Growth, Yield and Quality of Maize under Ozone and Carbon Dioxide Interaction in North West India

Maize is an important C4 crop and how it will respond to elevated atmospheric carbon dioxide and ozone levels is not well documented. To understand how the growth and nutritional quality of maize will be affected under elevated carbon dioxide (CO2) and tropospheric ozone (O3) interaction, a field experiment was conducted under free air O3 and CO2 enrichment rings (FAOCE) growing HQPM-1 and PMH-1 maize cultivars at New Delhi, India. Each cultivar was exposed to ambient and elevated CO2 (559 ppm) alone and along with ambient and elevated O3 (71.8 ppb) throughout the growing period. Elevated CO2 (EC) significantly increased the leaf area index (10.8–22.5%), chlorophyll (11.2–17.3%) and photosynthetic rate (12.1–16.5%) in the two cultivars over the ambient. O3 exposure of 27 ppm hr (AOT4O) under elevated O3 (EO) treatment led to a significant decline in yield (p < 0.01) by 9.2% in HQPM-1 and 9.8% in PMH-1. Under elevated CO2 the increase in grain yield was higher under HQPM-1 (25.4%) as compared to PMH-1 (9.04%). The protein content increased under EO (8.1–12.5%) and decreased under EC (13.4–13.6%) in the two maize cultivars due to yield dilution effect. Lysine, phosphorus and potassium content of the grain significantly decreased in both the cultivars under elevated CO2. Carbohydrate and amylose concentrations in grains increased (9.9–15.5%) under EC and decreased (10.8–16.7%) under EO, however, no significant change in yield, protein, amylase, carbohydrate, lysine, potassium and phosphorus was observed under the interaction treatment ECO as compared to the ambient. After two years of study we could conclude that elevated CO2 (559 ppm) was able to offset the negative effect of elevated O3 (71 ppb) on grain yield by 11.2% in PMH-1 and by 18.8% in HQPM-1 without significantly affecting the grain quality in both the maize cultivars.


INTRODUCTION
Climate change and air pollution are increasing threats to agriculture production and food security (Porter et al., 2014;Tai et al., 2014). Due to the incessant burning of fossil fuels and anthropogenic activities, the concentration of CO2 has increased by 40% (400 ppm) since preindustrial era (1750 AD) and may increase to 550 ppm by 2050 (IPCC, 2014). The greenhouse gases such as carbon dioxide (CO2) and tropospheric ozone (O3), also a phytotoxic air pollutant have bearings on plant growth and productivity. Atmospheric CO2 is a source of carbon assimilation in plants and works as a carbon fertilizer, resulting in improved plant growth and production (van der Kooi et al., 2016;Lopes et al., 2018). However, according to literature plants that use the C4 photosynthetic pathway are less responsive to elevated CO2 as compared to plants that use the C3 pathway. Maize being a C4 plant, may thus behave differently under elevated CO2 as compared to C3 crops and there have been contrasting reports about the impact of elevated CO2 in maize. Abebe et al. (2016) observed an increase of 53.7% in the grain yield of maize whereas, Leakey et al. (2006) observed reductions in photosynthesis, biomass and yield under elevated CO2 condition. Furthermore, CO2 enrichment leads to a decline in nutrients present in the grains such as zinc, iron and protein content (Myers et al., 2014;Broberg et al., 2017). Previously it has been reported that elevated CO2 reduced N, crude protein, P and K contents of grains (Abebe et al., 2016). Recently, Reich et al. (2018) observed a role reversal of the C3-C4 concept wherein they observed that after 12 year of experimentation, the biomass increased under elevated CO2 in C4 plants and decreased under C3 crops.
The concentration of ground level O3 is increasing continuously since industrial revolution and has reached 35-40 ppb globally and it may continue to increase rapidly in the developing countries in the 21st century (Frei, 2015;Proietti et al., 2016;Sicard et al., 2017) According to Environmental Protection Agency if global emissions of O3 precursors continue at present day rate then the tropospheric O3 may reach more than 70 ppb level by 2050 (Pfister et al., 2014). O3 is produced in the troposphere due to photochemical reactions of the oxides of nitrogen (NOx) and the volatile organic compounds (VOCs) including methane and CO2 (Bhatia et al., 2012). Several researchers reported that higher concentration of tropospheric O3 reduced carbon storage of crops, thereby reducing plant growth and yield (Bhatia et al., 2011;Zhu et al., 2011;Tai et al., 2014;Broberg et al., 2015;Tomer et al., 2015). The elevation of tropospheric O3 in the free air significantly increases P, K and protein concentrations in the cereals . However, most of the research on the effect of elevated tropospheric O3 on photosynthesis and stomatal function has been carried out in C3 species only (Ainsworth, 2017).
Besides the individual impacts of elevated CO2 and O3, the combined interaction of the elevated CO2 and O3 influence the crop growth and development. The enrichment of CO2 mitigated the adverse impacts of elevated O3 in many studies (Kumari et al., 2015;Phothi et al., 2016;Singh et al., 2018). Torbert et al. (2004) in a study in grain sorghum reported that elevated CO2 increased the total biomass production, however, the interactive influences of elevated CO2 and O3 have very rarely been studied in C4 crops. Bhatia et al. (2014) observed a significant reduction in the growth and yield of maize under elevated O3 in a study conducted in open top chambers. They reported that exposure of elevated (ambient+40-45 ppb) O3 resulted in greater susceptibility of the maize plant to pest attack. Very few studies have been carried out in C4 crops relative to C3 crops and thus there is a generally a poorer understanding of C4 responses to climate change (Leakey, 2009).
Maize a C4 cereal crop is the third most important crop after rice and wheat across the world. It is being produced in 189.3 million hectares and contributes 36% (1078 Mt) to the world food production (FAO, 2016) and about 70% of it is grown in the developing nations. In India, maize is mainly grown in Kharif season (June-October) with a total production of 24.4 million tons and 2.5 ton ha -1 productivity, accounting for 9% of the total food grain production (Government of India, 2017). Due to the declining ground water tables and poor soil fertility in the rice growing areas of Indo-Gangetic Plains, maize has been suggested as an alternate crop to diversify from rice for improving soil quality and reducing the global warming potential of soils in this region. However, increasing concentration of tropospheric O3 in India, especially in the Indo-Gangetic Plains (IGP) is of major concern as it poses a threat to the quality and productivity of crops (Singh and Agrawal, 2017) and may reach 70 ppb by 2050 in South Asia (IPCC, 2014). Apart from this most of the studies on the impact of tropospheric O3 on crops have been conducted in Europe and America (temperate regions). Very few studies have been carried out in the tropical and semi-arid regions. Keeping in view the contrasting response obtained in C4 crops under elevated CO2, the present study was carried out with the aim to assess the impact of elevated CO2 and tropospheric O3 separately as well as in combination on the growth, yield and nutritional quality of maize under free air O3 and CO2 enrichment.

Site Description
The experiment was carried out in free air O3 and CO2 enrichment (FAOCE) rings at the farm of Indian Agricultural Research Institute (IARI), New Delhi, India, during the summer season of 2016 and 2017. The site is located at (28°37′N latitudes, 77°12′E longitudes at mean sea level of 228.6 meters in an overlap of monsoon-influenced humid subtropical and semi-arid climate with average annual precipitation of 797 mm. The weather parameters recorded at the site are given in Table1. The experimental site had a typic Ustochrept soil, having sandy loam texture. The soil had a total organic carbon, P, K and total N content of 0.45%, 25.5 kg ha -1 , 249 kg ha -1 and 188 kg ha -1 , respectively.

Treatment and Experiment Design
Two maize (Zea mays L) cultivars HQPM-1 (high quality protein) and PMH-1 (normal) were sown, in free air ozone and carbon dioxide enrichment (FAOCE hereafter) rings having an area of 19.6 m 2 (5.0-meter diameter) each. Four treatments were taken for the study: ambient CO2 and O3 (Amb), elevated CO2 and ambient O3 (EC), elevated O3 and ambient CO2 (EO) and elevated CO2 and elevated O3 (ECO). Each FAOCE ring was divided in two halves (9.8 m 2 ) and each cultivar was sown in one half of the ring. There were two replicate rings for each of the four treatments. The crops were grown during 2016 and 2017 and were exposed to elevated levels of O3 and CO2 during the entire crop growth period. The targeted levels were 550 ppm of elevated CO2 and 70 ppb of tropospheric O3. The ambient CO2 ranged from 388 to 416 ppm and from 389 to 419 ppm during 2016 and 2017 respectively. The elevated CO2 ranged from 505 to 623 ppm and from 508 to 613 ppm in the FAOCE rings during 2016 and 2017 respectively. The wind vane and anemometer were installed in the center of the FAOCE at 1-meter height to monitor the wind direction and speed, respectively. For achieving the targeted elevated CO2 level (550 ppm), pure CO2 was released from commercial grade compressed 30 kg pure CO2 gas cylinders, fitted with a regulator (DURA, ESAB, India) through solenoid valves and PVC tubing to the base of each FAOCE plot with laser holes (0.3mm diameter) at a distance of 30 cm to spray CO2 in the rings uniformly. Releasing of the elevated concentration of compressed CO2 was controlled by the solenoid valves as per the wind direction and speed in the FAOCE rings. O3 was produced by an ozone generator (Eltech Engineers, Mumbai) by converting atmospheric oxygen into O3 using UV lamps. Air blowers were used to mix O3 with air and the mixture of air and O3 was controlled by the flappers using solenoid valves. Air from the center of each ring was monitored for the concentrations of CO2 by using a CO2 analyzer (NDIR, Topak USA) and an ozone analyzer (2B Technologies) was used for monitoring O3 levels. Microprocessors were used to perform control, logging and operation of CO2 and O3 concentrations through digital input and output modules as per wind direction and velocity on real time basis. The exposure of elevated CO2 and tropospheric O3 was given during day time (10.00 am to 17.00 pm) when the plant photosynthesis was taking place to save on costs as maintaining elevated CO2 was very costly. The crop was exposed to elevated tropospheric O3 for 7 hours as has been most commonly used practice in studies worldwide (Ashmore, 2005). Maize cultivars were sown on June, 24 and June, 25 in 2016 and 2017, respectively. Planting was done at 20 cm plant to plant and 50 cm row to row distances. The crops were fertilized with nitrogen (N) (150 kg ha -1 ), P2O5, and K2O (70, kg ha -1 ) and zinc sulphate (Zn SO4) (25 kg ha -1 ). The complete doses of Zn SO4, P2O5 and K2O and 50% of the nitrogen (N) were applied at the time of crop planting. The remaining amount of nitrogen was given in two equal parts at the knee high (35 DAS) and tasseling stage (65 DAS) of crop growth.

AOT40 Calculation
The O3 concentration during crop growth period ranged from 4 to 41.1 ppb and from 6 ppb to 38.5 ppb in the ambient air during 2016 and 2017, respectively (Fig. 1). The elevated O3 ranged from 60 to 88 ppb and from 55 to 87 ppb in the FAOCE rings during 2016 and 2017, respectively. The average O3 in the ambient air during crop growth period was 30.07 and 28.63 ppb whereas average elevated O3 in the FAOCE rings was 72.75 and 71.04 ppb during 2016 and 2017. AOT40 which is the sum of hourly average values of O3 concentration beyond 40 ppb  was calculated from the elevated levels of O3 in the FAOCE rings during the complete crop growth period and was 27441 ppb hr and 26292 ppb hr in 2016 and 2017 respectively.

Measurement of Plant Growth and Yield Parameters
Leaf area index (LAI) was measured by plant canopy analyzer (LAI-2200 C) at tasseling and silking stages of crop growth. The photosynthetic rates and stomatal conductance were also measured at tasseling and at silking stages on the third leaf of the plant from the top around 11.00 am by using portable photosynthesis system (Model No. LI-6400 XT; LI-COR, USA). For evaluating yield attributes of crops, maize plants from 1 m 2 area were harvested at maturity from each replicate. The cob length (cm), cob diameter (cm), grains number (cob -1 ), grain weight (g cob -1 ), biological yield (g m -2 ), grain yield (g m -2 ) and 100 grain weight (g) were determined as per standard procedures. Yield data was adjusted to 10% moisture content after drying (65°C) in an oven. The ratio of grain yield and biological yield multiplied by 100 was expressed as harvest percent.

Plant and Grain Sample Analysis
Chlorophyll content of leaves of both the cultivars was determined at tasseling as well as silking using procedure described by Wellburn (1994). Nutritional quality parameters such as potassium (K) and phosphorus (P) were determined as per method described by Jakson (1973). The crude protein content was calculated by multiplying the conversion coefficient 5.95 with the total N concentration (Juliano, 1993) which was determined by the Kjeldahl method. The carbohydrate concentration of the grains was determined by the method given by Sadasivam and Minickam (1992). The lysine content of grains was estimated as per procedure described by Tsai et al. (1972). The amylose content was determined by procedure described by Hu et al. (2010).

Statistical Analysis
Using the 9.3 SAS statistical software, growth, yield, yield components and quality attributes of maize were analyzed. Split plot design was used to conduct the experiment. Two-way analysis of variance (ANOVA) was used to analyze the impact of cultivar and the four treatments. The statistically significance of data was compared based on their 95% confidence level (p < 0.05) (Gomez and Gomez, 1984).

Impact of Elevated CO2 and O3 on Photosynthetic Rate, Stomatal Conductance, Chlorophyll Content and Leaf Area Index
The mean photosynthetic rates (Pn) ranged from 33-34.8 µmole CO2 m -2 s -1 under EO and from 41.8-44.4 µmole CO2 m -2 s -1 under EC across the crop growth stages and cultivars. It increased significantly by 12.1-16.6% (p < 0.05) in HQPM-1 and by 12.4-15.5% (p < 0.05) in PMH-1 under EC treatment across the crop growth stages in the two years (average data) (Table 2). However, increased tropospheric O3 (EO) significantly reduced the Pn of HQPM-1 (7.8-9.4%) and PMH-1 (6.7-15.4%) (p < 0.05) ( Table 2) at both crop stages as compared to the ambient. The Pn in the interactive treatment ECO increased by 11.9-12.4% for HQPM-1 and by 10.9-18.5% for PMH-1 (across the stages) over elevated O3 alone and was found to be at par with the ambient.
The stomatal conductance (gs) significantly decreased (p < 0.01) in both the cultivars under EC, EO and ECO as compared to the ambient ( Table 2). The lowest gs was observed in EC treatment at tasseling stage in PMH-1 cultivar.
Similar to the Pn the total chlorophyll content of leaves increased under EC and reduced under EO as compared to the ambient in both the cultivars (Table 2). In the ECO treatment, elevated CO2 negated the effect of O3 on the chlorophyll content in HQPM-1 (9.8-11%) and PMH-1 (4.8-10.4%) over elevated O3 alone.
Significant difference in LAI was observed between the cultivars. Increased concentration of EO led to a higher decline in LAI of HQPM-1 (13.2-16.2%) as compared to PMH-1 (7.5-8.5%) at tasseling and silking stages. Under ECO, a higher increase in LAI was observed for HQPM-1 (12.1-16.1%) as compared to PMH-1 (5.4-8.5 % in PMH-1) over EO across the years and stages.

Impact of Elevated CO2 and O3 on Yield Components
The cob length (cm) and grains (cob -1 ) reduced in both cultivars under EO. Elevated CO2 significantly (p < 0.01) increased the cob length as well as grains (cob -1 ) in both crop cultivars as compared to the ambient (Amb). Under the interactive treatment (ECO) averaged cob length increased by 9.4% for HQPM-1 and by 5.8% for PMH-1 over EO alone and was found to be at par with the Amb (Table 3). Average grain weight (cob -1 ) under EC significantly increased in HQPM-1 (22.1%)-and in PMH-1 (5.32% (Table 3). While, EO reduced the grain weight (cob -1 ) by 5.8% in HQPM-1 and by 11.3% in PMH-1 as compared to the Amb (Table 3). Under the interactive treatment ECO grain weight (cob -1 ) increased by 12.2% in HQPM-1 as compared to EO alone, whereas there was no significant difference in PMH-1. The 100 grain weights of HQPM-1 and PMH1 were not-significantly different under ECO treatment as compared to the ambient during both the years.
Grain yield of HQPM-1 increased significantly in the first (33.3%, p < 0.01) and the second year (17.4%, p < 0.01) under EC as compared to the Amb. Similarly, grain yield of PMH-1 responded positively to the increased CO2 during the first (10.5%) (p < 0.01) and the second year (7.5%) ( Table 3). Under the interactive treatment ECO, averaged grain yield increased by 18.8% in HQPM-1 and by 11.2% in PMH-1 as compared to the elevated O3 alone.

Impact of Elevated CO2 and O3 on Quality of the Grains
The mean of the two years' data revealed that the elevated CO2 concentration in free air declined the crude protein by 8.1% in HQPM-1 and by 12.5% in PMH-1. On the contrary, the enrichment of O3 increased the crude protein significantly (p < 0.05) by 13.5% in HQPM-1 and by 13.4% in PMH-1 as compared to the ambient (Fig. 2(a)). The EC treatment diminished the positive influence of elevated O3 on protein concentration and it decreased by 8.2 and 8.8% for HQPM-1 and PMH-1, respectively under the ECO treatment as compared to EO alone. The protein contents in both the crop cultivars under the interactive treatment (ECO) were at par with the Amb (Fig. 2(a)).
Carbohydrate content of grains increased in HQPM-1 (4.6%) and PMH-1 (15.5%) under the EC, but responded negatively to EO in HQPM-1 (15.3%) and PMH-1 (16.6%) as compared to the ambient treatment ( Fig. 2(b)). No significant change was obtained in the carbohydrate content    in maize grains under ECO in both the cultivars as compared to the Amb. Phosphorus (P) (16.2-23.6%) and potassium (K) (8.9-17.3%) concentrations significantly decreased (p < 0.01) under elevated CO2 treatment in both the cultivars as compared to the Amb (Table 4(c)). However, P (2.3-4.9%) and K (2.5-19.3%) increased under elevated tropospheric O3. The interactive environment under ECO did not lead to any change in the P and K concentrations of grains as compared to the ambient (Figs. 2(c) and 2(d)). The amylose content also varied similar to the carbohydrate content of grains in the different treatments ( Fig. 2(f)).
Lysine is an important α amino acid that is needed for the biosynthesis of proteins. It ranged under from 0.30 to 0.32% under elevated O3 (EO) and from 0.27 to 0.28% under elevated CO2 (EC). Lysine contents of maize grains decreased under EC by 3.7-8.4% and increased by 6.5-7.3% under EO over the ambient treatment in both HQPM-1 and PMH-1 maize cultivars (Fig. 2(e)).

Varietal Response to Yield and Quality under Different Treatments
During the first year AOT 40 of 27741 ppb hr of O3 exposure led to decline in grain yield in HQPM-1 by 9.5%, and by 15.7% % in PMH-1, while during the second year AOT exposure of 26292 ppb hr, decreased the grain yield of HQPM-1 by 8.7% and by 3.8% in PMH-1. On an average the exposure of AOT4O of 27016-ppbhr O3 led to the yield loss of 9.2% in HQPM1 and 9.8% in PMH1 during the two years of the study. Under EC there was higher increase in grain yield of HQPM-1 as compared to PMH-1. Under ECO the grain yield of HQPM-1 increased by (6.5-9.5%) over ambient but in case of PMH-1 it was not significantly different than ambient during the two years. There was higher increase in biological yield under EC in HQPM-1 (11.4-28%) as compared to PMH-1 (15.9-17.3%) (p < 0.05) during the two years as compared to the Amb. The increase in grain number cob -1 under elevated CO2 was higher for HQPM-1 (10.1-21.6%) as compared to PMH-1 (5.3-15%) (p < 0.05) over Amb. Under the interactive treatment ECO grain weight cob -1 increased by 12.2% in HQPM-1 as compared to EO alone, whereas there was no significant difference in PMH-1. No significant difference in protein, carbohydrate, P and K content was observed under ECO treatment as compared to Amb in any of the crop cultivars (Fig. 2).

DISCUSSIONS
Present study was carried out under free air field condition to study the responses of two varieties of maize, grown under elevated levels of tropospheric O3 and CO2 individually as well as in combination. Due to low humidity, high light intensity, and high temperature in the Indo-Gangetic Plains (IGP) there are high concentrations of tropospheric O3 along with atmospheric CO2 (IPCC, 2014;Mills et al., 2018) in this region. Leakey et al. (2009) reported that C4 crops under elevated CO2 did not increase the photosynthetic rate directly and only stimulated the uptake of carbon during drought or at midday when short-term water stress developed under high atmospheric vapour pressure deficits and temperatures in temperate climatic conditions. However, our results and also those of Abebe et al. (2016) and Pingale et al. (2017) confirmed that the elevated CO2 increased Pn of maize grown under tropical semi-arid environment. Higher daytime temperatures in tropical conditions, may result in higher vapour pressure deficit resulting in short term water stress, thereby increasing the CO2 uptake even in C4 maize.
EO significantly reduced the photosynthetic rate in both the crop cultivars and that might be due to entering of O3 molecules through stomatal pores during normal gas exchange process that created the reactive oxygen species (ROS), which may lead to programmed cell death (McGrath et al., 2015;Yendrek et al., 2017). No significant change in photosynthetic rate was observed under the interactive treatment (ECO) over the ambient (Amb), which could be due to reduced penetration of higher concentration of CO2 and tropospheric O3 through the stomata (Table 4(a)), because elevated CO2 andO3 both prohibit the opening of stomatal pores of the leaves, that minimize O3 flux in plant apoplast (Oikawa and Ainsworth, 2016). EC and EO conditions significantly reduced the gs in both crop varieties, and it might be attributed to reduced stomatal density and less opening of the stomatal pores (Xu et al., 2016;Singh et al., 2018). In previous research it was documented that gs reduced under CO2 condition in C4 crops (Harley and Baldocchi, 1995;De Souza et al., 2015;Hoshika et al., 2015). However, O3 induced damage may be less for C4 species, given their intrinsically lower stomatal conductance, as well as for plants under rising CO2 (McGrath et al., 2015).
Leaf area index and chlorophyll content of leaves were also significantly impacted under the different treatments however, the cultivar treatment interaction was not significantly different at both the crop stages (Table 4(a)). Adishesha et al. (2017) reported that the chlorophyll content of maize increased under elevated CO2 environment and reduced under the tropospheric O3 concentration (Bhatia et al., 2014). Deterioration in chlorophyll content of leaves under EO might be because of damage to the chloroplast due to change in membrane permeability (Zhu et al., 2011). EC (550 ppm) and EO (70 ppb) treatment significantly (p < 0.01) impacted the crop yield parameters such as cob length, grains (cob -1 ), biological yield and grain yield (Table 4(b)). The cultivar * treatment interaction did not significantly impact crop yield components except cob length and 100 grain weights (Table 4(b)). Higher grain number (cob -1 ) and cob length (cm) of HQPM-1 by 11.5% and 9.7%, respectively in the first year as compared to the second year resulted in higher grain yield in the first year. Improvement in the biological yield and grain yield of cultivars under EC condition was due to the increased photosynthetic rate (12.1-16.6%), cob length (9.3-10.4%), number of grains (cob -1 ) (10.2-15.8%) and test weight. In previous studies it was reported that cob length, grains (cob -1 ), biological yield and 100 grain weights increased significantly under the EC and vice-versa in the EO condition Ainsworth, 2017). Basu et al. (2014) assessed the responses of 23 maize simulation models and concluded that the doubling of CO2 from 360 to 720 ppm would increase grain yield by 7.5% on an average across the sites.  The fumigation of elevated O3 throughout the plant growth, reduced photosynthetic rate, grain weight (cob -1 ), cob length, grains (cob -1 ) and 100 grain weight of crop cultivars and led to reduced grain yield in the EO treatment. It may be attributed to the entry of O3 molecules in plants through leaf stomata and its quick reaction with the molecules of the intercellular space and production of reactive oxygen species (ROS) in the apoplast (Ainsworth, 2017). The produced ROS in apoplast trigger metabolic defense mechanism and causing programme cell death that promote leaf senescence, resulting in diversion of resources during growth and seed production and ultimately influences the crop yield. Feng et al. (2015) and Leisner and Ainsworth (2012) observed that the elevated tropospheric O3 damages the crop during the reproductive stage of growth due to increased abiotic stress sensitivity and high demand of resources for seed development. In our study the physiological maturity was reached earlier by 4 days under the EO treatment in both the crop cultivars, and it was delayed by five days in the elevated CO2 treatment as compared to the ambient and this may have contributed to the decrease or increase in maize yield under EO and EC respectively. Similar to our findings Broberg et al. (2015) and Pleijel et al. (2015) pointed out that the senescence promoted the negative effects of tropospheric O3 on the duration of grain filling, resulting in lower grain mass of wheat and reduced yield.
An increase in grain yield of maize was observed under the interactive treatment ECO over the ambient, implying that 559 ppm of elevated CO2 was able to offset the negative impact of elevated O3 (71 ppb) on the grain yield. The mitigative effect of EC under the combined interactive treatment may be attributed to the reduced stomatal conductance and declined stomatal density which reduced the influx of O3 into the plant leaves (Gray et al., 2000;Ainsworth et al., 2008).
Besides, impacts on growth and grain yield, elevated carbon dioxide (EC) and elevated tropospheric (EO) also impacted quality of grains. In this study phosphorus (P), potassium (K), ). It has been reported in other crops that the concentration of P, K and crude protein of the grains reduced significantly under the elevated CO2 and increased under elevated O3 (Hogy et al., 2009;Zhang et al., 2013;Zhou et al., 2015;Zhu et al., 2018). Mishra et al. (2013) noticed a negative correlation between grain yield and grain protein content due to the growth dilution effect. The average decline in yield over the two years under elevated O3 in both the cultivars was similar, however they responded differently to elevated CO2. The increase in grain yield was more in HQPM1 as compared to PMH-1 under EC treatment. Thus, under the interaction ECO the offsetting of grain yield as compared to decline in yield due to EO was much higher in case of HQPM-1. The increase in grain yield was higher in HQPM-1 under EC due to higher increase in 100 grain weight. The differences obtained in the grain weights (cob -1 ) of crop cultivars under various treatment was due to the number of grains (cob -1 ) and 100 grain weight (g).

CONCLUSIONS
In our study the presence of elevated CO2 increased the carbon uptake and yield of maize, but the increased carbohydrates in the grain led to a decline in the nutrient content (P, K & crude protein) due to the dilution effect and lowered the grain quality. Just the opposite was observed under elevated O3. The elevated CO2 mitigated the adverse impacts of elevated O3 on the maize growth and yield under the interaction treatment. After two years of study we could conclude that elevated CO2 (559 ppm) was able to offset the negative effect of elevated O3 (71 ppb) on growth parameters (4.8-18.3%) and yield (11.2-18.8%) without significantly affecting the grain quality in both the maize cultivars. This amelioration was primarily due to a reduction in stomatal conductance, as a consequence of CO2 elevation. The yield and the grain nutrient content of maize grown under the interaction of elevated CO2 and O3 (ECO) was observed to be at par with the ambient. Therefore, the yield and nutrient content of maize would be the least affected under the future climatic scenario of increasing atmospheric CO2 and O3 concentrations in tropical environments.