have a relatively high concentrations of one or more bioactive compounds (lycopene, lutein, omega-3 fatty acids, phenolics, sterols, and/or flavonoids) were selected for the study. Food samples were sent overnight to the Food Composition Laboratory of the Linus Pauling Institute at Oregon State University (Corvallis, OR) for bioactive compound analysis. Three packages of each product were blended together for the analysis to reduce package-to-package variability. All ISS food items and commercial foods were analyzed initially and after 12 and 24 months of 21degC storage. Food development occurred in a staggered fashion, so data collection for the newly developed foods continues. Lastly, sensory evaluation and additional temperature storage data (4degC, 35degC) for select foods were collected to establish additional stability parameters. Efficacious concentrations of lycopene, lutein, and omega-3 fatty acids were measured in limited spaceflight foods; two grams of sterols a day may be difficult to achieve with the current space diet. Total polyphenol delivery appears stable and adequate, but individual phenolic compounds vary in stability and were not specifically evaluated in this study. The data suggests that some bioactive compounds, like lycopene and lutein, degrade and then plateau at some equilibrium concentration. The anthocyanin stability appears to be related to storage temperature and food matrix, and lutein stability in leafy vegetables may be impacted by storage temperature. Because of the limited number of foods with high concentrations of the bioactive compounds, additional menu variety, formulation optimization, and reduced temperature storage will be required to ensure delivery of several bioactive compounds in the space food system. Validation of stability to five years will enable provisioning of these functional foods within the space food system for a mission to Mars.

Cooper, M. R.↗
NASA NTRS2017

Effects of White Leds on Growth and Phytonutrients of Outredgeous Romaine Lettuce When Supplemented with Various Monochromatic Wavelengths

Growing plants in space will be an essential part of sustaining astronauts during long-range missions. To drive photosynthesis, light-emitting diodes (LEDs) are becoming superior because of their efficiency, longevity, small size, safety, and wavelength versatility. Isolating the effects of certain wavelengths on plant growth when combined with white light is attracting attention. To optimize crop production/quality in space, this study has aimed to configure novel light recipes for the Advanced Plant Habitat currently aboard the International Space Station (ISS). By using white light as a background to maintain normal growth, the addition of monochromatic wavelengths provides a clearer understanding of how each part of the visible spectrum affects plant growth. By growing Outredgeous lettuce under six treatments of White (W) LEDs, W + blue (B), W+ green (G), W + red (R), W + far red (FR), and RGB + FR LEDs with ratios similar to natural sunlight, this investigation has assessed differences in biomass, morphology, chlorophyll, and the synthesis of key phytonutrients. The potential for Outredgeous to produce anthocyanin, lutein, potassium, magnesium, and iron is paramount to maintaining astronaut health. The crop responses to each treatment have been evaluated and the optimal LED combination for both plant yield and nutrient content will be presented.

Food Production↗
NASA NTRS2019

Dwarf Tomato and Pepper Cultivars for Space Crops

Several dwarf tomato and pepper varieties were evaluated under ISS-simulated growth conditions (22ºC, 50% RH, 1500 ppm CO2, and 300 μmol m(exp -2) s(exp -1) of light for 16 h per day) with the goal of selecting those with the best growth, nutrition, and organoleptic potential for use in a pick and eat salad crop system on ISS and future exploration flights. Testing included six cultivars of tomato (Red Robin, Scarlet Sweet ’N’ Neat, Tiny Tim, Mohamed, Patio Princess, and Tumbler) and six cultivars of pepper (Red Skin, Fruit Basket, Cajun Belle, Chablis, Sweet Pickle, and Pompeii). Plants were grown to an age sufficient to produce fruit (70 to 106 days for tomato and 109 days for pepper). Tomato fruits were harvested when they showed full red color, beginning ca. 70-days age and then at weekly intervals thereafter, while peppers were grown until numerous fruits showed color and all fruits (green and colored) were harvested once at the end of the test. Plant sizes, yields, and nutritional attributes were measured and used to down-select to three cultivars for each species. In particular, we were interested in cultivars that were short (dwarf) but still produced high yields. Nutritional data included elemental (Ca, Mg, Fe, and K) composition, vitamin K, phenolics, lycopene, anthocyanin, lutein, and zeaxanthin. The three down-selected cultivars for each species were evaluated for sensory attributes, including overall acceptability, appearance, color intensity aroma, flavor and texture. The combined data were compared and given weighting factors to rank the cultivars as potential candidates for testing in space. For tomato, the ranking was 1) cv. Mohamed, 2) cv. Red Robin, and 3) cv. Sweet N’ Neat. For pepper, the ranking was 1) cv. Pompeii, 2) cv. Red Skin, and 3) cv. Fruit Basket. These rankings are somewhat subjective but provide a good starting point for conducting higher fidelity testing with these crops (e.g., testing with LED lighting similar to the Veggie plant unit), and ultimately conducting flight experiments.

Spencer, LaShelle E.↗
DOE OSTI2024

Structural and Interactional Analysis of the Flavonoid Pathway Proteins: Chalcone Synthase, Chalcone Isomerase and Chalcone Isomerase-like Protein

Chalcone synthase (CHS) and chalcone isomerase (CHI) catalyze the first two committed steps of the flavonoid pathway that plays a pivotal role in the growth and reproduction of land plants, including UV protection, pigmentation, symbiotic nitrogen fixation, and pathogen resistance. Based on the obtained X-ray crystal structures of CHS, CHI, and chalcone isomerase-like protein (CHIL) from the same monocotyledon, Panicum virgatum, along with the results of the steady-state kinetics, spectroscopic/thermodynamic analyses, intermolecular interactions, and their effect on each catalytic step are proposed. In addition, PvCHI’s unique activity for both naringenin chalcone and isoliquiritigenin was analyzed, and the observed hierarchical activity for those type-I and -II substrates was explained with the intrinsic characteristics of the enzyme and two substrates. The structure of PvCHS complexed with naringenin supports uncompetitive inhibition. PvCHS displays intrinsic catalytic promiscuity, evident from the formation of p-coumaroyltriacetic acid lactone (CTAL) in addition to naringenin chalcone. In the presence of PvCHIL, conversion of p-coumaroyl-CoA to naringenin through PvCHS and PvCHI displayed ~400-fold increased Vmax with reduced formation of CTAL by 70%. Supporting this model, molecular docking, ITC (Isothermal Titration Calorimetry), and FRET (Fluorescence Resonance Energy Transfer) indicated that both PvCHI and PvCHIL interact with PvCHS in a non-competitive manner, indicating the plausible allosteric effect of naringenin on CHS. Significantly, the presence of naringenin increased the affinity between PvCHS and PvCHIL, whereas naringenin chalcone decreased the affinity, indicating a plausible feedback mechanism to minimize spontaneous incorrect stereoisomers. These are the first findings from a three-body system from the same species, indicating the importance of the macromolecular assembly of CHS-CHI-CHIL in determining the amount and type of flavonoids produced in plant cells.

59 BASIC BIOLOGICAL SCIENCES↗