Utilizing plasma-generated N2O5 gas from atmospheric air as a novel gaseous nitrogen source for plants

Fixing atmospheric nitrogen for use as fertilizer is a crucial process in promoting plant growth and enhancing crop yields in agricultural production. Currently, the chemical production of nitrogen fertilizer from atmospheric N2 relies on the energy-intensive Haber–Bosch process. Therefore, developing a low-cost and easily applicable method for fixing nitrogen from the air would provide a beneficial alternative. In this study, we tested the utilization of dinitrogen pentoxide (N2O5) gas, generated from oxygen and nitrogen present in ambient air with the help of a portable plasma device, as a nitrogen source for the model plant Arabidopsis thaliana. Nitrogen-deficient plants supplied with medium treated with N2O5, were able to overcome nitrogen deficiency, similar to those provided with medium containing a conventional nitrogen source. However, prolonged direct exposure of plants to N2O5 gas adversely affected their growth. Short-time exposure of plants to N2O5 gas mitigated its toxicity and was able to support growth. Moreover, when the exposure of N2O5 and the contact with plants were physically separated, plants cultured under nitrogen deficiency were able to grow. This study shows that N2O5 gas generated from atmospheric nitrogen can be used as an effective nutrient for plants, indicating its potential to serve as an alternative nitrogen fertilization method for promoting plant growth. Supplementary Information The online version contains supplementary material available at 10.1007/s11103-024-01438-9.


Introduction
Nitrogen is an essential element for plant growth, and the availability of nitrogen directly impacts crop yield (Cui et al. 2018;Luo et al. 2020).Plants cannot directly utilize atmospheric nitrogen as a nitrogen source, and therefore they rely on nitrogen sources that have undergone nitrogen fixation, converting atmospheric nitrogen into a form that plants can absorb (Charles et al. 2010;Pikaar et al. 2017).The Haber-Bosch process has significantly increased the supply of nitrogen for crop production, leading to higher crop yields and supporting a larger population.Nitrogen fixation through this process surpasses the amounts of nitrogen fixed by living organisms (Pikaar et al. 2017).
As the global population continues to grow, the demand for nitrogen fertilizers rises and will reach to 150 Mton by 2050 (Pikaar et al. 2022).However, the Haber-Bosch method has several drawbacks.Its production requires approximately 2% of the world's annual energy consumption and emits over 300 million tons of CO 2 due to the chemical conversion of highly stable N 2 molecules through reactions at high temperatures and pressures (350-550 °C, 150-350 atm) (Patil et al. 2015).The requirement for large-scale facilities leads to high expenses and the need for construction sites.As a result, nitrogen fertilizers currently represent the largest cost factor in agriculture in industrialized countries and their use is becoming increasingly difficult in developing countries.Furthermore, a significant portion (50-75%) of the supplied nitrogen is not absorbed by plants and is instead lost through leaching into the soil.The nitrogen forms that remain in the soil are a cause of excessive accumulation in soil, which affects plant growth adversely (Liu et al. 2020).Consequently, there is a growing need to explore new, sustainable, and environmentally friendly alternatives to the Haber-Bosch method of nitrogen fixation and nitrogen supplementation (Cherkasov et al. 2015;Fradgley et al. 2021;Chen et al. 2021).
Plasma-based nitrogen fixation is gaining attention as an alternative procedure to supply nitrogen (Patil et al. 2015;Li et al. 2018).Plasma is a highly energetic state of matter that can generate nitrogen oxides by dissociating nitrogen and oxygen molecules present in air (Graves 2012;Ochi et al. 2017).This process can be performed under relatively moderate reaction conditions, such as atmospheric pressure and ambient temperature, and requires minimal energy consumption and start-up time to initiate the process which ensures compatibility with renewable energy sources.Therefore, plasma-based nitrogen fixation is being widely investigated as a promising approach for establishing sustainable agricultural systems (Graves 2012;Patil et al. 2015;Puač et al. 2018;Gao et al. 2022).Traditional plasmabased nitrogen fixation has been employed for the synthesis of NH 3 and NOx (Bogaerts and Neyts 2018).More recently, a plasma device has been developed to generate dinitrogen pentoxide (N 2 O 5 ) from atmospheric nitrogen and oxygen (Sasaki et al. 2021).N 2 O 5 is a highly reactive compound that decomposes at high temperatures and is hygroscopic, readily reacting with water and rapidly converting into two nitrate ions (NO 3 − ).This knowledge inspired us to investigate whether plasma-generated N 2 O 5 gas could function as a nitrogen source for plant culture, i.e. an alternative form of nitrogen fixation.
In this study, we evaluated the potential of plasma-generated N 2 O 5 gas as a nitrogen fertilizer.Addition of N 2 O 5 gas to growth medium supported growth of plants cultured in otherwise nitrogen-deficient medium, however direct and prolonged exposure of plants to the gas itself inhibited growth.To overcome this problem, we successfully tested several alternative approaches for supplying plasma-generated N 2 O 5 gas to the plants.We conclude that plasmageneration of N 2 O 5 has potential to serve as a novel nitrogen fixation procedure that will allow the use of atmospheric nitrogen in agricultural production.

Plant growth conditions
In this research, a series of experiments were carried out using the model plant Arabidopsis thaliana.Plants were grown in a growth chamber (LPH-410S; Nippon Medical & Chemical Instruments Co. Ltd.) set to 16 h light, 8 h dark, 100-120 mmol of photons m −2 s −1 with daylight LEDs, and 22 °C, on vermiculite as substrate.To support growth, each plant was supplied with 100 ml/week of liquid medium by placing the pot holding the plants and vermiculite into a larger container containing the medium.Nitrate sufficient medium (+N medium)

Plasma gas generation with portable plasma N 2 O 5 device
A portable plasma N 2 O 5 generation device was used as previously described to produce N 2 O 5 from air (Sasaki et al. 2021).N 2 O 5 gas generated from dried air contained approximately 310 ppm of N 2 O 5 and was delivered at a flow rate of 2 L/min.When the dry N 2 O 5 gas (2 L/min) was mixed with humidified air (0.2 L/min), N 2 O 5 gas was converted to HNO 3 gas during a 21 s-reaction process in a perfluoroalkoxy alkanes tube (10-m length and 10-mm inner diameter).N 2 O 5 gas and HNO 3 gas were flown through a gas cell with 5 m long multi-optical path for Fourier-transform infrared spectroscopy (FT-IR) for gaseous composition analysis.The detailed quantification procedure was previously described (Kimura et al. 2019).

Supply of N 2 O 5 to plants via liquid medium
Plants were grown with +N medium for 1 week, then pots were transferred to -N medium and grown for another 2 weeks (= nitrogen-deficient plants).To supply N 2 O 5 , 50 ml of -N medium per plant was placed in a container, and N 2 O 5 gas was supplied for 1 (ca. 2 L), 4 (ca.8 L) or 7 min (ca.14 L).The plants with their pots were then transferred to this medium.The N 2 O 5 -dissolved medium was supplied every 3-4 days for a total of three times.The plants were grown for an additional 2 weeks after the last supply and then sampled.As a positive control, +N medium was supplied to nitrogen-deficient plants and, as a negative control, -N medium was supplied to nitrogen-deficient plants.

Supply of N 2 O 5 gas to plants
Plants were grown on +N medium for 1 week, transferred to -N medium and then grown for another 2 weeks (= nitrogendeficient plants).At that point, plants were placed with their pots into secondary containers, and exposed to N 2 O 5 gas directly at approximately 300 ppm at 2 L/min for 1, 4, or 7 min every 3-4 days for a total of three times.After the final treatment, plants were grown for another 2 weeks and then sampled.For the mock treatment, dry air was supplied for 7 min to nitrogen-deficient plants.Wave number (cm -1 ) Gaseous composition of plasma-generated N 2 O 5 gas and HNO 3 gas.A, B N 2 O 5 gas was generated from extremely dry air using an atmospheric pressure plasma device, developed recently (Sasaki et al. 2021).C, D HNO 3 gas was generated by mixing humidified air with the extremely dry N 2 O 5 gas.A, C A typical Fouriertransform infrared spectroscopy (FT-IR) absorbance spectrum of the plasma-generated N 2 O 5 gas and HNO 3 gas (upper half).The best-fit spectrum was obtained with the least square error method.The bestfit spectrum was the summation of the absorbance spectra of the individual species, as shown by the downward absorbance proportional to each density (lower half).B, D Typical densities of individual gaseous reactive species in dry N 2 O 5 gas B and in humidified HNO 3 gas obtained from FT-IR spectra in A and C, respectively.

Supply of N 2 O 5 and HNO 3 to plants
Plants were grown on +N medium for one week, transferred to -N medium and grown for 2 weeks more.The plants were then placed into secondary, conical containers and supplied with N 2 O 5 gas or humidified gas containing HNO 3 and N 2 O 5 (see description of plasma gas generation above) for 4 min every 3-4 days for a total of three times.The plants were grown for another 2 weeks after the final treatment and before sampling.

Long-term exposure to a batch volume of N 2 O 5 gas
Plants grown on -N medium for 2 weeks after sowing were used.N 2 O 5 and HNO 3 gases generated by the plasma device were supplied for 20 min into a 35 L container containing several 0.125 L boxes.A plant was then placed into each 0.125 L box, the box was quickly covered with a lid and placed in a growth chamber for 24 h.The lid was then opened, and the plants remained in the growth chamber, supplied with -N liquid medium until the next treatment.This gas treatment was repeated twice per week, for a total of six treatments.After the final treatment, plants were grown for one more week with -N medium before sampling.

N 2 O 5 is an effective nitrogen source for Arabidopsis thaliana
We employed an atmospheric pressure plasma portable device, which we recently developed, to selectively generate N 2 O 5 gas from ambient air with low power consumption (< 100 W) (Sasaki et al. 2021).The composition of the generated gas was highly dependent on the humidity of the air used as substrate.When dry air was used with the device, most of the generated gas consisted of N 2 O 5 (Fig. 1A, B).The resulting dry gas contained 311 ppm of N 2 O 5 .When and the following plant culture.B Plants were initially grown with a nitrogen-sufficient medium containing Ca(NO 3 ) 2 (+N) for a week (gray box) and then shifted to nitrogen-deficient medium (N) for 2 weeks (white box).After that, plants were supplied with medium treated with N 2 O 5 gas (for 1, 4 or 7 min, see Methods for details) every 3-4 days for a total of three times (green box).Following these treatments, the plants were grown on N medium for two more weeks (white box) before being photographed and sampled for phenotype analysis.For further details refer to Methods and Supplemental Table S1.C Photograph of plants taken at the end of the experiment.D Fresh weight of the plants.This experiment was repeated twice.
The data are presented as mean ± s.d. of n ≥ 3 plants, analyzed using one-way ANOVA ▸ the dry N 2 O 5 containing air was mixed with humidified air, the N 2 O 5 gas immediately converted to HNO 3 , and the main product was HNO 3 (Fig. 1C, D).The resulting humidified gas contained 433 ppm of HNO 3 and 73 ppm of N 2 O 5 .
To evaluate the potential of N 2 O 5 gas to be used as a nitrogen source for plant growth, we applied the generated N 2 O 5 gas to nitrogen-free medium for 1, 4 or 7 min and used this medium to treat nitrogen-deficient plants (Fig. 2A).Specifically, after culturing plants with nitrogen-sufficient medium for 1 week, they were cultured with nitrogen-free medium for 2 weeks to induce nitrogen-deficiency (Fig. 2B and Supplemental Table S1).Then, the plants in the pots were supplied with medium that had been exposed to N 2 O 5 gas (for 1, 4 or 7 min).Plants remained in this medium for 1 week and were then transferred back to medium without nitrogen source.Growth of plants treated with medium exposed to N 2 O 5 gas for 1 or 4 min was similar to that of plants grown under nitrogen-deficient conditions (Fig. 2C,  D).However, plants treated with medium exposed to N 2 O 5 gas for 7 min grew as well as those grown with nitrogensufficient medium that contained Ca(NO 3 ) 2 (Fig. 2C, D).Furthermore, accumulation of anthocyanins, a symptom of nitrogen deficiency, decreased the longer the medium had been exposed to N 2 O 5 (Fig. 2C).This suggested that plasmagenerated N 2 O 5 gas can be used as a nitrogen source for plant growth.

Treating plants directly with plasma-generated N 2 O 5 gas has negative effects on growth
We next investigated whether N 2 O 5 gas could be used to promote plant growth by directly supplying it to the plants as a gas.Considering the quick conversion of N 2 O 5 gas to NO 3 − in water (Fig. 1), we predicted that N 2 O 5 gas might be readily absorbed by the vermiculite and plant tissues (leaves and stems).N 2 O 5 gas generated by the same portable plasma device was applied to plants over a short period of time (1-9 min).Under those conditions, symptoms of plant damage were observed after about 4 min of treatment, and significant changes in the plants were evident after 7 min of the treatment (Fig. 3B).Plants exposed to the gas for 7 min began to wilt and change their leaf color within minutes (Fig. 3B and Supplemental Fig. S1).Therefore, the direct supply N 2 O 5 through the portable plasma device had negative impacts on plant health.
To supply a sufficient amount of nitrogen to the plants without prolonged direct exposure to N 2 O 5 gas, we instead supplied N 2 O 5 gas to the plants for a limited period of 1, 4, or 7 min at three separate times, 3-4 days apart (Fig. 4A, B and Supplemental Table S1).Reproductive-stage plants grown under nitrogen-deficient conditions were used for this experiment.Leave damage was observed in plants exposed to N 2 O 5 gas for 4 min and 7 min, and fresh weight was reduced by 71% and 89% when plants were treated three times for 4 min and 7 min, respectively, compared to the mock-treated plants (Fig. 4C, D).However, plants exposed to N 2 O 5 gas three times for 4 or 7 min had green leaves, and symptoms of nitrate deficiency disappeared (Fig. 4C).Plants exposed to N 2 O 5 three times for 1 min showed accumulation of anthocyanin in their leaves although their fresh weight was almost the same as that of the mock-treated plants.
We evaluated the effects of mixing N 2 O 5 gas with moist air on plant growth (Fig. 4e) because under high humidity N 2 O 5 was converted to HNO 3 gas (Fig. 1).Air was presaturated with humidity by flowing through a Gas washing bottle bubbler, and then mixed with N 2 O 5 gas (Fig. 4E).The resulting gas mixture was added to the plant growth chamber every 3-4 days for 4 min, a total of three times (Fig. 4B, Table S1).Control treatment with N 2 O 5 mixed with dry air was done the same as shown in Fig. 2. When plants were exposed to N 2 O 5 gas mixed with moist air, they produced less biomass but continued to look healthy without chlorosis (Fig. 4F).Plants treated with dry N 2 O 5 gas had an 88% increase in fresh weight, compared with plants treated with the humidified N 2 O 5 gas (Fig. 4G).These data suggested that treating plants with dry N 2 O 5 gas was better for plant growth than treatment with N 2 O 5 gas mixed with moist air.

Alternative modes of application of N 2 O 5 to plants
The direct supply of N 2 O 5 gas as shown in Figs. 3 and 4 inhibited the growth of plants.Therefore, we tested an alternative method of supplying nitrogen directly from the N 2 O 5 generator to plants in nitrogen-free medium.For this, we physically separated the plant culture process from the treatment with N 2 O 5 (Fig. 5A, B). the device (either mixed with dry or moist air) was introduced for 20 min into a container holding several smaller containers but no plants.After that, nitrogen-deficient plants were placed into the smaller containers which were covered with lids.Note that this process was feasible since N 2 O 5 gas has a higher density than air.The covered containers with the plants were moved to a growth chamber.After 24 h, the lids were removed, and plants were again supplied with nitrogen-free medium (Fig. 5A).This procedure was repeated twice per week, for a total of six times, followed by an additional week of growth with nitrogen-free medium at the end (Fig. 5B and Supplemental Table S2).In this result, plants treated with dry air could hardly grow on vermiculite and this suggested vermiculite did not have sufficient amount of nitrogen for plant growth.Plants treated with N 2 O 5 gas had increased fresh weight, similar to the plants treated with HNO 3 gas (resulting from mixing N 2 O 5 gas with moist air) (Fig. 5C, D).These results showed that supplying N 2 O 5 gas to plants can be an effective method for supplying nitrogen fertilizers to plants without relying on traditional nitrogen fixation processes.

Discussion
N 2 in the air is an inert gas that is not readily available as a plant nutrient but instead requires conversion by energyintensive conventional nitrogen fixation processes.Our study shows that N 2 O 5 gas generated with a portable plasma device can be a new avenue of supplying a nitrogen source to plants.This method can be expected to be useful as an alternative nitrogen fertilization procedure to support plant growth while preventing the excessive use of conventional nitrogen fertilizers.However, direct administration of N 2 O 5 to plants inhibited growth (Fig. 3).Hence, it was crucial for us to develop an effective method for delivering N 2 O 5 to the plants.In this study, we explored various methods for administering N 2 O 5 gas to plants.When plants were cultured using liquid medium that had been exposed to N 2 O 5 gas, their growth was not impaired (Fig. 2).But repeated shortterm application of the N 2 O 5 gas directly from the plasma device to nitrogen-starved plants damaged shoot tissues and only partially improved plant growth (Fig. 4).These problems could be avoided by placing nitrogen-deficient plants into a chamber pre-filled with N 2 O 5 , and subsequently growing them in medium without a nitrogen source (Fig. 5).Our results demonstrated that plants could be grown from seeds on nitrogen-free medium, with nitrogen supplied from the air, in the form of N 2 O 5 gas if the duration of contact with the plants or the concentration of the gas was reduced.
The portable plasma device used in this study consumes less than 100 W of power, which can be efficiently supplied from renewable energy sources, and the only material used is air.This device excels in its portability and ability to produce a nitrogen source from air using minimal power, without the need for large facilities.The device is an efficient N 2 O 5 generator that can selectively produce N 2 O 5 , minimizing waste (Fig. 1) (Sasaki et al. 2021).On the other hand, the energy efficiency of nitrogen fixation by the plasma device is about 72 MJ/(mol N) (Sasaki et al. 2021), and that by the Haber-Bosch method is about 0.5 to 0.6 MJ/(mol N) (Rouwenhorst et al. 2021), indicating that the Haber-Bosch method is still superior to the portable plasma device in terms of energy efficiency per mole of nitrogen.
The generated N 2 O 5 gas contained some impurities including O 3 (Fig. 1), which is known to cause cell death and photosynthesis inhibition in Arabidopsis even at very low concentrations at the ppb level.As shown in Fig. 3 and  4, direct exposure of plants to N 2 O 5 gas produced by the device resulted in growth impairment, potentially due to the presence of O 3 or rapid acidification due to the chemical reaction of N 2 O 5 and H 2 O.However, when the supply of plasma-generated N 2 O 5 gas was more limited, the plants grew well (Fig. 5).
Foliar application of liquid fertilizers is used in modern agriculture.This technique is especially effective in plants with weakened roots and under continuous cloud cover (high humidity) where transpiration does not take place (Fernández et al. 2021;Naqve et al. 2021).Therefore, foliar application allows nitrogen to be supplied directly to the tissues that need it.Foliar nitrogen is supplied as water-soluble compounds including NO 3 − , NH 4 + , urea and amino acids (Lv et al. 2021).This method has the disadvantage of not supplying nitrogen Fig. 4 Direct exposure to plasma-generated N 2 O 5 gas impaired plant growth.A Experimental set-up for the treatment of plants with N 2 O 5 and the following plant culture.B Plants were grown on +N medium for 1 week (grey box), followed by growth on -N medium for two weeks (white box), and then treated with dry air (mock) or with N 2 O 5 gas added directly to the vapor phase of the container.The gas was injected for 1, 4, or 7 min each time, every 3-4 days for a total of three times (green box).After the final treatment, the plants remained on -N medium for two more weeks (white box) before being photographed and sampled for phenotype analysis.C Photographs of plants taken at the end of the experiment.D Fresh weight of the plants.This experiment was repeated twice.The data are presented as mean ± s.d. of n = 3 plants, analyzed using one-way ANOVA.E Experimental setup for the treatment of plants with dry or humidified N 2 O 5 gas and the following plant culture, providing a simplified method of supplying N 2 O 5 and HNO 3 to plants.The experimental schedule was essentially the same as B except for the N 2 O 5 treatment.Three-week-old plants were either exposed to generated N 2 O 5 directly or to N 2 O 5 that had been additionally passed through humid air for 4 min.The treatment was performed every 3-4 days for a total of three times before plants were photographed and sampled for phenotype analysis.F Photographs of plants taken at the end of the experiment.G Fresh weight of the plants.This experiment was repeated twice.The data are presented as mean ± s.d. of n = 3 independent plants P < 0.05, ns not significant (P > 0.05); Student t-test.Scale bars = 1 cm ◂ uniformly due to high leaf overlap and plant density.In contrast, gaseous nitrogen fertilizers can even penetrate into small spaces between leaves.The chemical polarity of N 2 O 5 is significantly lower than that of HNO 3 .This lower polarity may promote the infiltration of N 2 O 5 across the leaf surface, consisting of the cuticular wax and cell wall layers.Once taken up into the cells, the buffering capacity of the cytoplasm may ameliorate any acidification resulting from the conversion of N 2 O 5 to HNO 3 .This may explain why treatment with dry N 2 O 5 was more effective in enhancing plant growth than when N 2 O 5 was mixed with moist air (as shown in Fig. 4F,  G).Moreover, it was recently reported that exposure of Arabidopsis thaliana to N 2 O 5 gas leads to the induction of an immune response during a pathogen inoculation test (Tsukidate et al. 2022).This indicates that N 2 O 5 is absorbed by the surface of the plant tissue.The immuneinducing effect of N 2 O 5 also increases its superiority for use as a nitrogen fertilizer for plants.Our study supports the idea that air-derived N 2 O 5 has potential to be used as a foliar nitrogen fertilizer altered to or with conventional nitrogen fertilizer although N 2 O 5 was also absorbed by the soil and the medium.
In conclusion, we have successfully applied N 2 O 5 gas as alternative nitrogen source that improves plant growth.This ability to directly utilize N 2 , which is abundantly or HNO 3 gas generated according to the process shown in Fig. 1B and containers remained sealed for one day.After that, the plant pots were removed from the containers and placed directly into the growth chamber for 2 or 3 days.This treatment cycle was repeated five times for a total of 6 treatments over the course of 3 weeks (green box, arrowheads indicate treatments).After the final treatment, plants were grown for another one week in a growth chamber (white box) before being photographed and sampled for phenotype analysis.Further details refer to Materials and Methods and Supplemental TableS2 .C Photographs of plants at the end of the experiment.Scale bars = 1 cm.D Fresh weight of the plants.This experiment was repeated twice.The data are presented as mean ± s.d. of n = 4 plants, analyzed using one-way ANOVA present in the Earth's atmosphere, as a gaseous nitrogen source is a promising development for plant cultivation and agriculture.The method established in this study will even make it feasible to reliably supply nitrogen to regions of the world where nitrogen fertilizer production has been challenging or impossible, such as in less developed regions.
Fig.1Gaseous composition of plasma-generated N 2 O 5 gas and HNO 3 gas.A, B N 2 O 5 gas was generated from extremely dry air using an atmospheric pressure plasma device, developed recently(Sasaki et al. 2021).C, D HNO 3 gas was generated by mixing humidified air with the extremely dry N 2 O 5 gas.A, C A typical Fouriertransform infrared spectroscopy (FT-IR) absorbance spectrum of the plasma-generated N 2 O 5 gas and HNO 3 gas (upper half).The best-fit

Fig. 2
Fig.2Plasma-generated N 2 O 5 is an effective nitrogen source for plants.A Experimental set-up for the treatment of plants with N 2 O 5 and the following plant culture.B Plants were initially grown with a nitrogen-sufficient medium containing Ca(NO 3 ) 2 (+N) for a week (gray box) and then shifted to nitrogen-deficient medium (N) for 2 weeks (white box).After that, plants were supplied with medium treated with N 2 O 5 gas (for 1, 4 or 7 min, see Methods for details) every 3-4 days for a total of three times (green box).Following these treatments, the plants were grown on N medium for two more weeks (white box) before being photographed and sampled for phenotype analysis.For further details refer to Methods and Supplemental TableS1.C Photograph of plants taken at the end of the experiment.D Fresh weight of the plants.This experiment was repeated twice.The data are presented as mean ± s.d. of n ≥ 3 plants, analyzed using one-way ANOVA

Fig. 3
Fig. 3 Effects of direct supply of N 2 O 5 gas to plant seedlings.A Experimental set-up for the treatment of plants with N 2 O 5 and the following plant culture.B Plants grown for 1 week on +N medium were transferred to -N medium and grown for another 2 weeks.Then N 2 O 5 gas was directly supplied to each plant for 1 to 9 min.Mock treatment consisted of dry air supplied to the plants for 9 min.C Photographs of plants, taken after one treatment First, N 2 O 5 gas generated by

Fig. 5
Fig. 5 Supplementation of generated N 2 O 5 to plants without direct exposure to N 2 O 5 gas.A Experimental set-up for the treatment of plants with N 2 O 5 and the following plant culture.B After germinating seeds on -N medium and growing them for 2 weeks (white box),the pots were placed in containers filled with plasma-generated N 2 O 5 or HNO 3 gas generated according to the process shown in Fig.1Band containers remained sealed for one day.After that, the plant pots were removed from the containers and placed directly into the growth chamber for 2 or 3 days.This treatment cycle was repeated five times