Aluminum oxide nanoparticles affect the cell wall structure and lignin composition slightly altering the soybean growth

Highlights

• Al₂O₃ NPs slightly affected the growth of soybean plants.

• Al₂O₃ NPs were internalized by the root tips.

• Root and stem lignification were increased by Al₂O₃ NPs.

• Lignin monomer composition was altered by Al₂O₃ NPs.

• Cell-wall esterified hydroxycinnamic acids were altered by Al₂O₃ NPs.

Abstract

Aluminum oxide (Al₂O₃) nanoparticles (NPs) are among the nanoparticles most used industrially, but their impacts on living organisms are widely unknown. We evaluated the effects of 50–1000 mg L⁻¹ Al₂O₃ NPs on the growth, metabolism of lignin and its monomeric composition in soybean plants. Al₂O₃ NPs did not affect the length of roots and stems. However, at the microscopic level, Al₂O₃ NPs altered the root surface inducing the formation of cracks near to root apexes and damage to the root cap. The results suggest that Al₂O₃ NPs were internalized and accumulated into the cytosol and cell wall of roots, probably interacting with organelles such as mitochondria. At the metabolic level, Al₂O₃ NPs increased soluble and cell wall-bound peroxidase activities in roots and stems but reduced phenylalanine ammonia-lyase activity in stems. Increased lignin contents were also detected in roots and stems. The Al₂O₃ NPs increased the p-hydroxyphenyl monomer levels in stems but reduced them in roots. The total phenolic content increased in roots and stems; cell wall-esterified p-coumaric and ferulic acids increased in roots, while the content of p-coumaric acid decreased in stems. In roots, the content of ionic aluminum (Al⁺³) was extremely low, corresponding to 0.0000252% of the aluminum applied in the nanoparticulate form. This finding suggests that all adverse effects observed were due to the Al₂O₃ NPs only. Altogether, these findings suggest that the structure and properties of the soybean cell wall were altered by the Al₂O₃ NPs, probably to reduce its uptake and phytotoxicity.

Introduction

Nanoparticles (NPs) are materials having one of their dimensions ranging from 1 to 100 nm (Amist et al., 2017; Dayem et al., 2017). They have a high surface to volume ratio, which gives them special properties, such as increased chemical reactivity, conductivity, ductility, toughness, and strength (Amist et al., 2017). These properties have enabled their use in a wide range applications (Singh et al., 2017), especially in industry, medicine, and agriculture (Marchiol, 2018). Due to this, current studies on NPs have been intensified. In fact, about 239,000 papers are found in PubMed website using the term “nanoparticles”, 50% of which have been published in the last 5 years.

With annual world production estimated to reach 100,000 tons in 2020, Al₂O₃ NPs are among the most widely utilized (Asztemborska, 2018). Their unique physicochemical properties allow their application in drug delivery, chemical synthesis, catalysis, energy storage, hydrogen production, rocket propellants, explosives, high performance ceramics, sunscreens, packaging materials, cutting tools, plastics, and others (Martin et al., 2018; Shabnam and Kim, 2018; Yanık and Vardar, 2015). Undoubtedly, the largest employment sector that uses Al₂O₃ NPs is the production of paints and coatings (Asztemborska, 2018). However, all these advantages have a cost, and, unfortunately, Al₂O₃ NPs can reach the soil, water, and air during their production, transport, storage, or disposal (Srikanth et al., 2015). Therefore, studies aiming to investigate their toxicity to living organisms and the environment are imperative.

Whether Al₂O₃ NPs are phytotoxic or not to plants is still a controversial issue, since positive, negative, and negligible effects have all been reported. Factors that may affect the results of toxicity studies are the sizes and concentrations of the NPs, as well as the methods of plant cultivation and treatment (Asztemborska, 2018). Positive effects of Al₂O₃ NPs on growth have been reported in Lemna minor (Juhel et al., 2011) and Glycine max under flooding (Yasmeen et al., 2016). These NPs were able to induce the transcription of several genes involved in the growth and nutrient uptake without affecting the photosynthesis and growth of Arabidopsis thaliana (Jin et al., 2017). In turn, Nicotiana tabacum exposed to Al₂O₃ NPs showed changes in microRNA levels, chlorosis, and had decreased leaf area and root weight (Burklew et al., 2012). Inhibitory effects on growth of Triticum aestivum, Zea mays, Cucumis sativus, Brassica oleracea, and Daucus carota exposed to Al₂O₃ NPs have also been reported (Yang and Watts, 2005; Yanık and Vardar, 2015). Finally, Vigna radiata growth was not affected by Al₂O₃ NPs (Shabnam and Kim 2018).

Plant cell wall is a complex and dynamic structure consisting of many different polysaccharides, aromatic compounds, and proteins (Carpita et al., 2015). Cellulose is the main structural component, whose microfibrils are cross-linked by hemicelluloses. Lignin is a heteropolymer of simple phenolic acids that confers rigidity and mechanical resistance to the plant cell wall (Boerjan et al., 2003; Marchiosi et al., 2020). Hydroxycinnamic acids such as p-coumaric and ferulic acids, produced in the phenylpropanoid pathway, covalently bind lignin to proteins and polysaccharides, and act as connectors between cell wall polymers (de Oliveira et al., 2015). Although apparently rigid, the plant cell wall architecture and composition can be modified in response to several biotic and abiotic stresses (Loix et al., 2017; Parrotta et al., 2015). For instance, increased lignification occurs in response to low temperatures, water deficit, salt stress, light, mineral nutrition, and heavy metals (Moura et al., 2010). It has recently been shown that increased lignin deposition produces a physical barrier able to restrict the uptake of heavy metals such as cadmium and, later, its phytotoxicity (Loix et al., 2017; Parrotta et al., 2015). Similarly, the involvement of lignification in plant responses to NPs needs to be investigated.

Some works have revealed increased lignification in a variety of plant species exposed to silica, silver, iron oxide, copper oxide, and zinc oxide NPs (Asgari et al., 2018; Bernard et al., 2015; Lopes et al., 2018; Nazaralian et al., 2017; Prakash and Chung, 2016). However, studies focusing on lignification responses to Al₂O₃ NPs exposure are scarce. In this work, we evaluated the effects of Al₂O₃ NPs on the growth and lignin metabolism and composition of soybean plants. Impacts of Al₂O₃ NPs on the morphology and ultrastructure of roots were analyzed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The contents of lignin and its monomeric composition, total phenolics, cell-wall-esterified hydroxycinnamic acids, and activities of soluble and cell wall-bound peroxidases (POD) in addition to phenylalanine ammonia-lyase (PAL) were determined to evaluate the effects at structural and metabolic levels.