بررسی اثر محلول‌پاشی نانوذره‌ی روی و ترکیبات محرک رشد بر صفات فیزیولوژیکی و عملکرد چغندرقند تحت شرایط کم‌آبی

Extended Abstract
 Introduction
Sugar beet is a crucial crop for global sugar production, but its yield is significantly hampered by drought stress, which can lead to a reduction of up to 30% by interfering with physiological and biochemical functions. The use of nano-fertilizers such as zinc oxide nanoparticles (ZnO-NPs) and plant biostimulants like chitosan and proline offers a promising strategy to improve drought resistance in crops. ZnO-NPs can enhance nutrient absorption and boost antioxidant defenses, whereas chitosan serves as an elicitor to reinforce plant defense systems, and proline acts as an essential osmolyte for osmotic balance. We hypothesized that the combined foliar application of ZnO-NPs, chitosan, and proline would synergistically enhance the physiological performance, yield, and sugar concentration of sugar beet under conditions of limited irrigation more effectively than when applied individually.
 
Materials and Methods
A two-year field study (2021-2022 and 2022-2023) was conducted using a split-split plot design arranged in Randomized Complete Blocks with three replications. The main plots were assigned to two irrigation regimes: normal (irrigation after 90 mm evaporation) and deficit irrigation (irrigation after 180 mm evaporation). Sub-plots received foliar applications of ZnO-NPs at three concentrations (0, 2, and 4 mg L⁻¹). The sub-sub plots received four biostimulant combinations: control (water), chitosan (200 mg L⁻¹), proline (150 ppm), and a combination of chitosan + proline. Key parameters assessed included leaf relative water content (RWC), stomatal conductance, leaf proline content, root yield, sugar content, and white sugar yield. Data analysis was performed using SAS software (v. 9.1), and mean comparisons were conducted with the LSD test at a 5% significance level.
 
Results
The experimental findings demonstrated that the imposition of deficit irrigation significantly impaired key physiological and agronomic parameters, leading to a substantial reduction in leaf relative water content (RWC), stomatal conductance, and ultimately, root yield. However, the foliar application of the various treatments, particularly the combined formulations, effectively mitigated these adverse effects. Under drought stress conditions, an analysis of physiological traits revealed that the application of 4 mg L⁻¹ ZnO-NPs alone resulted in the highest recorded leaf proline content (0.46 mg g⁻¹ FW). Furthermore, the binary combination of chitosan and proline under the same stress conditions provided a significant boost, increasing proline content by 4.76% and enhancing stomatal conductance by 13.32% compared to the stressed control plants, indicating an improved osmotic adjustment and gas exchange capacity. Regarding yield and quality parame ters, the most effective single treatment under normal irrigation was 2 mg L⁻¹ ZnO-NPs, which increased root yield by 11.07%. Under the constraint of deficit irrigation, the chitosan and proline combination emerged as particularly potent, significantly elevating root yield by 11.76%, sugar content by 15.67%, and white sugar yield by a remarkable 32.71% compared to the non-treated stressed control. The most compelling evidence of an interactive effect was observed with the triple-combination treatment. The synergistic application of 2 mg L⁻¹ ZnO-NPs+ Chitosan+ Proline was the most effective overall strategy. It produced the highest values across multiple critical metrics: stomatal conductance (16.51 mol m⁻² s⁻¹), root yield (68.77 t ha⁻¹), sugar content (16.34%), and white sugar yield (9.65 t ha⁻¹). This superior performance across the board clearly demonstrates a powerful synergistic interaction among the components, where their combined effect surpasses the sum of their individual contributions, establishing this integrated approach as a highly effective strategy for enhancing sugar beet productivity and resilience under water-deficit conditions.
 
Discussion
The findings of this study robustly confirm our initial hypothesis that the combined foliar application of zinc oxide nanoparticles (ZnO-NPs), chitosan, and proline confers a greater advantage in mitigating drought stress in sugar beet than the application of any single component. The observed synergy suggests that each element contributes a unique, complementary mechanism to a comprehensive defense strategy, leading to the significant improvements in physiological resilience and yield parameters. The enhanced physiological performance can be attributed to a multi-faceted mechanism of action. Firstly, zinc oxide nanoparticles are instrumental in bolstering the plant's biochemical defenses. They are known to enhance the activity of key antioxidant enzymes, thereby reducing the oxidative damage inflicted by reactive oxygen species (ROS) under drought conditions. Concurrently, ZnO-NPs contribute to improved photosynthetic efficiency, potentially by protecting the photosynthetic apparatus and facilitating better light utilization. Secondly, chitosan functions primarily as a potent elicitor, priming the plant's innate defense systems. This priming effect leads to a more robust and rapid response to water scarcity, including improved stomatal regulation. By optimizing stomatal aperture, chitosan helps maintain a critical balance between CO₂ uptake for photosynthesis and water conservation through transpiration. Thirdly, the role of proline is fundamental to osmotic adjustment. As a highly soluble osmolyte, proline accumulates in the cytoplasm, lowering the osmotic potential and helping to maintain turgor pressure within cells. This process is vital for sustaining cell expansion and metabolic activity under dehydrating conditions. Furthermore, proline acts as a molecular chaperone, stabilizing proteins and membranes, and as a potent antioxidant, directly scavenging free radicals to protect cellular structures from oxidative damage. The powerful synergy among these components creates a cascade of benefits. The elicitor activity of chitosan primes the plant's stress response pathways, potentially making it more receptive to the benefits of ZnO-NPs and proline. The ROS-scavenging and photosynthetic support from ZnO-NPs, combined with the osmotic and protective functions of proline, collectively lead to superior plant water status, a more robust antioxidant defense system, and ultimately, a more efficient partitioning of photo-assimilates. This redirected flow of carbohydrates towards the roots as a primary sink is the direct cause of the observed enhancements in both root yield and sugar quality under drought stress. This integrated approach, which leverages multiple physiological pathways, aligns with the growing body of literature advocating for integrated nutrient and biostimulant strategies as a cornerstone of sustainable and climate-resilient agriculture.
 
Conclusion
This study concludes that the combined foliar application of zinc oxide nanoparticles (2 mg L⁻¹), chitosan, and proline is a highly effective agronomic strategy for alleviating the detrimental impacts of drought stress on sugar beet. This treatment combination synergistically improves the plant's physiological resilience, leading to significantly higher root yield and sugar productivity. It is therefore recommended as a practical and sustainable approach for cultivating sugar beet in water-limited environments to ensure economic stability for farmers.
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