Abstract
Effects of Fe-based nanoparticles (Fe2O3 and Fe3O4) on germination parameters were studied for some seedling characteristics of Oriental beech (Fagus orientalis) seeds. Fe2O3 and Fe3O4 nanoparticle applications were made at concentrations of 400, 800, 1200, 1600, and 2000 mg/L on Fagus orientalis seeds collected from 10 different populations, and some germination and seedling characteristics were evaluated. Preliminary results generally indicated that low-dose nanoparticle applications positively affected germination and seedling characteristics, while increases in doses led to decreases in these parameters. Values obtained from high-dose nanoparticle applications were generally lower than those from the control group. The iron nanoparticles affected the parameters to different extents, Fe2O3 nanoparticles showed a significant positive effect on germination rate and radicle length, while exhibiting a significant negative effect on germination percentage and plumule length. The populations least affected by high-dose iron nanoparticle applications were Bursa Inegol, Karabuk-Yenice, and Ordu Akkus, while the most affected were the Bartin-Kumluca and Kahramanmaras-Andirin populations.
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Effects of Adding Fe2O3 and Fe3O4 Nanoparticles to Soil on Germination and Seedling Characteristics of Oriental Beech
Hakan Sevik,a,* Handan Ucun Ozel,b Yafes Yildiz,c and Halil Barış Özel c
Effects of Fe-based nanoparticles (Fe2O3 and Fe3O4) on germination parameters were studied for some seedling characteristics of Oriental beech (Fagus orientalis) seeds. Fe2O3 and Fe3O4 nanoparticle applications were made at concentrations of 400, 800, 1200, 1600, and 2000 mg/L on Fagus orientalis seeds collected from 10 different populations, and some germination and seedling characteristics were evaluated. Preliminary results generally indicated that low-dose nanoparticle applications positively affected germination and seedling characteristics, while increases in doses led to decreases in these parameters. Values obtained from high-dose nanoparticle applications were generally lower than those from the control group. The iron nanoparticles affected the parameters to different extents, Fe2O3 nanoparticles showed a significant positive effect on germination rate and radicle length, while exhibiting a significant negative effect on germination percentage and plumule length. The populations least affected by high-dose iron nanoparticle applications were Bursa Inegol, Karabuk-Yenice, and Ordu Akkus, while the most affected were the Bartin-Kumluca and Kahramanmaras-Andirin populations.
DOI: 10.15376/biores.20.1.70-82
Keywords: Nanoparticle; Fe2O3; Fe3O4; Fagus orientalis; Germination
Contact information: a: Department of Environmental Engineering Faculty of Engineering and Architecture, Kastamonu University, Türkiye; b: Department of Environmental Engineering Faculty of Engineering, Architecture and Design, Bartın University, Türkiye; c: Department of Forest Engineering, Faculty of Forestry, Bartın University, Türkiye; *Corresponding author: hakansevik@gmail.com
INTRODUCTION
Plants are the basis of all living life on Earth. Therefore, understanding and directing plant growth and development is one of the primary study topics for human beings (Ozel et al. 2021; Özdikmenli et al. 2024). As is known, the growth and development of plants, as in all living organisms, are shaped under the influence of genetic structure (Hrivnak et al. 2024) and environmental conditions (Yayla et al. 2022; Sevik et al. 2024). The main environmental factors affecting plant growth are climatic and edaphic (Key et al. 2023; Koc et al. 2024). Edaphic factors include many components, such as soil nutrient content, pH, and soil texture and structure (Erdem et al. 2023).
The chemical structure of the soil is the most important factor affecting and shaping plant growth. The presence and amount of nutrients required for plant growth are essential. However, metals, essential as nutrients, can become lethal to plant biology at high concentrations. Therefore, it is crucial to provide plants with the optimum amounts of metals so that normal metabolic functions can be maintained without metals deficiency or phytotoxicity (Natasha et al. 2022). Moreover, the optimum amount of each metal for plants needs to be determined individually for each plant. The optimum metal concentration and toxicity limit for each plant is different (Pavlovic et al. 2021; Kaur et al. 2023).
Iron is one of the metals absolutely necessary for plant growth. Iron (Fe) is an essential element for plants and plays many important roles in physiological and metabolic processes. It is a redox active, highly reactive element, and thus higher concentrations of Fe can be toxic to plants (Zuo and Zhang 2011; Kuzmina et al. 2023). Although Fe is present in high concentrations in soil, it is not readily taken up by plants. Fe is the third most limiting nutrient for plant growth and metabolism, primarily due to the low solubility of the oxidized ferric form in aerobic environments (Zuo and Zhang 2011). The plant growth depends on the Fe availability that is subjected to several factors including physico-chemical properties of soil and microbial Fe metabolism. Plants secrete protons, phenolics, and metabolites (e.g. mugineic acid) to aid conversion of Fe from Fe3+ to Fe2+ and to increase chelation of Fe for uptake (Ishimaru et al. 2011). The optimum amount of Fe and the level of toxicity have been determined in many cases. For example, for Triticum aestivum, 10 to 50 mg L-1 level is optimum, while 250 mg L-1 level is toxic and 1000 mg L-1 level is lethal (Kaur et al. 2023). In Oryza sativa, Fe levels that cause toxicity range from as low as 10 mg Fe L-1 to 500 mg Fe L-1 or higher (Sahrawat 2005).
In plant development and especially in cultivated plants, the elimination of elemental deficiency in the soil due to long-term production is of great importance in terms of ensuring product continuity. Fertilization for this purpose is vital for plant development (Erdem et al. 2023). Soil fertilization with nanoparticles shows promising results for agriculture, especially for crop production (Mielcarz-Skalska et al. 2021). In recent years, nanoparticles have been used intensively in many fields, not only for fertilization (Özel et al. 2024).
Among the most widely used nanoparticles, Fe2O3 and Fe3O4 nanoparticles are used for the recovery of contaminated water and soil, offering great promise for use in biomedicine and water treatment due to their superparamagnetic and adsorptive properties. Recent studies suggest that Fe-based nanoparticles may also be beneficial for improving plant growth and Fe nutrient accumulation in agricultural plants. However, the potential adverse effects of nanoparticle applications on other organisms remain a serious concern, despite the fact that it generally has positive effect on agricultural plants (Tombuloglu et al. 2022). As a result of the reaction of iron (III) and oxygen, there are about 16 different iron oxide species in nature (Nanography 2024). Iron oxide NPs are often reported in literature using the formula Fe3O4, but they also can have different forms such as magnetite (Fe3O4), maghemite (γ-Fe2O3), hematite (α-Fe2O3), FeO, ε-Fe2O3, and βFe2O3. The different forms possess different magnetic behavior. For instance, magnetite and maghemite are ferromagnetic or superparamagnetic; however, hematites are weakly ferromagnetic or antiferromagnetic (Tombuloglu et al. 2022). For example, magnetic Fe3O4 nanoparticles are superparamagnetic below the size of 20 nm. As the nanoparticle size decreases, this property tends towards paramagnetic or superparamagnetic magnetization (Ajinkya et al. 2020). This magnetic property is very valuable for many applications when produced with controlled size and crystal structure.
Nanoparticle compounds can show different properties from those in nature (Nanography 2024). Nanoparticles can be defined as nano-sized powder grains or particles having dimensions in the range of 1 to 100 nm. Compared to their bulk structures, nanoscale materials exhibit different properties due to quantum size effects, size dependence of the electronic structure, and the number of surface atoms. Nanoparticles have enormous surface energies due to their enormous surface area to volume ratio. In this way, while their chemical and physical properties change, their functionality also changes. Therefore, naturally occurring compounds and nanoparticles are compounds of different character (Seyhan 2022).
Iron nanoparticles can have adverse effects, especially on soil organisms, stimulate abundant ROS (reactive oxygen species) production in plants, cause oxidative stress damage, and thus cause inhibition of plant growth. They can cause serious disruptions in photosynthesis, especially in chlorophyll synthesis, leading to adverse effects on plants (Tao et al. 2023). It has also been determined that cell density decreases dose-dependently (Ameen et al. 2021).
Despite having various studies conducted in agricultural areas, the number of studies on the effects of nanoparticles on forest ecosystems and forest elements is much more limited. In this study, it was aimed to identify the preliminary effects of Fe2O3 and Fe3O4 nanoparticles on the germination parameters of seeds collected from different locations of oriental beech (Fagus orientalis), which is one of the important native tree species in the authors’ country, at different doses.
Iron-based nanoparticles can have different effects in soluble ionic forms, and this effect may differ depending on the plant species. Gui et al. (2015) observed that inhibition of rice root phytohormones under hydroponic conditions was positively correlated with nFe2O3 concentration up to 200 mg/L. However, nFe2O3 was reported to cause oxidative stress in maize and roots of Citrus maxima and also reduced leaf chlorophyll content (Li et al. 2016; Hu et al. 2017). Yang et al. (2020) reported that exposure of soybean to nFe2O3 did not cause any toxicity stress or physiological disorders, on the contrary, exposure to nFe2O3 significantly improved physiological performance by causing increases in chlorophyll content, plant biomass and root growth indices.
However, high amounts of nanoparticles may result in the presence of higher concentrations of iron ions of various types adjacent to the nanoparticles in moist soil due to diffusion mechanism, which may negatively affect plant growth. Therefore, the main hypothesis of the study was “Increasing dosage of Fe-based nanoparticles negatively affects germination and seedling characters in Fagus orientalis seeds”.
EXPERIMENTAL
Materials and Method
The seeds collected within the scope of the study were obtained from the oriental beech forests of Türkiye. Information regarding the regions and populations, from which the seeds were collected, is given in Table 1.
After being subjected to health tests and showing no issues in their embryo and endosperm parts, and demonstrating healthy and normal development performance, the collected seeds were used in germination tests. One of the main objectives of the research was to identify the effects of iron nanoparticles on the germination parameters of oriental beech seeds as well as to establish toxic threshold values. For this purpose, under sterile and hygienic conditions in the laboratory, five different nanoparticle concentrations for both Fe2O3 (average diameter: <30 nm, purity: >99.5%) and Fe3O4 (average diameter: <20 nm, purity: >99.5%) nanoparticles were prepared in the amounts of 400, 800, 1200, 1600, and 2000 mg/L, thus a total of ten concentrations were then stored in sterile concentration bottles to be applied to the seeds.
Table 1. Populations from Which the Seeds Were Collected
Table 1. Populations from Which the Seeds Were Collected
The beech seeds treated with prepared concentrations in five different doses were placed in single-use sterile petri dishes prepared with quantitative filter papers to be used in germination tests. Five repetitions were performed for each dose, and nanoparticle treatment was applied to 150 seeds for each dose, with 30 healthy seeds per repetition. A total of 900 seeds, including the control group, were used in the germination tests. Germination tests were conducted in a 3M Climacell brand germination cabinet. The temperature of the germination medium in the cabinet was set to 20 °C, relative humidity to 70%, and the lighting duration to 12 h.
Oriental beech seeds, placed in 100-mL petri dishes in a way not to touch each other, were monitored for germination by exposing them to 10 mL of nanoparticle solution daily. On the 7th day of application, the number of germinated seeds was counted to calculate the germination rate (GR). The applications continued for 35 days, and at the end of the 35th day, the seedling height (SH), root collar diameter (RCD), plumule length (PL), radicle length (RL), and radicle thickness (RT) were measured using a digital micro-meter compass. Non-germinated seeds were cut to check if they were healthy, and the germination percentage (GP) was calculated by comparing the total number of germinated seeds to the total number of healthy seeds. Similarly, the germination rate was calculated as the ratio of the number of germinated seeds on the 7th day to the number of healthy seeds. The data obtained were analysed using SPSS 22.0 software package, and variance analysis and Duncan test were applied to the data.
RESULTS
Data showing the changes in germination rate is presented in Table 2.
Table 2. Changes in Germination Percentage (%)
Pop.; Population number, Cont.; Control, Av.; Average, StD.; Standard deviation
When the changes in germination percentage were examined, it was observed that in both nanoparticle applications, the germination percentage significantly increased at low doses and began to decrease with increasing doses. The germination at a dose of 400 mg/L increased by approximately 50% in the Fe3O4 nanoparticle application, while it increased more than twice in the Fe2O3 nanoparticle application. Afterwards, the germination started to decrease with increasing doses, but even at the highest dose application of 2000 mg/L, it did not fall below the values in the control group.
Table 3. Changes in Germination Percentage (%)
When the changes in germination percentage was examined based on population, it was observed that the highest germination rates were obtained in the P4 and P9 populations, while the lowest germination rates were obtained in the P7 population. The trend showing the changes in germination percentage is presented in Table 3.
It was observed that the application of Fe nanoparticles significantly affected the germination percentage. In both nanoparticle applications, the germination percentage increased significantly at low doses, with this increase being higher in the Fe3O4 nanoparticle application. However, in both applications, the germination percentage decreased significantly at the application of 1200 mg/L, and it continued to decrease with increasing doses. While the germination percentage in Fe3O4 nanoparticles was generally higher than that in the control group even at high doses, the germination percentages obtained at 2000 mg/L application in Fe2O3 nanoparticles were generally lower than those in the control group.
When the changes in germination percentage were examined based on population, it was observed that the highest germination percentages were obtained in the P4, P8, and P9 populations. The lowest germination percentages were obtained in the P3 and P7 populations. The graph showing the changes in seedling height is presented in Table 4.
Table 4. Changes in Seedling Height (cm)
Similar to germination values, it was determined that Fe nanoparticle applications significantly affected the seedling height as well. In both nanoparticle applications, seedling height increased significantly at low doses, with this increase being higher in the Fe3O4 nanoparticle application. However, seedling height decreased with increasing nanoparticle levels. In the Fe2O3 nanoparticle application, it was determined that the decrease in seedling height with increasing doses was at higher levels.
Seedling height was also observed to be varying significantly on a population basis. The highest seedling heights were observed in the P4, P8, and P9 populations, while the lowest seedling heights were observed in the P3, P7, and P10 populations. The graph showing the changes in root collar diameter is presented in Table 5.
Table 5. Changes in Root Collar Diameter (mm)
As shown in Table 5, the values of root collar diameter generally ranged between 0.8 and 1.0 mm, and it was not possible to say that there was a significant change depending on both the population and the nanoparticles applied. According to these results, it can be said that the root collar diameter did not change noticeably depending on the nanoparticle applications. The graph showing the changes in plumule length is presented in Table 6.
Table 6. Changes in Plumule Length (cm)Formun Üstü
It can be said that the changes in plumule length were not significant in the Fe3O4 application, while they tended to decrease with increasing doses in the Fe2O3 application. It was observed that the plumule length, which was in the range of 0.9 to 1 mm in the control group for the Fe2O3 application, dropped to the level of 0.8 mm in most populations with the application of Fe2O3. The graph showing the changes in radicle thickness is presented in Table 7.
Table 7. Changes in Radicle Thickness (mm)Formun Üstü
As shown in Table 7, the radicle thickness significantly decreased after the application of Fe nanoparticles, when it was 1 mm or more in the control group, and it continued to decrease with increasing doses. This decrease was much faster in the Fe2O3 application. In the Fe2O3 application, from the dose of 1200 mg/L onwards, the average radicle length in most populations is observed to be around 0.8 mm. The graph showing the changes in radicle length is presented in Table 8.
Table 8. Changes in Radicle Length (cm)Formun Üstü