Spectral quality may be used to alter plant disease development in CELSS.

Plants were grown under light emitting diode (LED) arrays with different spectral qualities to determine the effects of light on the development of tomato mosaic virus (ToMV) in peppers and powdery mildew on cucumbers. One LED array supplied 100% of the photosynthetic photon flux (PPF) at 660 nm, a second array supplied 90% of the PPF at 660 nm and 10% at 735 nm, and a third array supplied 98% of the PPF at 660 nm with 2% in the blue region (380-500 nm) supplied by blue fluorescent lamps. Control plants were grown under metal halide (MH) lamps. Pepper plants inoculated with ToMV and grown under 660 and 660/735 LED arrays showed marked increases in both the rate and the severity of symptoms as compared to inoculated plants grown under the MH lamp or 660/blue array. Pepper plants grown under the 660/blue array did not develop symptoms as rapidly as inoculated plants grown under the 660 or 660/735 arrays, but they did develop symptoms faster than inoculated plants grown under the MH lamp. The numbers of colonies of powdery mildew per leaf and the size of each colony were greatest on inoculated cucumber plants grown under the MH lamp.     

Effects of air velocity on photosynthesis of plant canopies under elevated CO2 levels in a plant culture system.

To obtain basic data for adequate air circulation for promoting plant growth in closed plant production modules in bioregenerative life support systems in space, effects of air velocities ranging from 0.1 to 0.8 m s-1 on photosynthesis in tomato seedlings canopies were investigated under atmospheric CO2 concentrations of 0.4 and 0.8 mmol mol-1. The canopy of tomato seedlings on a plug tray (0.4 x 0.4 m2) was set in a wind-tunnel-type chamber (0.6 x 0.4 x 0.3 m3) installed in a semi-closed-type assimilation chamber (0.9 x 0.5 x 0.4 m3). The net photosynthetic rate in the plant canopy was determined with the differences in CO2 concentrations between the inlet and outlet of the assimilation chamber multiplied by the volumetric air exchange rate of the chamber. Photosynthetic photon flux (PPF) on the plant canopy was kept at 0.25 mmol m-2 s-1, air temperature at 23 degrees C and relative humidity at 55%. The leaf area indices (LAIs) of the plant canopies were 0.6-2.5 and plant heights were 0.05-0.2 m. The net photosynthetic rate of the plant canopy increased with increasing air velocities inside plant canopies and saturated at 0.2 m s-1. The net photosynthetic rate at the air velocity of 0.4 m s-1 was 1.3 times that at 0.1 m s-1 under CO2 concentrations of 0.4 and 0.8 mmol mol-1. The net photosynthetic rate under CO2 concentrations of 0.8 mmol mol-1 was 1.2 times that under 0.4 mmol mol-1 at the air velocity ranging from 0.1 to 0.8 m s-1. The results confirmed the importance of controlling air movement for enhancing the canopy photosynthesis under an elevated CO2 level as well as under a normal CO2 level in the closed plant production modules.     

CO2 crop growth enhancement and toxicity in wheat and rice.

The effects of elevated CO2 on plant growth are reviewed and the implications for crop yields in regenerative systems are discussed. There is considerable theoretical and experimental evidence indicating that the beneficial effects of CO2 are saturated at about 0.12% CO2 in air. However, CO2 can easily rise above 1% of the total gas in a closed system, and we have thus studied continuous exposure to CO2 levels as high as 2%. Elevating CO2 from 340 to 1200 micromoles mol-1 can increase the seed yield of wheat and rice by 30 to 40%; unfortunately, further CO2 elevation to 2500 micromoles mol-1 (0.25%) has consistently reduced yield by 25% compared to plants grown at 1200 micromoles mol-1; fortunately, there was only an additional 10% decrease in yield as the CO2 level was further elevated to 2% (20,000 micromoles mol-1). Yield increases in both rice and wheat were primarily the result of increased number of heads per m2, with minor effects on seed number per head and seed size. Yield increases were greatest in the highest photosynthetic photon flux. We used photosynthetic gas exchange to analyze CO2 effects on radiation interception, canopy quantum yield, and canopy carbon use efficiency. We were surprised to find that radiation interception during early growth was not improved by elevated CO2. As expected, CO2 increased quantum yield, but there was also a small increase in carbon use efficiency. Super-optimal CO2 levels did not reduce vegetative growth, but decreased seed set and thus yield. The reduced seed set is not visually apparent until final yield is measured. The physiological mechanism underlying CO2 toxicity is not yet known, but elevated CO2 levels (0.1 to 1% CO2) increase ethylene synthesis in some plants and ethylene is a potent inhibitor of seed set in wheat.     

Effect of CO2 levels on nutrient content of lettuce and radish.

Atmospheric carbon-dioxide enrichment is known to affect the yield of lettuce and radish grown in controlled environments, but little is known about CO2 enrichment effects on the chemical composition of lettuce and radish. These crops are useful model systems for a Controlled Ecological Life-Support System (CELSS), largely because of their relatively short production cycles. Lettuce (Lactuca sativa L.) cultivar 'Waldmann's Green' and radish (Raphanus sativus L.) cultivar 'Giant White Globe' were grown both in the field and in controlled environments, where hydroponic nutrient solution, light, and temperature were regulated, and where CO2 levels were controlled at 400, 1000, 5000, or 10,000 ppm. Plants were harvested at maturity, dried, and analyzed for proximate composition (protein, fat, ash, and carbohydrate), total nitrogen (N), nitrate N, free sugars, starch, total dietary fiber, and minerals. Total N, protein N, nonprotein N (NPN), and nitrate N generally increased for radish roots and lettuce leaves when grown under growth chamber conditions compared to field conditions. The nitrate-N level of lettuce leaves, as a percentage of total NPN, decreased with increasing levels of CO2 enrichment. The ash content of radish roots and of radish and lettuce leaves decreased with increasing levels of CO2 enrichment. The levels of certain minerals differed between field- and chamber-grown materials, including changes in the calcium (Ca) and phosphorus (P) contents of radish and lettuce leaves, resulting in reduced Ca/P ratio for chamber-grown materials. The free-sugar contents were similar between the field and chamber-grown lettuce leaves, but total dietary fiber content was much higher in the field-grown plant material. The starch content of growth-chamber lettuce increased with CO2 level.     

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.     

Changes in spectral reflectance of wheat leaves in response to specific macronutrient deficiency.

In wheat (Triticum aestivum L.) plants, deficiency of an essential element may drastically affect growth, appearance, and most importantly yield. Wheat, the focus of this study, is one of the crops studied in the CELSS program. Information about nutrient deficiencies in crops grown in controlled environment is essential to optimize food productivity. The main objective of this study was to determine whether deficiency of Nitrogen (N), Phosphorus (P), Potassium (K), Calcium (Ca) and Magnesium (M) alters spectral reflectance properties of wheat leaves. Plants were grown in the greenhouse and growth chamber, in a modified Hoagland's nutrient solution. Spectral reflectance of fully expanded wheat leaves from 280 to 1100 nm, nutrient concentrations (N, P, K, and Ca) and chlorophyll (Chl) were determined when deficiency symptoms were first evident (approximately 6-7 weeks). Chlorophyll content and fresh and dry weight, were used to assess the severity of the nutrient stress. All nutrient deficiencies affected chlorophyll content and generally increased reflectance in the visible (VIS) 400-700 nm and infrared (IR) 700-1100 nm ranges. Magnesium and nitrogen deficiencies had the most pronounced effect on chlorophyll concentration height, and reflectance. All macronutrient deficiencies tested reduced chlorophyll concentration, increase reflectance in the visible range and caused a shift in the position of the red edge (the point of maximum slope on the reflectance spectrum of vegetation between red and near-infrared wavelengths) toward shorter or longer wavelengths; depending upon the element. In the greenhouse, N and Mg induced the greatest increase in reflectance of 33% and 25% in the VI range and 86% and 53% in the IR range, respectively. However, in the growth chamber, an increase of 97% and 25% occurred in the VI range, and 20% and 33% in the IR range, respectively. In the IR range in the growth chamber, P, K, and Ca deficiency caused a reduction in reflectance (412-770 nm). This research indicates that mineral deficiencies and reflectance are not specific to one environment and could have important implications for the design of CELSS in space, and perhaps the future of terrestrial agriculture.     

CO2 enrichment influences yields of 'Florunner,' 'Georgia Red' and 'New Mexico' peanut cultivars.

Three peanut cultivars, 'Florunner,' 'Georgia Red,' and 'New Mexico,' were grown in reach-in chambers to determine response to CO2 enrichment. CO2 treatments were ambient (400 micromol mol-1) and 700 micromol mol-1. Growth chamber conditions included 700 micromol m-2 s-1 photosynthetic photon flux (PPF), 28/22C, 7O% RH, and 12/12 h photoperiod. Growth media consisted of a 1:1 mixture (v/v) of vermiculite and sterilized sand. Six 10 L pots of each cultivar were fertilized three times per week with 250 mL of nutrient solution containing additional Ca (10 mM) and NO3 (25 mM) and watered well. Beginning 21 days after planting (DAP) and every three weeks thereafter up to 84 days, the second leaf from the growing axis (main stem) was detached to determine CO2 effect on leaf area, specific leaf area (SLA) and dry weight. Plants were harvested 97 DAP, at which time total leaf area, leaf number, plant and root weights and pod production data were taken. Numbers of pods per plant, pod fresh and dry weights, fibrous root and plant dry weights were higher for all cultivars grown at 700 micromol mol-1 than at ambient CO2. Also, leaf area for all cultivars was larger with CO2 enrichment than at ambient. SLA tended to decline with time regardless of CO2 treatment. Percentage of total sound mature kernels (%TSMK) was similar for both treatments. Plants grown at 700 micromol mol-1 CO2 had slightly more immature pods and seeds at final harvest.     

Productivity and food value of Amaranthus cruentus under non-lethal salt stress.

Stress effects from the accumulation of metal salts may pose a problem for plants in closed biological systems such as spacecraft. This work examined the effects of salinity on growth, photosynthesis and carbon allocation in the crop plant, Amaranthus. Plants were germinated and grown in modified Hoagland's solution with NaCl concentrations of 0 to 1.0%. Plants received salt treatments at various times in development to assess effects on particular life history phases. For Amaranthus cruentus, germination, vegetative growth, flowering, seed development and yield were normal at salinities from 0 to 0.2%. Inhibition of these phases increased from 0.2 to 0.4% salinity and was total above 0.5%. 1.0% salinity was lethal to all developmental phases. Onset of growth phases were not affected by salinity. Plants could not be adapted by gradually increasing salinity over days or weeks. Water uptake increased, while photosynthetic CO2 uptake decreased with increasing salinity on a dry weight basis during vegetative growth. Respiration was not affected by salinity. After flowering, respiration and photosynthesis decreased markedly, such that 1.0% NaCl inhibited photosynthesis completely. Protein levels were unchanged with increasing salinity. Leaf starch levels were lower at salinities of 0.5% and above, while stem starch levels were not affected by these salinities. The evidence supports salt inhibition arising from changes in primary biochemical processes rather than from effects on water relations. While not addressing the toxic effects of specific ions, it suggests that moderate salinity per se need not be a problem in space systems.     

Regulating plant/insect interactions using CO2 enrichment in model ecosystems.

The greenhouse environment is a challenging artificial ecosystem in which it is possible to study selected plant/insect interaction in a controlled environment. Due to a combination of "direct" and "indirect" effects of CO2 enrichment on plant photosynthesis and plant development, canopy productivity is generally increased. In this paper, we discuss the effects of daytime and nighttime CO2 enrichment protocols on gas exchange of pepper plants (Capsicum annuum L, cv Cubico) grown in controlled environments. In addition, we present the effects of thrips, a common Insect pest, on the photosynthetic and respiratory activity of these plant canopies. Carbon dioxide has diverse effects on the physiology and mortality of insects. However, our data indicate that thrips and whiteflies, at least, are not killed "directly" by CO2 levels used to enhance photosynthesis and plant growth. Together the data suggest that the insect population is affected "indirectly" by CO2 and that the primary effect of CO2 is via its effects on plant metabolism.     

Carbon dioxide exchange of lettuce plants under hypobaric conditions.

Growth of plants in a Controlled Ecological Life Support System (CELSS) may involve the use of hypobaric pressures enabling lower mass requirements for atmospheres and possible enhancement of crop productivity. A controlled environment plant growth chamber with hypobaric capability designed and built at Ames Research Center was used to determine if reduced pressures influence the rates of photosynthesis (Ps) and dark respiration (DR) of hydroponically grown lettuce plants. The chamber, referred to as a plant volatiles chamber (PVC), has a growing area of about 0.2 m2, a total gas volume of about 0.7 m3, and a leak rate at 50 kPa of <0.1%/day. When the pressure in the chamber was reduced from ambient to 51 kPa, the rate of net Ps increased by 25% and the rate of DR decreased by 40%. The rate of Ps increased linearly with decreasing pressure. There was a greater effect of reduced pressure at 41 Pa CO2 than at 81 Pa CO2. This is consistent with reports showing greater inhibition of photorespiration (Pr) in reduced O2 at low CO2 concentrations. When the partial pressure of O2 was held constant but the total pressure was varied between 51 and 101 kPa, the rate of CO2 uptake was nearly constant, suggesting that low pressure enhancement of Ps may be mainly attributable to lowered partial pressure of O2 and the accompanying reduction in Pr. The effects of lowered partial pressure of O2 on Ps and DR could result in substantial increases in the rates of biomass production, enabling rapid throughput of crops or allowing flexibility in the use of mass and energy resources for a CELSS.     

Carbon dioxide exchange of lettuce plants under hypobaric conditions.

Growth of plants in a Controlled Ecological Life Support System (CELSS) may involve the use of hypobaric pressures enabling lower mass requirements for atmospheres and possible enhancement of crop productivity. A controlled environment plant growth chamber with hypobaric capability designed and built at Ames Research Center was used to determine if reduced pressures influence the rates of photosynthesis (Ps) and dark respiration (DR) of hydroponically grown lettuce plants. The chamber, referred to as a plant volatiles chamber (PVC), has a growing area of about 0.2 m2, a total gas volume of about 0.7 m3, and a leak rate at 50 kPa of <0.1%/day. When the pressure in the chamber was reduced from ambient to 51 kPa, the rate of net Ps increased by 25% and the rate of DR decreased by 40%. The rate of Ps increased linearly with decreasing pressure. There was a greater effect of reduced pressure at 41 Pa CO2 than at 81 Pa CO2. This is consistent with reports showing greater inhibition of photorespiration (Pr) in reduced O2 at low CO2 concentrations. When the partial pressure of O2 was held constant but the total pressure was varied between 51 and 101 kPa, the rate of CO2 uptake was nearly constant, suggesting that low pressure enhancement of Ps may be mainly attributable to lowered partial pressure of O2 and the accompanying reduction in Pr. The effects of lowered partial pressure of O2 on Ps and DR could result in substantial increases in the rates of biomass production, enabling rapid throughput of crops or allowing flexibility in the use of mass and energy resources for a CELSS.     

Growth of wheat under one tenth of the atmospheric pressure.

Wheat plants were grown in twin closed growth chambers under normal and reduced atmospheric pressures. For the first 22 days from sowing, the reduced pressure was maintained at 200 hPa, and at 100 hPa for the remaining 27 days until harvest. These pressures were obtained by evacuation of the chamber and adding oxygen (170 and 79 hPa respectively) and carbon dioxide (0.65 and 1.0 hPa respectively; about 2 and 3 times above the control). Eighty-seven per cent of the final dry mass was produce under 100 hPa treatment. Growth and development of wheat are not negatively affected by low pressure treatment. Compared to the control, final dry mass increased by 76%, leaf number by 133%, and ear number by 35%, probably due to elevation of CO2. Shortening of shoot parts and increases in chlorophyll and proteins content are not in accordance with a predicted CO2 effect and could be attributed to the N2 removal and the subsequent alteration in gas diffusion rate.     

Carbon balance and productivity of Lemna gibba, a candidate plant for CELSS.

The photosynthesis and productivity of Lemna gibba were studied with a view to its use in Controlled Ecological Life Support Systems (CELSS). Photosynthesis of L. gibba floating on the nutrient solution could be driven by light coming from either above or below. Light from below was about 75% as effective as from above when the stand was sparse, but much less so with dense stands. High rates of photosynthesis (ca. 800 nanomoles CO2 g dry weight (DW)-1 s-1) were measured at 750 micromoles m-2 s-1 PPF and 1500 micromoles mol-1 CO2. This was attained at densities up to 660 g fresh weight (FW) m-2 with young cultures. After a few days growth under these conditions, and at higher densities, the rate of photosynthesis dropped to less than 25% of the initial value. This drop was only partly alleviated by thinning the stand or by introducing a short dark period at high temperature (26 degrees C). Despite the drop in the rate of photosynthesis, maximum yields were obtained in batch cultures grown under continuous light, constant temperature and high [CO2]. Plant protein content was less than reported for field grown Lemna. When the plants were harvested daily, maintaining a stand density of 600 g FW m-2, yields of 18 g DW m-2 d-1 were obtained. The total dry weight of L. gibba included 40% soluble material (sugars and amino acids), 15% protein, 5% starch, 5% ash and 35% cellulose and other polymers. We conclude that a CELSS system could be designed around stacked, alternate layers of transparent Lemna trays and lamps. This would allow for 7 tiers per meter height. Based on present data from single layers, the yield of such a system is calculated to be 135 g DW m-3 d-1 of a 100% edible, protein-rich food.     

Modeling gas exchange in a closed plant growth chamber.

Fluid transport models for fluxes of water vapor and CO2 have been developed for one crop of wheat and three crops of soybean grown in a closed plant growth chamber. Correspondence among these fluxes is discussed. Maximum fluxes of gases are provided for engineering design requirements of fluid recycling equipment in growth chambers. Furthermore, to investigate the feasibility of generalized crop models, dimensionless representations of water vapor fluxes are presented. The feasibility of such generalized models and the need for additional data are discussed.     

Plant responses to reduced air pressure: advanced techniques and results.

Knowledge on air pressure impacts on plant processes and growth is essential for understanding responses to altitude and for comprehending the way of action of aerial gasses in general, and is of potential importance for life support systems in space. Our research on reduced air pressure was extended by help of a new set-up comprising two constantly ventilated chambers (283 L each), allowing pressure gradients of +/-100 kPa. They provide favourable general growth conditions while maintaining all those factors constant or at desired levels which modify the action of air pressure, e.g., water vapour pressure deficit and air mass flow over the plants. Besides plant growth parameters, transpiration and CO2 gas exchange are determined continuously. Results are presented on young tomato plants grown hydroponically, which had been treated with various combinations of air pressure (400-700-1000 hPa), CO2 concentration and wind intensity for seven days. At the lowest pressure transpiration was enhanced considerably, and the plants became sturdier. On the other hand growth was retarded to a certain extent, attributable to secondary air pressure effects. Therefore, even greater limitations of plant productivity are expected after more extended periods of low pressure treatment.     

Developing a vitamin greenhouse for the life support system of the International Space Station and for future interplanetary missions.

In order to evaluate the effects of gravity on growing plants, we conducted ground based long-term experiments with dwarf wheat, cultivar Apogee and Chinese cabbage, cultivar Khibinskaya. The test crops had been grown in overhead position with HPS lamp below root module so gravity and light intensity gradients had been in opposite direction. Plants of the control crop grew in normal position under the same lamp. Both crops were grown on porous metallic membranes with stable -1 kPa matric potential on their surface. Results from these and other studies allowed us to examine the differences in growth and development of the plants as well as the root systems in relation to the value of the gravity force influence. Dry weight of the roots from test group was decreased in 2.5 times for wheat and in 6 times - at the Chinese cabbage, but shoot dry biomass was practically same for both test and control versions. A harvest index of the test plants increased substantially. The data shows, that development of the plants was essentially changed in microgravity. The experiments in the space greenhouse Svet aboard the Mir space station proved that it is possible to compensate the effects of weightlessness on higher plants by manipulating gradients of environmental parameters (i.e. photon flux, matric potential in the root zone, etc.). However, the average productivity of Svet concerning salad crops even in ground studies did not provide more than 14 g fresh biomass per day. This does not provide a sufficient level of supplemental nutrients to the crew of the ISS. A cylindrical design of a space plant growth chamber (SPGC) allows for maximal productivity in presence of very tight energy and volume limitations onboard the ISS and provides a number of operational advantages. Productivity from this type of SPGF with a 0.5 kW energy utilization when salad growing would provide approximately 100 g of edible biomass per day, which would almost satisfy requirements for a crew of two in vitamin C and carotene and partly vitamin B group as well as rough fiber.     

Growth of soybean and potato at high CO2 partial pressures.

Soybean and potato plants were grown in controlled environments at carbon dioxide (CO2) partial pressures ranging from 0.05 to 1.00 kPa. The highest yields of edible biomass occurred at 0.10 kPa for both species, with higher CO2 levels being supraoptimal, but not injurious to the plants. Stomatal conductance rates of upper canopy leaves were lowest at 0.10 kPa CO2, while conductance rates at 0.50 and 1.00 kPa were significantly greater than 0.10 kPa. Total water use by the plants was greatest at the highest CO2 pressures (i.e. 0.50 and 1.00 kPa); consequently, water use efficiencies (biomass produced/water used) were low at the highest CO2 pressures. Based on previous CO2 studies in the literature, the increased conductance and water use at the highest CO2 pressures were surprising and pose interesting challenges for managing plants in a CELSS, where CO2 pressures may exceed optimal levels.     

Instrumentation for plant health and growth in space.

The present-day plant growth facilities ("greenhouses") for space should be equipped with monitors and controllers of ambient parameters within the chamber because spacecraft environmental variations can be unfavorable to plants. Moreover, little is known about the effects of spaceflight on the greenhouse and rooting media. Lack of information about spaceflight effects on plants necessitates supplying space greenhouses with automatic, non-invasive monitors of, e.g., gas exchange rate, water and nutrient ion uptake, plant mass, temperature and water content of leaves. However, introduction of an environmental or plant sensor into the monitoring system may be reasonable only if it is justified by quantitative evaluation of the influence of a measured parameter on productivity, efficacy of illumination, or some other index of greenhouse efficiency. The multivariate adaptive optimization in terrestrial phytotrons appears to be one of the best methods to assess environmental impacts on crops. Two modifications of greenhouses with the three-dimensional adaptive optimization of crop photosynthetic characteristics include: (1) irradiation, air temperature and carbon dioxide using a modified simplex algorithm; and (2) using irradiation, air temperature, and humidity with sensitivity algorithms with varying frequency of test exposures that have been experimentally developed. As a result, during some stages of plant ontogensis, the photosynthetic productivity of wheat, tomatoes, and Chinese cabbage in these systems was found to increase by a factor of 2-3.     

Incineration of biomass and utilization of product gas as a CO2 source for crop production in closed systems: gas quality and phytotoxicity.

This study addressed the recycle of carbon from inedible biomass to CO2 for utilization in crop production. Earlier work identified incineration as an attractive approach to resource recovery from solid wastes because the products are well segregated. Given the effective separation of carbon into the gaseous product stream from the incinerator in the form of CO2 we captured the gaseous stream produced during incineration of wheat inedible biomass and utilized it as the CO2 source for crop production. Injection rate was based on maintenance of CO2 concentration in the growing environment. The crop grown in the closed system was lettuce. Carbon was primarily in the form of CO2 in the incinerator product gas with less than 8% of carbon compounds appearing as CO. Nitrogen oxides and organic compounds such as toluene, xylene, and benzene were present in the product gas at lower concentrations (< 4 micromol mol-1); sulfur containing compounds were below the detection limits. Direct utilization of the gaseous product of the incinerator as the CO2 source was toxic to lettuce grown in a closed chamber. Net photosynthetic rates of the crop was suppressed more than 50% and visual injury symptoms were visible within 3 days of the introduction of the incinerator gas. Even the removal of the incinerator gas alter two days of crop exposure and replacement with pure CO2 did not eliminate the toxic effects. Both organic and inorganic components of the incinerator gas are candidates for the toxin.