Graphene ameliorates saline-alkaline stress-induced damage and improves growth and tolerance in alfalfa (Medicago sativa L.)

Highlights

• Positive and negative effects of graphene on alfalfa were provided.

• Nano-C (5 g kg⁻¹) can promote photosynthesis, and affect antioxidant system.

• Results of the model analyses showed optimal application was 6.57 g kg⁻¹ Nano-C.

• The findings help to find sustainable nano-enabled plant protection strategies.

Abstract

Nanoscience is leading to the production of nanotech applications to enhance growth and environmental stress tolerance of plants. However, there are few studies on the morphological and physiological regulative mechanism of coupling effects between graphene (Nano-C) and pH in alfalfa, especially both in promotion and phytotoxicity effect by Nano-C under abiotic stress. The coupling effects of Nano-C and pH on plant growth, photosynthetic parameters and enzymes of the antioxidant defence system on leaves and roots were investigated at graphene concentrations of 0, 5, 10, and 20 g kg⁻¹ in pots trials. The result showed a significantly promoted the plant growth at treatment of 5 g kg⁻¹ Nano-C, whereas 10-20 g kg⁻¹ Nano-C harmed the physiological and morphological characterizations. Under salt and alkali stresses, the dry weight, fresh weight, seedling length and antioxidant enzymes activities were increased significantly at 5 g kg⁻¹ Nano-C treatment. These findings revealed that 5 g kg⁻¹ Nano-C enhanced alfalfa tolerance by modulating various morphological and physiological characteristics. Meanwhile, through models analyses, the equivalent coupling effects of Nano-C and pH were located at rectangular area defined by 5.43–7.80 Nano-C combining 8.59–8.96 pH value, and the final optimal application of Nano-C was at 6.57 g kg⁻¹ when high exposure dose of Nano-C indeed resulted in phytotoxicity to the plant. Results of the model analyses also showed significant synergistic effects on the photosynthesis and antioxidant system. Overall, our findings suggested fundamental insights into carbon nanomaterials-mediated physiological adaptation in plants.

Introduction

Soil salinization and alkalization is a major global environmental concern that not only limits plant growth and development, but also contributes to land degradation. For instance, the world's arable land area is 1.5 × 10⁹ ha, but 23% (0.34 × 10⁹ ha) of the area is saline, and 37% (0.56 × 10⁹ ha) is sodic. Neutral salts (NaCl and Na₂SO₄) cause salt stress, whereas alkali stress is caused by alkaline salts (NaHCO₃ and Na₂CO₃) (Yang et al., 2007). Alkaline salt stress is best defined as alkali stress in general, whereas salt stress refers to neutral salt stress only. To survive under those adverse conditions, via a series of dramatic physiological and biochemical changes, plants can use complex defensive mechanisms (Wu et al., 2018). Soil salinity and alkalinity initially restrict plant growth in the form of osmotic stress and later induce ionic stress (Munns and Tester, 2008). During the initial step, cell expansion, cell wall synthesis, protein synthesis and stomatal conductance are all inhibited (Apel and Hirt, 2004). The later step of ion aggregation promotes the generation of reactive oxygen species (ROS). In mitigating oxidative damage, antioxidant enzymes, including catalase (CAT), peroxidase (POD), superoxide dismutase (SOD) and ascorbate peroxidase (APX), are important components of the ROS scavenging method (Wang et al., 2016). Moreover, photosynthetic activity in plant leaves also will be reduced under salt or alkali stress. For instance, salt stress can damage light-capturing pigment complexes on photosystem I and photosystem II (LHC I and LHC II), resulting in the reduced chloroplast's ability to absorb light and transfer electrons, and reduce energy conversion effects, which ultimately results in the reduced photosynthesis' absorption, transmission and conversion of energy efficiency (Mu and Chen, 2020).

The use of nano technology, which can be effective and economical in contrast with traditional methods, is a new age in the field of plant fertilization and environmental remediation (Sabet and Mortazaeinezhad, 2018). Studies have shown that the nano-scale use of nutrients (nano-fertilization) can improve the efficiency of nutrient use and lead to less fertilizer use, increased yield production, and decreased environmental pollution (Chen and Yada, 2011, Husen and Siddiqi, 2014, Khot et al., 2012). A growing number of studies have shown that nanoparticles have positive effects on plants, such as promoting seed germination and plant development (Chen and Yada, 2011), improving nitrogen metabolism and plant photosynthesis (Husen and Siddiqi, 2014), promoting nutrient absorption, and fertilizer utilization rates (Khot et al., 2012). Graphene, a popular two-dimensional carbon-based nanomaterial, has received increasing attention due to its unique physical and structural properties (Avouris, 2010). Low sulfonated graphene concentration may scavenge ROS in the roots, change root morphology and improve the health of maize seedlings (Ren et al., 2016). High graphene concentrations inhibit rice growth, eventually reducing its biomass (Liu et al., 2015). Some studies also summarized the potential key mechanisms of phytotoxicity of graphene family nanomaterials, include physical effects (mechanical damage and physical obstruction) and biochemical and physiological effects (increase in performance of reactive oxygen species (ROS), antioxidant activity generation and inhibition, metabolic disorders, and photosynthesis suppression by decreasing chlorophyll biosynthesis) (Wang et al., 2019). However the influence mechanism of nanomaterials on plants is complicated, as it is not only related to class, concentration and surface characteristics of nanomaterials, but also to types of plants and seeds size (Wu et al., 2012), there are limited studies on the specific application rate in the saline-alkaline soil which are because of different mobility and accessibility to plants.

Alfalfa (Medicago sativa L.) is widely distributed worldwide with high protein, vitamin, and mineral contents and is predominantly located in the northern, northwestern, and northeastern regions of China (Jiang et al., 2019, Peng et al., 2008). However, in those regions, soil salinization is significantly increasing, reducing the productivity and persistence of alfalfa (Munns and Tester, 2008, Yang et al., 2007). Therefore, for the use of these saline areas, breeding and finding salt-alkali tolerant alfalfa varieties is important. Using the relatively salt-tolerant alfalfa Zhongmu NO.1, we analyzed the responses of Zhongmu NO.1 to salt and alkali stresses under graphene (Nano-C) treatment at both in terms of physiological and morphological responses. And, we used the two main factors, Nano-C and pH, to construct the models. The main objectives were (1) to evaluate the effects of non-uniform Nano-C treatments on the growth of alfalfa; (2) to evaluate the effects of Nano-C on the morphological and physiological regulation of the leaves and roots under salt and alkali stresses, and (3) to investigate the mechanisms of coupling effects and develop optimal Nano-C under salt-alkaline stress by constructing suitable models.