Effects of ethylenediurea and ozone on the antioxidative systems in beans (Phaseolus vulgaris L.)

To study the biochemical mechanism of EDU protection against ozone injury, peroxidase, ascorbate-dependent peroxidase, and catalase activities, and the contents of ascorbic acid, dehydroascorbic acid, malondialdehyde and soluble protein were measured in Phaseolus vulgaris L. cv. Lit exposed to ozone and ethylenediurea (EDU) in open-top chambers. Plants not treated with EDU showed foliar bronzing due to ozone, while EDU-treated plants were not affected. EDU application modified the reaction of biochemical parameters to ozone. Soluble protein content was elevated by EDU. Peroxidase activity increased with ozone exposure in untreated plants only, while ascorbate-dependent peroxidase activity was lower in EDU treated plants. Catalase activity decreased in EDU-untreated plants. The ratio of ascorbic acid to dehydroascorbic acid was significantly increased in EDU treated plants. These results suggest that EDU might induce ascorbic acid synthesis and therefore provide the plant with a very potent antioxidant. Or the content of hydrogen peroxide was reduced due to other unknown processes and caused a delay in foliar senescence, regardless of whether these processes were ozone-induced or due to natural aging processes.     

Inheritance of ambient ozone insensitivity in common bean (Phaseolus vulgaris L.)

The inheritance of ozone (O(3)) insensitivity in common bean (Phaseolus vulgaris L.) was evaluated using F(2) and F(3) populations under ambient conditions. This study was conducted over two growing seasons (1987, 1988) at Virginia State University, Randolph Research Farm, Petersburg, Virginia. Two populations were obtained by crossing insensitive plant introductions with sensitive commercial cultivars. Ratings on the scale of 1 to 5 (1 = 0 to 20% leaf injury, 2 = 21 to 40%, 3 = 41 to 60%, 4 = 61 to 80%, and 5 > 80%) were made on 160 F(2), F(3) progenies, and parental lines. Population mean injury ratings were recorded and estimates of genotypic, environmental, and phenotypic variances were computed. Estimates of heritability in the broadsense and of genetic advance were calculated for each population using F(2) and family component variance methods. Population means of the F(2) and F(3) progenies were not significantly different from their mid-parent values, suggesting that genetic variance was primarily additive. Broad-sense heritability estimates using F(2) variance method ranged from 51.4 to 70.5% and using family component variance method ranged from 62.1 to 75.6%. In this study, the computed genetic advance values closely parallel those of heritability estimated values. The high heritable nature of insensitivity would indicate that effective levels of insensitivity could be transferred to agronomically superior cultivars in a relatively short time.     

Cutleaf coneflower (Rudbeckia laciniata L.) response to ozone and ethylenediurea (EDU)

Cutleaf coneflower (Rudbeckia laciniata L.) seedlings were placed into open-top chambers in May, 2004 and fumigated for 12 wks. Nine chambers were fumigated with either carbon-filtered air (CF), non-filtered air (NF) or twice-ambient (2×) ozone (O₃). Ethylenediurea (EDU) was applied as a foliar spray weekly at 0 (control), 200, 400 or 600 ppm. Foliar injury occurred at ambient (30%) and elevated O₃ (100%). Elevated O₃ resulted in significant decreases in biomass and nutritive quality. Ethylenediurea reduced percent of leaves injured, but decreased root and total biomass. Foliar concentrations of cell-wall constituents were not affected by EDU alone; however, EDU × O₃ interactions were observed for total cell-wall constituents and lignocellulose fraction. Our results demonstrated that O₃ altered the physiology and productivity of cutleaf coneflower, and although reducing visible injury EDU may be phytotoxic at higher concentrations.     

A flux-based assessment of the effects of ozone on foliar injury, photosynthesis, and yield of bean (Phaseolus vulgaris L. cv. Borlotto Nano Lingua di Fuoco) in open-top chambers

Stomatal ozone uptake, determined with the Jarvis' approach, was related to photosynthetic efficiency assessed by chlorophyll fluorescence and reflectance measurements in open-top chamber experiments on Phaseolus vulgaris. The effects of O₃ exposure were also evaluated in terms of visible and microscopical leaf injury and plant productivity. Results showed that microscopical leaf symptoms, assessed as cell death and H₂O₂ accumulation, preceded by 3-4 days the appearance of visible symptoms. An effective dose of ozone stomatal flux for visible leaf damages was found around 1.33 mmol O₃ m⁻². Significant linear dose-response relationships were obtained between accumulated fluxes and optical indices (PRI, NDI, ΔF/F_m^′). The negative effects on photosynthesis reduced plant productivity, affecting the number of pods and seeds, but not seed weight. These results, besides contributing to the development of a flux-based ozone risk assessment for crops in Europe, highlight the potentiality of reflectance measurements for the early detection of ozone stress.     

Effects of nitrogen dioxide on growth and yield of black turtle bean (Phaseolus vulgaris L.) cv. 'Domino'

Twenty-six-day-old black turtle bean cv. 'Domino' plants were exposed to nitrogen dioxide (0.0, 0.025, 0.05 and 0.10 microl liter(-1)), 7 h per day for 5 days per week for 3 weeks, under controlled environment. Data were collected on net photosynthesis rate (PN), stomatal resistance (SR), and dark respiration rate (DR), immediately after exposure, 24 h after the termination of exposure and at maturity (when the leaves had just started turning yellow), using a LICOR 6000 Portable Photosynthesis System. Chlorophyll-a (Ch-a), chlorophyll-b (Ch-b), total chlorophyll (tot-Ch) and leaf nitrogen were measured immediately after exposure and at maturity. Growth characteristics-relative growth rate (RGR), net assimilation rate (NAR), leaf area ratio (LAR) and root: shoot ratio (RSR)-were computed for treated plants. Net photosynthesis rate increased by 53% in 0.10 microl liter(-1) NO2 treated plants immediately after exposure compared to control plants. Dark respiration rates were also higher in treated plants. Ch-a, Ch-b and tot-Ch showed significant increases with 0.1 microl liter(-1) NO2 treatment immediately after exposure. Foliar nitrogen content showed an increase in treated plants both immediately after exposure and at maturity. Increases were also seen in RGR and NAR. Plant yield increased by 86% (number of pods), 29% (number of seeds) and 46% (weight of seeds), respectively. Nitrogen dioxide stimulated the overall plant growth and crop yield.     

Effects of ozone on the yield of spring wheat (Triticum aestivum L., cv. Albis) grown in open-top field chambers

Spring wheat (Triticum aestivum L., cv. Albis) was grown in the field at a site located in central Switzerland, and exposed to chronic doses of ozone (O(3)) in open-top chambers to study impacts on yield. The experiment was carried out in 1986, 1987 and 1988. The treatments used included charcoal-filtered air (CF), non-filtered air (NF) and non-filtered air to which constant amounts of O(3) (two levels, O(3)-1 and O(3)-2) were added daily from 09.00 until 17.00 local time. Mean solar radiation-weighted O(3) concentrations during the fumigation period were in the range 0.016-0.022 microl litre(-1) (CF), 0.036-0.039 microl litre(-1) (NF), 0.057-0.058 microl litre(-1) (O(3)-1, used in 1987 and 1988 only) and 0.078-0.090 microl litre(-1) (O(3)-2). Fumigation was maintained from the three-leaf stage until harvest. Ambient plots were used as a reference. Plant characteristics examined included straw yield, grain yield, number of grains per head, number of heads per surface area, weight of individual grains and harvest index (ratio of grain weight to total dry weight). Pollutant concentrations and other environmental parameters were monitored continuously inside and outside the chambers. In 1986 and 1987, enclosure mostly increased the values of different parameters, while in 1988, they were decreased. The negative enclosure effect was due to extremely turbulent winds, which caused lodging inside the chambers. In all 3 years, increasing O(3) concentrations negatively affected the parameters studied, except for the number of heads per surface area, which showed no treatment response. Grain yield showed a very sensitive response to O(3). The effect of O(3) on grain yield was due to an effect primarily on grain size and secondarily on grain number. The relative response of grain yield to O(3) was similar in all 3 years, despite year-to-year differences in climatic conditions and enclosure effects. The analysis of the data for combined years revealed an increase of about 10% in grain yield due to air filtration. The corresponding increase in straw yield was only about 3.5%. Exposure-response models were developed for individual years and combined years. It is concluded that, in the study area, ambient O(3) may affect grain yield in spring wheat.     

Responses of sensitive and tolerant bush beans (Phaseolus vulgaris L.) to ozone in open-top chambers are influenced by phenotypic differences, morphological characteristics, and the chamber environment

Responses of bush bean (Phaseolus vulgaris L.) lines 'S156' (O(3)-sensitive) and 'R123' (O(3)-tolerant), and cultivars 'BBL 290' (O(3)-sensitive) and 'BBL 274' (O(3)-tolerant) to ambient ozone (O(3)) were investigated during the 2001 and 2002 growing seasons. Seedlings were grown in pots inside open-top chambers (OTCs), with charcoal filtered (CF) and non-filtered (NF) ambient air, and in non-chambered ambient air (AA) plots. Growth parameters from individual plants were evaluated after harvests at the end of vegetative (V(4)) and reproductive (R(10)) growth phases. Results at V(4) indicated that CF did not provide additional benefits over NF in 'S156' in 2001 and 2002. In contrast, exposure to CF significantly impaired the growth of 'R123'. At the end of R(10), 'S156' produced more pods, most of which remained immature, and contained fewer seeds or were more frequently aborted, whereas pods produced in 'R123' reached pod maturation and senescence more consistently. Despite increased seed weights inside the OTCs, as observed in 'S156', differences between the two lines were insignificant when grown outside OTCs. Results from the 'BBL 290'/'BBL 274' pair, especially at V(4) phase, remained inconclusive. Plant morphological characteristics, variabilities in environmental conditions, and 'chamber effects' inside OTCs were influential in determining plant response to ambient O(3).     

Effects of various ozone exposures on the susceptibility of bean leaves (Phaseolus vulgaris L.) to Botrytis cinerea

The effects of various ozone exposures in predisposing bean leaves (Phaseolus vulgaris L.) to Botrytis cinerea have been investigated under laboratory conditions. Seedlings of two bean cultivars were exposed to incremental ozone concentrations (120, 180 and 270 microg m(-3) for 8-h day(-1)) for five days and primary leaves were subsequently inoculated with conidia suspended in water or in an inorganic phosphate solution (Pi), and with mycelium. Ozone injury increased with increasing ozone concentration and was much higher in the ozone-sensitive cultivar 'Pros' than in the ozone-insensitive 'Groffy'. Ozone only increased the number of lesions on leaves of Pros after inoculation with either of the conidial suspensions. The Pi-stimulated infection in Groffy was reduced by the lower ozone concentrations. Ozone decreased lesion expansion after inoculation with mycelium. In a chronic fumigation experiment, plants of the two cultivars were exposed to 90 microg m(-3) (7-h day(-1)) and the primary and the oldest tree trifoliate leaves were inoculated after five and seven weeks of exposure. Ozone enhanced the senescence-related injury only in Pros. The number of lesions was not influenced by ozone for either cultivar, conidial suspension or inoculation date. Lesion expansion after inoculation with mycelium was generally reduced in exposed plants. Thus, contrasting effects of ozone on the susceptibility of bean leaves to B. cinerea were observed depending on the cultivar, the conidial suspension, the disease parameter and the ozone exposure pattern. In extrapolating the laboratory results to the field, it is suggested that episodic and chronic exposures to ambient ozone are of minor importance in increasing the susceptibility of bean leaves to B. cinerea.     

The rapid inhibition of root respiration after exposure of bean (Phaseolus vulgaris L.) plants to ozone

The metabolic activity of the roots was very sensitive to the changes induced in the leaves by O₃. Respiratory activity began to decrease well before visible injury appeared on the leaves, and the per cent reduction of respiration was much greater than the per cent leaf injury. The triphenyl tetrazolium chloride (TTC) staining technique revealed changes in root tips very quickly, was generally more sensitive to changes in respiratory activity and was a convenient technique for handling large numbers of samples. Reducing foliar injury from O₃ with low levels of SO₂ reduced the effects on the roots indicating the effect of O₃ is on processes in the leaf.     

Effects of elevated carbon dioxide and ozone on the growth and yield of potatoes (Solanum tuberosum) grown in open-top chambers

Potato (Solanum tuberosum cv. Bintje) was grown in open-top chambers under three carbon dioxide (ambient and seasonal mean concentrations of 550 and 680 mumol mol-1 CO2) and two ozone concentrations (ambient and an 8 h day-1 seasonal mean of 50 nmol mol-1 O3) between emergence and final harvest. Periodic non-destructive measurements were made and destructive harvests were carried out at three key developmental stages (24, 49 and 101 days after emergence) to establish effects on growth and tuber yield. Season-long exposure to elevated O3 reduced above-ground dry weight at final harvest by 8.4% (P < 0.05), but did not affect tuber yields. There was no significant interaction between CO2 and O3 for any of the growth and yield variables examined. Non-destructive analyses revealed no significant effect of elevated CO2 on plant height, leaf number or green leaf area ratio. However, destructive harvests at tuber initiation and 500 degrees Cd after emergence showed that above-ground dry weight (8 and 7% respectively) and tuber yield (88 and 44%) were significantly increased (P < 0.05) in the 550 mumol mol-1 CO2 treatment. Responses to 550 and 680 mumol mol-1 CO2 were not significantly different for most parameters examined, suggesting the existence of an upper limit to the beneficial influence of CO2 enrichment. Significant effects on above-ground dry weight and tuber yield were no longer apparent at final harvest, although tuber numbers were increased (P < 0.05) under elevated CO2, particularly in the smaller size categories. The results show that the O3 treatment imposed was insufficient to reduce tuber yields and that, although elevated CO2 enhanced crop growth during the early stages of the season, this beneficial effect was not sustained to maturity.     

Dose-response studies with the antiozonant ethylenediurea (EDU), applied as a soil drench to two growth substrates, on greenhouse-grown varieties of Phaseolus vulgaris L

To study plant growth and yield effects of the antiozonant ethylenediurea (EDU), which is frequently used for ozone crop loss assessments, dose-response studies were carried out with potted bean plants under greenhouse conditions in winter and spring. Two cultivars of Phaseolus vulgaris L., differing in sensitivity to ozone (O(3)), were grown in unfiltered air on a sandy loam rich in organic matter and on a vermiculite-clay mixture. Four treatments of EDU at concentrations from 300 to 800 mg liter(-1) were given as a soil drench during plant development. Foliar symptoms of EDU phytoxicity were observed at all doses, and plant biomass, particularly pod dry weight, was considerably reduced to increasing doses of EDU. Primary and first trifoliate leaf weight in EDU-treated plants increased as did the number of buds, indicating an extension of vegetative growth and a delay of reproductive processes. 'BBL 290' beans, which are O(3)-sensitive, were injured by EDU more than the O(3)-tolerant 'BBL 274'. The phytotoxic effects of EDU were more pronounced in the synthetic growth substrate than in field soil. In a second experiment, EDU was applied in concentrations from 100 to 400 mg liter(-1) to 'BBL 290' plants, exposed to filtered air or simulated levels of O(3) pollution. In field soil, plant growth and biomass partitioning in filtered air was only slightly altered by EDU, although leaf injury due to EDU occurred. In the vermiculite-clay mix, the biomass of most plant organs, particularly that of roots, was linearly reduced with increasing EDU doses. O(3) did not cause any alteration in plant biomass in field soil-grown and EDU-treated plants. Ozone leaf injury, which affected 67% of primary leaf area in non-treated plants, was completely suppressed by EDU doses as low as 100 mg liter(-1). This indicates that low concentrations of EDU, which do not affect plant growth in field soil, provide sufficient protection from O(3) injury. The need for careful EDU dose-response studies prior to field assessments is emphasized.     

Effects of ozone on 14C translocation velocity and growth of spring wheat (Triticum aestivum L.) exposed in open-top chambers

Growth and yield were reduced but (14)C translocation velocity was not affected by increasing levels of ozone in spring wheat exposed in open top chambers to the following treatments: charcoal filtered air (CF), non-filtered ambient air (NF), or NF with addition of 30 microl litre(-1) ozone, 8 h daily (NFO). Destructive harvests were performed at anthesis and at maturity. Parts of the flag leaf or the second leaf were exposed to (14)CO(2) in small cuvettes for 5 min before, during and after anthesis. The translocation velocity was followed by autoradiography and scintillation counting of the plants frozen and lyophilized at different times after labelling. The label was transported at the same velocity in all the treatments. Ozone induced changes in carbon allocation or partitioning should probably be explained as amounts of carbon transported (mg s(-1)), rather than as transportation velocity (mm s(-1)). The amount translocated may be governed by source conditions under O(3) stress: reduced healthy green biomass and photosynthesis, but perhaps also by impairment of phloem loading because of membrane damage.     

Gravitational infusion of ethylenediurea (EDU) into trunks protected adult European ash trees (Fraxinus excelsior L.) from foliar ozone injury

Adult ash trees (Fraxinus excelsior L.), known to be sensitive or insensitive to ozone, determined by presence or absence of foliar symptoms in previous years, were treated with ethylenediurea (EDU) at 450 ppm by gravitational trunk infusion on six occasions at 21-day intervals in summer 2005 at Turin, Italy. At the end of the season, foliar ozone injury on EDU-treated trees was not complete, but was greatly and significantly reduced when compared to results from trees infused with water. Significant symptom reduction occurred at any crown level in the treated trees suggesting that EDU protected whole crowns. Gravitational infusion of EDU resulted in protection from ozone injury for ozone-sensitive ash trees. The amount of EDU needed to provide protection is assumed to be in the range 13-26 mg m(-2) leaf.     

Effects of ethylenediurea (EDU) on ozone-induced acceleration of foliar senescence in potato (Solanum tuberosum L.)

Experiments were conducted to examine the effects of the anti-ozonant ethylenediurea (EDU) and chronic ozone (O3) exposure on leaf physiology and senescence in an O3-sensitive potato cultivar (Solanum tuberosum L. cv. Norland). A dose-response experiment showed that an EDU concentration of 15 mg l(-1) soil (given as a soil drench) provided complete protection from accelerated foliar senescence induced by exposure to 0.1 microl l(-1) O3 for 5 h day(-1) for 11 days. EDU doses of 45 and 75 mg active ingredient l(-1) soil also gave protection but were associated with symptoms of toxicity and delayed senescence. In further experiments, plants were given 0 or 15 mg EDU l(-1) soil and exposed to clean air or 0.1 microl l(-1) O3 for 5 h day(-1) for 14 days. Chronic O3 exposure in the absence of EDU resulted in accelerated foliar senescence, characterized by early declines in net photosynthesis and Rubisco quantity in O3-treated plants relative to controls. EDU in the presence of O3 gave complete protection against symptoms of accelerated senescence. Senescence was not delayed in plants that received EDU in the absence of O3, and no symptoms of EDU toxicity were evident. The results suggest that EDU-induced tolerance to O3 was not based on 'anti-senescent' properties of this anti-ozonant.     

Use of ethylenediurea (EDU) to ameliorate ozone effects on purple coneflower (Echinacea purpurea)

Purple coneflower plants (Echinacea purpurea) were placed into open-top chambers (OTCs) for 6 and 12 weeks in 2003 and 2004, respectively, and exposed to charcoal-filtered air (CF) or twice-ambient (2x) ozone (O3) in 2003, and to CF, 2x or non-filtered (NF), ambient air in 2004. Plants were treated with ethylenediurea (EDU) weekly as a foliar spray. Foliar symptoms were observed in >95% of the plants in 2x-treated OTCs in both years. Above-ground biomass was not affected by 2x treatments in 2003, but root and total-plant biomass decreased in 2004. As a result of higher concentrations of select cell wall constituents (% ADF, NDF and lignin) nutritive quality was lower for plants exposed to 2x-O3 in 2003 and 2004 (26% and 17%, respectively). Significant EDU x O3 interactions for concentrations of cell wall constituents in 2003 indicated that EDU ameliorated O3 effects on nutritive quality. Interactions observed in 2004 were inconsistent.     

Assessing the impact of ambient ozone on growth and yield of a rice (Oryza sativa L.) and a wheat (Triticum aestivum L.) cultivar grown in the Yangtze Delta, China, using three rates of application of ethylenediurea (EDU)

Foliar applications of ethylenediurea (abbreviated as EDU) were made at 0, 150, 300 or 450 ppm to field-grown rice and wheat in the Yangtze Delta in China. Rice and wheat responded differently to ambient ozone and EDU applications. For wheat, some growth characteristics, such as yield, seed number per plant, seed set rate and harvest index, increased significantly at 300 ppm EDU treatment, while for rice no parameters measured were statistically different regarding EDU application. The reason may be that the wheat cultivar used may be more sensitive to ozone than the rice cultivar. EDU was effective in demonstrating ozone effects on the wheat cultivar, but not on the rice cultivar. Cultivar sensitivity might be an important consideration when assessing the effects of ambient ozone on plants.     

Comparison of a diurnal vs steady-state ozone exposure profile on growth and yield of oilseed rape (Brassica napus L.) in open-top chambers in the Yangtze Delta, China

Most available exposure-response relationships for assessing crop loss due to elevated ozone (O₃) have been established using data from chamber and open-top chamber experiments, using a simulated constant O₃ concentration exposure (square wave), which is not consistent with the diurnal variation of O₃ concentration that occurs in nature. We investigated the response of oilseed rape (Brassica napus L.) to O₃ as affected by two exposure regimes: one with a diurnal variation (CF100D) and another with a constant concentration (CF100). Although the two exposure regimes have the same mean O₃ concentration and accumulated O₃ concentration above 40 ppb (AOT40), our results show that O₃ at CF100D reduced biomass and number of pods/plant more than O₃ at CF100. Both O₃ exposures resulted in larger seed weights/100 pods compared to CF. Numbers of seeds/100 pods were reduced by CF100, while numbers of seeds/100 pods in the CF100D chambers were comparable to those in CF. Our results suggest that chamber experiments that use a constant O₃ exposure may underestimate O₃ effects on biomass and yields.     

Differential protection of ethylenediurea (EDU) against ambient ozone for five cultivars of tropical wheat

The antiozonant EDU (ethylenediurea) was used to assess the impact of ambient O₃ under field conditions on five cultivars of tropical wheat (Triticum aestivum L.). EDU solution (0 ppm and 400 ppm) was applied as soil drench (100 ml plant^?1) 10 days after germination (DAG) at an interval of 12 days. EDU-treated plants showed significant increments in stomatal conductance, photosynthetic rate, variable fluorescence, total chlorophyll, ascorbic acid, proline and protein contents and protective enzymes (POX, SOD and APX) activities in HUW468, HUW510 and HUW234 cultivars, while, a reverse trend was observed for lipid peroxidation. EDU application restored grain yield significantly by maintaining higher levels of antioxidants, metabolites and enzymes in cultivars HUW468 and HUW510. Sonalika and PBW343 showed least response of measured parameters under EDU treatment suggesting their greater resistance to O₃. EDU, thus proved its usefulness in screening suitable wheat cultivars for areas experiencing elevated concentrations of O₃.     

Growth and yield responses of spring wheat (Triticum aestivum L. CV. Turbo) grown in open-top chambers to ozone and water stress

Spring wheat (Triticum aestivum L.) cv. Turbo was exposed to different levels of ozone and water supply in open-top chambers in 1991. The plants were grown either in charcoal filtered air (CF), not filtered air (NF), in charcoal filtered air with proportional addition of ambient ozone (CF1), or in charcoal filtered air with twice proportional addition of ambient ozone (CF2). The mean seasonal ozone concentrations (24 h mean) were 2.3, 20.6, 17.3, and 24.5 nl litre(-1) for CF, NF, CF1, and CF2 treatments, respectively. Ozone enhanced senescence and reduced growth and yield of the wheat plants. At final harvest, dry weight reductions were mainly due to reductions in ear weight. Grain yield loss by ozone mainly resulted from depressions of 1000 grain weight, whereas numbers of ears per plant and of grains per ear remained unchanged. Pollutants other than ozone did not alter the response to ozone, as was obvious from comparisons between CF1 and NF responses. Water stress alone did not enhance senescence, but also reduced growth and yield. However, yield loss mainly resulted from reductions in the number of ears per plant; 1000 grain weight was not influenced by water stress. No water supply by ozone treatment interactions were detected for any of the estimated parameters.     

Protection of plants from ambient ozone by applications of ethylenediurea (EDU): A meta-analytic review

A meta-analysis was conducted to quantitatively assess the effects of ethylenediurea (EDU) on ozone (O₃) injury, growth, physiology and productivity of plants grown in ambient air conditions. Results indicated that EDU significantly reduced O₃-caused visible injury by 76%, and increased photosynthetic rate by 8%, above-ground biomass by 7% and crop yield by 15% in comparison with non-EDU treated plants, suggesting that ozone reduces growth and yield under current ambient conditions. EDU significantly ameliorated the biomass and yield of crops and grasses, but had no significant effect on tree growth with an exception of stem diameter. EDU applied as a soil drench at a concentration of 200-400 mg/L has the highest positive effect on crops grown in the field. Long-term research on full-grown tree species is needed. In conclusion, EDU is a powerful tool for assessing effects of ambient [O₃] on vegetation.