Abstract

Reducing the leaf senescence rate could improve the grain yield in wheat. In the present study, responses of photosynthesis, stomata conductance, SPAD, and yield traits of winter wheat. The results indicated from different nitrogen rate, and variety YH- 20410 significantly affected the yield, and grain quality. The P_N, C_i and E of flag leaves during grain filling, and decreased g_s of flag leaves. Nitrogen application significantly increased the P_N, C_i and E significantly decreased the g_s of post-anthesis flag leaves with 210 kg ha⁻¹ was the best, and SPAD was the best N280. Nitrogen application amount 210 kg ha^–1 combined to variety YH-20410 increased the soluble sugar content in grains at the filling stage. The difference between nitrogen application was significant on 1000-grain mass, Spike number, Grain number per spike, leaf area index nitrogen application rate of 210 kg ha^–1, and variety YH-20410 reached the highest at anthesis and maturity stage, promote the transpiration of flag leaves, intercellular CO₂, yield were the highest under N210 compared to others nitrogen rate. The yield, and three factors were the highest in N210 kg ha⁻¹, and enhances Photosynthetic Characteristics of flag leaves, the spike number, and yield had significant difference compared with other N application rates.

1. Introduction

Winter wheat (Triticum aestivum L.) is one of the most important food crops in the world. It is an important food, and accounting for about one-fifth of food production in China. Nevertheless, the yields in this area is often unstable owing to limited and uneven precipitation distribution in fallow and growing season ¹. Wheat are the most important food crops in Asia, providing food grains for more than 20% of the population worldwide ^{2, 3}. With the rising atmospheric CO₂ concentrations, many scholars pay extensive attention to the ‘CO₂ fertilization effect, the promotion of plant growth or productivity by the increase of atmospheric CO₂ concentrations, in the hope that it will compensate for the crop yield reduction caused by climate change to a certain extent ³. Under drought conditions, stomata are closed, and the CO₂ uptake is reduced, which affects the rate of photosynthesis and reduces growth and yield ⁴. The high density plant populations also decrease the photosynthetic efficiency and starch accumulation. Leaf senescence involves a reduction in the leaf area at the bottom layer and eventually the whole plant ^{5, 6}. Environmental factors (nutrient stress, water, and temperature) regulate leaf senescence in crops ⁷. The green leaf area, and duration have significant effects on the grain yield, and shaded leaves become senescent earlier compared to unshaded leaves ⁸. Accelerated leaf senescence leads to a lower grain yield in maize because of nutritional and hormonal signals ⁹. It was reported that N fertilizer SPAD significantly reduced the content of P_N, g_s, and E, as well as increasing the intercellular carbon dioxide concentration (C_i) in wheat cultivars. Nitrogen (N) deposition has dramatically altered terrestrial ecosystem properties and processes, such as plant nutrient cycling, photosynthetic carbon assimilation, and species diversity ^{10, 11}. Only a small fraction (10%) of these studies quantify the relationship between the leaf determined in vitro, and the SPAD readings. The studies that do perform such calibrations of the SPAD meter usually parameterize linear relationships ^{12, 13, 14}. Under nitrogen, the grain-filling process is sustained mainly by photosynthesis in the upper parts of wheat plants, such as the flag leaves and wheat ears. Short-term exposure to high CO₂ caused increased CO₂ assimilation. It has been reported in several studies that the decrease in g_s caused a more pronounced drop in E ¹⁵. The study it was resulted a much higher WUE in plants grown at high CO₂ than in those grown at ambient CO₂ concentrations. Nitrogen fertilizer is a major component of proteins and nucleic acids, and application regulates plant growth and development ^{15, 16}. Insufficient or Deficient N inhibits the photosynthetic and radiation use efficiencies to reduce the grain yield, whereas the optimum N application concentration can increased the grain yield ¹⁷. In a previous study reported that N enhanced the content, net photosynthetic rate (P_N), stomatal conductance (g_s), transpiration rate (E), and stomatal limitation value of wheat in different plant populations. It was also indicated that N also improved the yield and yield components in wheat ¹⁸. Thus, reducing accelerated leaf senescence in dense plant populations is important for increasing the production of wheat.

Nitrogen is important for enhancing the photosynthetic efficiency and leaf area ¹⁹. In different previous studies, researchers thought that photosynthesis of the flag leaves was supplying the majority of assimilates for grain filling. Now it is accepted that nitrogen ear photosynthesis is a major contributor to the final grain yield. It was proposed that high photosynthesis in wheat leaves is an important trait for nitrogen assimilation in wheat ²⁰. Fertilizers constitute an integral part for improved crop production technology. Nitrogen (N) is an essential mineral nutrient for plant growth, expands soil fertility and crop productivity proper amount of fertilizer application is considered a key for high crop production ²¹.

The objective of this study was to reveal the combined effects of reduced N rate N0, N210, and 280 kg N ha⁻¹, and increased variety on: (1) protein content and yield; (2) soluble sugar, sucrose, starch; and (3) SPAD nitrogen application rate of N280 kg ha^–1 was more beneficial. The results would provide a novel view for achieving the balance between 210 kg ha^–1 was beneficial to increase the content of gluten in grains, obtain higher protein content and protein yield in grains, and increase the content of dry gluten in grains.

2. Materials and Methods

The experiment was conducted in the maize experimental base of Shanxi agricultural in Shanxi Province, China (E112°34 'E, N37°25' N). The pond was planted in a pool 2 m deep, separated by concrete walls with a thickness of 20 cm, and an insulation layer of 10 cm added to the outer walls.

The experiment was conducted in the wheat experimental base of Shanxi agricultural university in Taigu, Shanxi Province, China (E112°34 'E, N37°25' N) from 2018 to 2019, which belonging to the temperate continental climate zone, with an average annual temperature of 10.4 °C. The pond was planted in a pool 2 m deep, separated by concrete walls with a thickness of 20 cm, and an insulation layer of 10 cm added to the outer walls. The experiment had a split-split-plot design with three replications. Three nitrogen treatments were 0, 210 and 280 kg N ha^–1 (N0, N210, and N280 respectively), Winter wheat (Triticum aestivum L.), cultivar ‘Yunhan 20410’‘Yunhan 618’ Before sowing, Pure P₂O₅ and K₂O were applied at the rate of 150 and 75 kg ha⁻¹ respectively, the nitrogen fertilizer was applied to the base fertilizer. The seeds were sown on November, 9, 2018, planted in manual with row spacing of 20cm and sowing quantity of 225 kg ha⁻¹. Irrigation was applied using a drip irrigation system, with application of 50 mm each time as measured with a water meter. All plants were harvested in June 1, 2019. The greenhouse was kept free from insects, pests, and diseases using pesticides as needed. During whole growing season, weed was well controlled by hand.

Twenty random plants with uniform growth were uprooted from the field and the plant height was measured by taking vertical distance from the rhizome to the top of the main stem. At the same time the length, width, and number of green leaves were measured and leaf area index (LAI) was calculated from following formula ²².

At maturity, 20 plants from each plot were randomly sampled from the inner rows for the determination of yield components such as ear number, seed number per ear and weight of thousand seed. Plot grain yield was determined by harvesting all plants in the area of 20m², shelled using machine and the grain was air-dried for the determination of grain yield.

The bromophenol blue water solution and isopropanol lactic acid mixture, and the settling values were determined by shock. The landing value was measured using the Landing Numerical Measurer (FN-IV). The Micro dough LAB, a micro powder instrument was produced by a [Swedish company Botone (SCB)] and it was measured the fluidity of bread. The wet gluten content and gluten index were measured using the Gluten Index Meter (MJZ-II) Mian Jin zhi–2 gluten index analyzer china) quality analyzer. For Quality analysis dough mixed from 200g flour was divided into small dough weighted based on 0.25g flour calculated as by using the following formula ²².

The dry gluten was obtained by drying the wet gluten in an oven (TD5G, Hunan Xiang Li Scientific instruments com., Lt China) to constant weight at 100°C for 24h using the air oven drying method. The dried gluten was left cool for 1 h before taking its weight as the dry gluten content. The percentage of dry gluten obtund was calculated using following formula ²².

The net photosynthetic rate (P_N), transpiration rate (E), intercellular CO₂ concentration (C_i), and stomatal conductance (g_s) of the flag leaf were measured by CI-340 hand-held photosynthesis measurement system (LI-COR, Inc., Lincoln, Nebraska, USA) from 9:00 to 11:00 h. In order to compare photosynthetic characteristics of leaves with a similar developmental age, gas-exchange measurements of each cultivar were conducted immediately after the flag leaves fully expanded. The measurements of P_N, G_s, C_i, and E, were acquired in a system with a leaf chamber temperature of 25°C and the carbon dioxide concentration in the leaf chamber was kept at 380 µmol (CO₂) mol⁻¹ by using a CO₂ injector with a high-pressure liquid CO₂ cartridge source. The photosynthetic active radiation was set at 1,500 µmol (photon) m⁻² s⁻¹. The external humidity was noted as 40 –50% ¹. The ear surface area = 3.8 × [(front width + side width) (ear length)] /2.

The SPAD-502 measurements were conducted in the field between 10:00 and 16:00 h. The adaxial side of the leaves was always placed toward the emitting window of the instrument and major veins were avoided. Leaves at different development stages were selected, including premature, fully mature and senescent leaves. Leaves were detached and collected immediately after the SPAD-502 Tester measurements and were kept cool (– 0C) until sub-samples were punched in the laboratory. The punched leaf material was weighted and then immediately frozen and kept at –25C until chlorophyll extraction. The area of the punched wheat leaf material was determined using an area meter (Delta-T Devices Ltd., Cambridge, England), while two circular 1.0 cm diameter leaf discs from one side on the midrib were punched for birch and potato. The spatial distribution of punched leaf material matched the distribution of the SPAD-502.

After flowering period listed growth consistent and the same day flowering of wheat spike and peeling grain was placed in the oven dried at 105°C for 20 min and then at 80°C for 12 hours for dry weight. The quality and speed of weighing samples greatly affect the overall quality of the test. Then the grain was weighed and phenol method was used to determine the content of sucrose, and ketone color method was used to determine the total soluble total sugar content. H₂SO₄ -H₂O₂ -Phenol blue color method was used to determine the seed protein and component content ²³.

Excel 2010 was used to analyze all the experimental data Analyze of variance (ANOVA) was proceed using DPS7.05 software. SD method was used for multiple comparison (α=0.05).

3. Results

The P_N increased with the amount of nitrogen time after anthesis showing a single peak increase trend Figure 3A. The 14_D P_N time after anthesis was still the highest at variety YH-20410, N280 kg ha⁻¹, and not significantly different from 0 kg ha⁻¹ variety YH-618. Time after anthesis P_N increased with N application. time after anthesis 5_D was still the highest with 210 kg ha⁻¹, but the difference was not significant compared to 0 kg ha⁻¹, variety YH-618 15_D.The addition of variety YH-20410 can significantly improve the P_N of the flag leaves time after anthesis, but the treatment effects of high N, 210 kg ha⁻¹, 280 kg ha⁻¹ in the later grouting was weakened, and 280 kg ha⁻¹ could sustain the whole growth stage.

The E of flag leaves decreased gradually with the grouting process, and the E of flag leaves increased Figure 3B. The 210 kg ha⁻¹, variety YH-20410 treatments were significantly higher than other treatments on 15_D and 10_D, time after anthesis and 0 kg ha⁻¹ was the lowest variety YH-618. After 15_D 280 kg ha⁻¹ was significantly higher than other treatments variety YH-20410, and 0 kg ha⁻¹ was the lowest. The increased N fertilizer can significantly enhance the E of flag leaves after flowering, which could last for the whole growth stage, and the best effect of 280 kg ha⁻¹ was obtained in the large.

The C_i in the flag leaves of wheat decreased gradually with the growing process, and C_i in the flag leaves decreased first and then increased with the increase of N fertilizer, variety YH-20410 different stages time after anthesis Figure 3C. The N level 280 kg ha⁻¹ treatment was significantly lower than other treatments on 10_D, and 15_D after flowering and 0 kg ha⁻¹ was the highest variety YH-20410, N210 kg ha⁻¹, and was the lowest 5_D variety YH-618 time after anthesis, but the difference was not significant compared to variety. The increase of nitrogen fertilizer can significantly reduce the C_i in the leaves of the flags time after anthesis.

The g_s of the flag leaves increased first of N fertilizer, variety YH-20410 at different stages time after anthesis Figure 3D. The variety YH-20410, N210 kg ha⁻¹ treatment was significantly higher than other treatments 15_D time after anthesis, and variety YH-618 N0 kg ha⁻¹ it was the lowest, and N280 kg ha⁻¹ was the highest at 5_D, but the difference was not significant N0 kg ha⁻¹. The increased N can significantly increase g_s of flower flag leaves, but the effect lasts until the middle stage of growth, and the effect weakened in the later stage.

The nitrogen rate SPAD value of flag leaf at after heading stage, SPAD value of flag leaf variety YH-20410 of wheat were increased N210 kg ha^–1, and N280 kg ha^–1 fertilizer compared to variety YH-20410, and the difference was significant when the amount of nitrogen fertilizer was 280 kg ha^–1 (Figure 4). The increase of nitrogen application, SPAD value of flag leaf increased at flowering stage, but the difference was not significant, and SPAD value of flag leaf lowest at N rate of N0 kg ha^–1 variety YH-618.

Effect of nitrogen fertilizer on the content anthesis, and maturity stage. The increase of nitrogen application rate, the leaf area index of wheat at flowering stage gradually increased after flowering, and increased first and then decreased at maturity stage (Figure 5). The leaf area index of wheat at flowering stage was the highest at 280 kg ha^–1, and the lowest at N0 kg ha^–1.The leaf area of wheat at mature stage was the highest at N rate of 210 kg ha^–1, and the lowest at N rate of N0 kg ha^–1 variety YH-618. The leaf area of wheat at flowering stage increased with the increase of N rate, and the highest variety YH-20410 However, there was no significant difference among treatments variety YH-618. In conclusion, nitrogen application rate of 210 kg ha^–1, and variety YH-20410 reached the highest at anthesis and maturity stage, which was beneficial to improve leaf area index at anthesis, and maturity stage and enhance photosynthetic capacity of wheat.

The soluble sugar of grains was the highest in the 280 kg ha^–1 treatment, and the lowest in the N0 kg ha^–1 treatment, (Table 1). However, there was no significant difference in grain soluble sugar between YH-618, and N0, but there was significant difference between them under 280 kg ha^–1 nitrogen application. The soluble sugar content of seeds in N280 kg ha^–1 nitrogen fertilizer, and 2.4 kg ha^–1 variety YH-20410 was the highest, and significantly higher than that in other treatments. There was N0 significant difference in grain sucrose between 210 kg ha^–1 treatments, but it was significantly higher than N0 kg ha^–1, and there was no significant difference between YH-618 treatments. The sucrose content of grain at maturity was significantly higher in high N treatment 210 kg ha^–1 combined to YH-618 than in low N treatment N0 kg ha^–1. The grain starch content at maturity stage was the highest at 210 kg ha^–1 and the lowest at N0 kg ha^–1. There was no significant difference between N0, and the 280 kg ha^–1 combined to variety YH-20410 of reached the highest. But no significant effect on starch content of grain at maturity stage. The effects of variety YH-618 on soluble sugar, sucrose and starch contents of mature grains were not significant. There was a significant interaction between the content of sucrose, and starch in the mature stage.

The Variety compared to nitrogen application rate 210 kg ha^–1 increased the number of ears by 16%-31%, the number of grains per ears by 5%-6%, the 1000-grain weight by 5%-9%, and the yield by 21%-40%, and the number of ears and yield reached significant levels (Table 2). Variety YH-20410 significantly increased the grain number per spike and yield by 4%-16% respectively. There was no significant N0 and Variety YH-618 grain number per spike, and 1000-grain weight. Variety YH-20410 significantly affected 1000-grain weight and yield. The interaction of nitrogen, and variety YH-20410 significantly affected grain number per spike, 1000-grain weight and yield. In conclusion, the nitrogen application rate 210 kg ha^–1 and N280 Variety YH-20410 significantly increased the number of per spike area, which was beneficial to the construction of a reasonable population, and thus increased the grain yield. Moreover, the interaction of nitrogen, and Variety YH-20410 could achieve high yield by regulating the number of panicles and 1000-grain weight

The grain protein content, and grain protein yield of variety YH-20410 were significantly higher than those of N210. The grain protein content and grain protein yield of N280 kg ha^–1 combined to variety YH-618 were the highest, and significantly higher than those of other treatments (Table 3). There was no significant difference in the ratio of gluten to alcohol among nitrogen treatments, but 2.4 kg ha^–1 variety YH-20410 was significantly higher than YH-618. N0 compared to variety YH-618 had no significant effect on grain albumin content. Variety YH-618 had no significant effect on globulin content in grains. Grain gliadin in N210 kg ha^–1 and N150 kg ha^–1 treatments was significantly higher than that in N0 kg ha^–1 treatments. The grain gliadin content of variety YH-20410 was significantly higher than that of YH-618, but there was no significant difference between them only under N280 kg ha^–1 nitrogen application. The glutenin content in variety YH-20410 was significantly higher than that in variety YH-618. The glutenin content of 210 kg ha^–1 nitrogen fertilizer combined to YH-20410 was the highest, and significantly higher than that of other treatments. Analysis of variance showed that nitrogen treatment had extremely significant effects on grain albumin, globulin, glutenin, glutento ratio, grain protein content and protein yield. The contents of gliadin and gluten in grains were significantly affected by variety YH-20410 treatment. There were significant interaction between grain gliadin, glutenin, protein content, and protein yield among treatments N210, and variety YH-20410.

4. Discussions

The nitrogen rate compared to Variety, and SPAD the leaf area index of winter wheat increased under N280 especially in the middle, and late growth stage. The leaf area under variety YH-20410 were also higher than the variety YH-618. The differences in leaf area under the different planting method could be attributed to the differences in the wheat distribution. ²⁴. Wheat with variety YH-20410 intercept light more properly with higher photosynthesis rate and leaf area index. The optimize doses of nitrogen to increase the yield of winter wheat crop. Thus, we used three nitrogen application concentrations set as N0, N210, N280, respectively to analyze the photosynthetic characteristics and yield-related traits in winter wheat. Therefore, the SPAD with N280 kg ha⁻¹ would be a valuable management practice to improve wheat ^{1, 25}. The use of the SPAD chlorophyll meter has increased dramatically within agriculture and research during the last decade. The studies that do perform such calibrations of the SPAD meter usually parameterize linear relationships ²⁶. Enhanced g_s of the three species under increased N availability improved the photosynthetic capacity by increasing CO₂ supply to the intercellular spaces ²⁷. The results of this study showed that the leaf area index was increased until bolting stage and then decreased. The reduction in leaf area index at later growth stage is due to leaf senescence ²⁸. Reported that the photosynthesis rate were higher under SPAD, and photochemical efficiency and chlorophyll contents at the heading and anthesis stages. The better plant distribution effectively intercept radiation N, and improves photosynthesis efficiency and growth. The results of this study showed that the Photosynthetic active radiation was found positively related with the leaf area index and number of spikes ²⁹.

The optimum nitrogen application rates varies with the increasing production, and effect of higher nitrogen fertilizer application rate proved better because fertility has improved the plant growth by promoting the absorption and transportation of fertilizer by the root system ³⁰. Wheat grain protein can be categorized as albumins, globulins, gliadins, and glutenins according to their solubility properties. Albumins and globulins are soluble proteins comprising various variety, and inhibitors, which have crucial structural, and metabolic functions during grain-filling ³¹. Gliadins and glutenins, called gluten proteins, interact to form a viscoelastic gluten network in dough, which are essential in determining the baking quality of wheat flour products ³². Previous studies showed that both grain protein content GPC and grain yield (GY) can be increased by N fertilizer simultaneously under low to medium N rate (0–200 kg N ha^–1), but grain protein content (GPC) was continuously increased with no further increase in GY under high N rate (210–280 kg N ha^–1) ³³. In China, the N application rate has been generally much higher than the recommended dose of 280 kg N ha^{– 1} in soft wheat production for higher grain yield GY. Reducing N rate can significantly decrease GPC and wet gluten content (WGC) of soft wheat, which help to form softer dough and produce crisper biscuits. N agronomic efficiency (NAE) is also enhanced by reducing N input. However, reducing N rate also increased the risk of GY loss mainly owing to reductions in both number of spikes and grains ^{34, 35}. Thus, it is very important to develop agronomic practices to balance grain quality and yield in soft wheat production.

The North China Plain showed that, compared to conventional farmer practice (210 kg N ha⁻¹), sensor-based N management strategy (280 kg N ha⁻¹) decreased residual soil mineral-N content after harvest on average by 44 % (across 2 years). Furthermore, N was considerably greater (by 368 %) for the sensor-based fertilizer recommendation than for common farmer practice. Also this strategy produced comparable grain yields ³⁶. The variable-rate fertilization (based on the reflected spectrum from wheat) reduced the variation of yield, ear numbers and dry biomass, but it did not increase crop yield and grain protein content significantly. An improved fertilizer N with comparable yield was also achieved in irrigated wheat ³⁷. More specifically, this strategy consisted of applying moderate prescriptive dose of fertilizer N at planting and crown root initiation stages, and a corrective sensor-guided application at two stages corresponding to 2nd or 3rd irrigation events ³⁸. Also showed that winter wheat N improved when mid-season fertilization was based on optically sensed in-season estimates of grain yield. Nitrogen rate, in fact, increased by more than 15 % compared with the Variety and N rate of 210 kg N ha⁻¹. As N increases, generally the ability of site-specific fertilization to maintain profitability with lower average N applications is expected to be improved ³⁹. The N increased in about 53 % of the grids and enabled the design of N prescriptions adapted to plants demand. In different studies, lower N doses were applied on winter wheat based on crop reflection methods, producing the best efficiency in terms of grain production (as highest ratio: yield/applied N) and grain yields equivalent to the current standard method ⁴⁰. These results were obtained despite differences between N treatments were always significant ^{41, 42}. Found that a large reduction in N inputs (up to 48.6 %) was due to in-season system to evaluate the crop and optimize N rates compared to the practices normally applied by farmers. A further reduction (up to 19.6 %) was possible through site-specific application. This method maximized spring N fertilizer use efficiency and reduced within-field grain yield variance, compared with greenhouse-specific management. Similarly, in a field trial in the UK, the application of N by using sensors saved 15 kg N ha⁻¹ without a negative influence on yield, which increased the N. In addition, there were potential environmental benefits through a 52 % reduction of the residual N in the soil ^{43, 44, 45}. From all the above, it can be summarized that greenhouse studies in which sensor-based N management systems were compared to common farmer practices have indicated significant increases in the N.

5. Conclusion

The Nitrogen application amount 210 kg ha−1 enhances the photosynthetic characteristics of flag leaves and promotes high yield. SPAD nitrogen application rate of N280 kg ha–1 was more beneficial to the different growth stages of wheat (Triticum aestivum L.). Nitrogen application amount 210 kg ha^–1 combined to variety YH-20410 increased the soluble sugar content in grains at the filling stage, which was conducive to the accumulation of sucrose, and starch in grains 210 kg ha^–1 was beneficial to increase the content of wheat gluten in grains, obtain higher protein content and yield in grains, and increase the content of dry gluten in grains.

Acknowledgements

This project was approved by the Shanxi Collaboration and Innovation Centre for Efficient Production of Specialty Crops in Loess Plateau”. The authors are thankful to China Agriculture Research System (No. CARS-03-01-24). The Central Government guides local science and Technology Development Fund projects (No.YDZJSX2021C016). General Project of SCO Institute of Modern Agricultural Development (No.SC021B004). State Key Laboratory of Sustainable Dryland Agriculture, Shanxi Agricultural University (No.202003-2). The Basic Research Program Project of Shanxi Province (20210302123410). The technology innovation team of Shanxi Province (No.201605D131041). The & quot; 1331 & quot; Engineering Key Laboratory of Shanxi Province for financial support of this study.

References