EFFECTS OF TIO₂ NANO-PRIMING ON TOMATO SEED GERMINATION AND PLANT DEVELOPMENT

A. K. Yagız¹* and M. E. Çalışkan¹

¹Nigde Omer Halisdemir University Faculty of Agricultural Sciences and Technologies Agricultural Genetic Engineering Department Nigde, Turkey

Corresponding author’s e-mail: ayagiz@ohu.edu.tr

The effect of nano-TiO₂ on seed germination and plant development as a priming agent has not been thoroughly studied. This study aimed to evaluate the effect of TiO₂ nano-priming on seed germination, plant growth and yield of tomato plants relying on experiments that have been conducted both under laboratory and greenhouse conditions. During the laboratory experiments, the seed germination, seed vigor index and water uptake of seeds were determined after 5, 10, 50 and 100 mg L^-1 of TiO₂ nano-priming while hydropriming was used as control. The biomass of tomato seedling was increased the most for the 10 mg L^-1 TiO₂ nano-priming condition. With follow-up experiments, the effect of 10 mg L^-1 TiO₂ nano-priming was investigated further under greenhouse conditions, where hydropriming was also used as control. The physiological traits of tomato plants, like chlorophyll content, stomatal conductance and transpiration rate were increased by TiO₂ nano-priming treatment. Although the photosynthesis rate was boosted by nano-priming, the yield was not affected. The application of 10 mg L^-1 TiO₂ as nano-priming agent increased plant development and chlorophyll content under both laboratory and greenhouse conditions without translocation in the plant, which is one of the most important concerts of using nanoparticles in plant production.

Seed germination has a vital role in crop production. Even if the seed germination is high, the homogeneous seed germination is irrefutable in plant cultivation. Moreover, the synchronization of plant development enables further treatments in agricultural production (Azimi et al., 2014; Khalaki et al., 2021). For these purposes, priming was developed as a useful technique and currently, there are various types of priming (Finch-Savage et al., 2004; Moammeri et al., 2018; Bekhrad et al., 2016). One of these techniques, nano-priming, is described simply as the soaking of seeds in nanomaterial suspension (Khalaki et al., 2021). Considering that the introduction of nanoparticles (NP) to agriculture does not go back a long way, the ethical and environmental issues about their use in agriculture is still under discussion. This especially relates to the uncertainties regarding NPs, such as their environmental effect as well as durability in nature. Studies have established the toxic effects of NPs, such as prohibition of seed germination, stem and root growth, decrease in the seedling numbers, and reduction of the yield (El-Temsah and Joner, 2010; Li et al., 2015). The toxicity of a NP depends on elemental composition, porosity, average size, surface area, surface charge, aggregation propensity, and hydrodynamic diameter (Peralta-Videa et al., 2011). However, the positive effects of the nano-priming in the plant production are undeniable. The treatment of NPs can increase the seed germination rate, seedling development, growth and yield in different plant types (Dutta, 2018; Feizi et al., 2013).

The combination of these two factors makes using nanoparticles as a priming agent attractive. Considering that a major concern is the uncontrolled release of NPs into nature, the advantages of using NPs as priming agent allows for preventing the spread of NPs to the environment. Furthermore, NPs both increase the solubility and decrease the premature degradation of active ingredients and can be designed of target focusing particles. For this purpose, two strategies, which are both not yet entirely understood, might be followed: developing the effect of active ingredients or introducing new ingredients (Kah et al., 2013).

NPs have unique properties in comparison to their bulk structure. Plant growth, resistance to abiotic stresses, photosynthetic rate and seed germination was increased after subjected to nano-size metals and their oxides such as TiO₂, SiO₂, carbon nanotubes (CNT), ZnO (Almutairi, 2016; Almutairi and Amjad, 2015; Dhoke et al., 2013; Gao et al., 2006). Many different studies have shown that TiO₂ increases the photosynthesis ratio and chlorophyll content of plants (Gao et al., 2006, 2008; Hong et al., 2005; Mahmoodzadeh, 2013). Hong et al (2005) pointed out that, TiO₂ raised the photosynthesis ratio of spinach by promoting photochemical reactions of chloroplast (Hong et al., 2005). Formerly, the studies performed by Gao et al. in 2006 and 2008 illuminated that TiO₂ caused a conformational change in rubisco activase enzyme, which increased the activity of the enzyme (Gao et al., 2006, 2008). TiO₂ nanoparticles were also used under N-deficient conditions and managed nitrogen deficiency in spinach by increasing the nitrate reductase activity (Yang et al., 2006, 2007).

In the presented study TiO₂ was used as the priming agent for tomato (Solanum lycopersicum L.) seeds with the aim to assess any change in plant development and growth. Therefore, tomato seeds were primed with TiO₂ nanoparticles at different concentrations and the effect of TiO₂ nano-priming on tomato seeds and plantlets’ development was investigated. Later on, the optimal concentration was selected and the experiments were continued under the greenhouse condition. The physiological responses and changes in the yield of tomato plants were observed.

Characterization of TiO₂ nanoparticles: The phase analysis of the TiO₂ nano powder was conducted with X-ray diffractometer (XRD) between 2θ: 20-80° with 2°/min speed and Cu-Kα radiation. The structure of the TiO₂ NPs was observed using a field emission scanning electron microscope (SEM) at the accelerating voltage of 5.0 kV.

Effect of TiO₂ on tomato seed germination and seedling growth: For nano-priming treatments, 0 (hydropriming as control), 5, 10, 50 and 100 mg L^-1 of TiO₂ nanoparticle suspensions were prepared using distilled water and sterilized in the autoclave at 121°C for 20 minutes. After sterilization the suspensions were incubated in the sonicator for 30 minutes for homogenization.

Experiments were laid out using a Completely Randomized Design with five replications, and each replication was conducted with 30 seeds. Tomato seeds (S. lycopersicum L. cv. Altar F1) were surface sterilized with 0.7% NaClO for 4 minutes and rinsed with sterile distilled water. The seeds were then soaked in TiO₂ suspensions for 24h at 120 rpm for priming. The seeds were germinated on quarter-strength Murashige and Skoog (MS) media supplied with 1.5% sucrose (Duchefa) and 8% phytoagar (Duchefa). The germination experiment was conducted in germination cabinets at 20°C under 16/8 h light/dark conditions.

On the 21^st day of germination, the germination ratio, length and biomass of the shoot and root were measured. Seed vigor index (SVI) was calculated according to the following formulas:

Seed Vigor Index I= Germination % x Seedling Length (mm)

Seed Vigor Index I= Germination % x Seedling Weight (g)

Water uptake (WU) of seeds was measured with five replications. The seeds were weighed and priming was conducted. After priming, seeds were blotted dry and weighed again and WU was calculated according to the following formula:

WU%= ((SW after priming-SW before priming)/SW before priming) * 100

SW: Seed weight

Effect of TiO₂ on tomato plant development and yield: According to the laboratory experiment, 10 mg L^-1 of TiO₂ was selected for further experiments due to having the highest SVI and hydropriming (0 mg L^-1) was used as a control. Experiments were laid out in Completely Randomized Design with 5 replications. Each experimental unit comprised three pots (12 L, 26 cm in diameter) with one plant in each pot. Before sowing the seeds, the seed nano-priming was conducted with the same method. The pots were filled with peat and perlite mixture in 2:1 ratio (v/v). A composed fertilizer of NPK (18+18+18+ microelements) was incorporated into the peat-perlite mixture before sowing. Three seeds were sown at 3 cm in each pot and thinned to one plant in each pot after emergence. The stem length of the tomato plants was measured at the flowering stage.

Gas exchange, photosynthesis rate and chlorophyll content measurements: Photosynthetic rate (Pn), transpiration rate (E) and stomatal conductance (Gs) were measured on the fully expanded youngest leaves using a portable photosynthesis system (LI-6400 XT, LICOR, USA). The measurements were performed under the following conditions: 400 µmol mol⁻¹ of CO₂ concentration and photosynthetically active radiation (PAR) at 1500 µmolm^–2s^–1. Three measurements were taken from one plant of each replication for both treatments on 28^th, 56^th, 96^th and 123^rd day after planting.

Chlorophyll index was measured on the fully-expanded youngest leaves with SPAD-502 Plus chlorophyll meter (Konica Minolta). An average of three measurements were taken for each replication. Thereafter, the same leaves, which were used for chlorophyll index measurement, were harvested for chlorophyll content determination. The fresh weight of the leaf samples was measured (around 0.2 g), and the leaves were ground very well using a tissue laser. Then, 1 ml of 80% acetone was added to the samples and shaken and incubated overnight at +4°C. After overnight incubation, the samples were centrifuged for 5 min. at 13.000 g and the supernatant was collected. The supernatant, which had the chlorophyll, was measured at 470, 646.8 and 663.2 nm. The chlorophyll content was calculated according to the following formulas:

Chlorophyll a content = (7,15 x A663.2)/1000/Fresh weight (A)

Chlorophyll b content = (18,71 x A646.8)/1000/Fresh weight (B)

Total chlorophyll content = A + B

Tomato fruits were harvested three times; on 110^th, 128^th and 154^th days after planting. The yield was calculated as the number and the weight of the fruits per plant.

The root, stem, leaf and fruit samples were collected after the final harvest and TiO₂ content of each sample was measured by X-Ray Fluorescence (XRF).

Statistical analysis: The data obtained from experiments were examined by ANOVA test by using MINITAB software. The differences between groups were determined by Fisher’s post-hoc analysis. A 2-paired t-test was performed by using MINITAB.

Characterization of TiO₂ nanoparticles: The purchased TiO₂ nanoparticles were analyzed with XRD and SEM. According to the XRD pattern, the nanoparticles showed strong peaks at 25° and 48° specific to the TiO₂ anatase phase, which confirmed the purification of the nanoparticles (Figure 1) (Theivasanthi and Alagar, 2013).

Besides, the image of TiO₂ nanoparticles was obtained using SEM and the sizes of TiO₂ nanoparticles were measured (Figure 2). According to the image, the sizes of particles were less than 100 nm and confirmed the suitable size for the treatments.

Effect of TiO₂ on tomato seed germination and seedling growth: The effect of nano-priming on seed germination rate and seed vigor index according to the length and the weight of plantlets can be seen in Table 1. The nano-priming did not affect the seed germination rate. The seeds had more than 97% of germination rate after all treatments. On the other hand, the seed vigor index was affected for hydro and nano-priming treatments. When the 10 mg L^-1 of TiO₂ nano-priming treatment increased seed vigor index by length, 50 mg L^-1 of TiO₂ nano-priming increased the seed vigor index by weight the most. On the other hand, 100 mg L^-1 of TiO₂ nano-priming decreased the seed vigor index by length. The water uptake ability of the seeds was increased for the TiO₂ nano-priming treatment. 10 mg L^-1 of TiO₂ treatment increased the water uptake ability of the tomato seeds the most.

Table 1. Effect of hydro and TiO₂ nano-priming on germination rate, seed vigor index.

Means, in each column, followed by a similar letter are not significantly different at the 5% probability level using Fisher’s Multiple Range Test.

GR: Germination rate, SVI-I: Seed vigor index by length, SVI-II: Seed vigor index by weight, WU: Water uptake, T₀: Hydropriming, T₅: 5 mg L^-1 TiO₂ nano-priming, T₁₀: 10 mg L^-1 TiO₂ nano-priming, T₅₀: 50 mg L^-1 TiO₂ nano-priming, T₁₀₀: 100 mg L^-1 TiO₂ nano-priming.

The effect of TiO₂ nano-priming on plantlets’ development was also observed (Figure 3). The shoot length was not affected by TiO₂ nano-priming. However, root length was increased with 10 mg L^-1 of TiO₂ nano-priming treatment. Besides, 100 mg L^-1 of TiO₂ nano-priming decreased the root length of the tomato plantlets. The shoot fresh weight of tomato plantlets was increased with TiO₂ nano-priming treatments at all concentrations, and 50 mg L^-1 of TiO₂ treatment holds the first rank. The heaviest shoot dry weight was measured after 10 mg L^-1 TiO₂ nano-priming and root dry weight after 5 and 100 mg L^-1 TiO₂ treatment (Table 2).

Table 2. Effect of hydro and TiO₂ nano-priming on shoot and root length, shoot and root fresh and dry weight.

Means, in each column, followed by a similar letter are not significantly different at the 5% probability level using Fisher’s Multiple Range Test.

SL: Shoot length, RL: Root length, SDW: Shoot dry weight, RDW: Root dry weight, T₀: Hydropriming, T₅: 5 mg L^-1 TiO₂ nano-priming, T₁₀: 10 mg L^-1 TiO₂ nano-priming, T₅₀: 50 mg L^-1 TiO₂ nano-priming, T₁₀₀: 100 mg L^-1 TiO₂ nano-priming.

Effect of TiO₂ on plant growth and yield: Primed seeds with 10 mg L^-1 of TiO₂ nanoparticles were planted under greenhouse conditions. The stem length of the plants was measured at the flowering period, and it was observed that the nano-priming treatment did not affect the stem length of the plants (Figure 4).

The stomatal conductance and transpiration rate showed the same trend under the same treatments (Figures 5a and 5b). The stomatal conductance increased after TiO₂ nano-priming treatment (Figure 5a). Likewise, the transpiration rate was positively affected by TiO₂ nano-priming treatment and the control group had a lower transpiration rate (Figure 5b).

TiO₂ nano-priming affected the chlorophyll content of tomato plants (Figures 5c-f). When the chlorophyll content of tomato plants was analyzed in detail, the chlorophyll a content increased with the TiO₂ treatment (Figure 5c). Chlorophyll b content also increased with the TiO₂ nano-priming treatment (Figure 5d). As a result, total chlorophyll content also increased when the chlorophyll a and chlorophyll b content were increased (Figure 5e). In contrast to chlorophyll content, relative chlorophyll content was not affected by TiO₂ nano-priming treatment (Figure 5f).

The Pn of plants developed from nanoprimed seeds were higher than the control group (Figure 6). On 56^th day after planting, the photosynthesis performance of the plants was 18% higher than the control (16,20 vs. 19,05). Besides, when the plants started to die, the Pn increased 55% (from 6,96 to 10,85) following the TiO₂ treatment.

The yield of tomato plants was not changed by treatment of TiO₂ nano-priming (Figures 7a and 7b). When the yield was analyzed by fruit number / plant, there was no effect of TiO₂ nano-priming on the yield of tomato plants (Figure 7a). In the same trend, yield of tomato plants as weight was not affected by TiO₂ nano-priming treatment (Figure 7b).

Translocation of TiO₂ nanoparticles in the tomato plant: Translocation of TiO₂ nanoparticles to different parts of the tomato plants was investigated. For this purpose, root, stem, leaf and fruit samples were collected and TiO₂ accumulation in these parts were analyzed. As result, accumulation of TiO₂ nanoparticles was observed in the root (Figure 8).

DISCUSSION

In this study, our focus has been on using TiO₂ NP as priming agent, in order to determine optimal concentration for increasing in tomato seed germination and plant development by minimizing the toxicity of NPs. TiO₂ nano-priming treatment did not change the germination rate of the tomato seeds because the seeds had already high germination rate. Yet, both weight- and length-dependent seed vigor index increased with the TiO₂ nano-priming treatment (Table 1). Nanoparticles have a higher binding ability to seed coat compared to other priming agents (Anand et al., 2019; Mahakham et al., 2017). Nano-priming treatment was shown to affect the hormonal balance in the seed (Yang et al., 2017). Seeds recognize the NP as an external agent and activate gibberellic acid (GA) and abscisic acid (ABA) dependent response mechanisms (Anand et al., 2019; Mahakham et al., 2017). When NPs enter the seed coat, ROS accumulate and seed dormancy is broken via the activation of GA synthesis (Dietz et al., 2016). An increase in ROS after nano-priming is reported in some studies (Chandrasekaran et al., 2020; Oracz and Karpiński, 2016). Thus, the accumulation of ROS probably promotes seed germination (Chandrasekaran et al., 2020).

On the other hand, nano-priming increases ɑ-amylase activity and starts rapid starch degradation (Mahakham et al., 2017; Man et al., 2013). The activity of ɑ-amylase depends on GA and this relationship increases with the integration of nanoparticles in the seed (Mahakham et al., 2017). The effect of nano-priming on phytohormones enhances sugar signaling factors, which promote seed germination (Laby et al., 2000). Even though the TiO₂ nano-priming treatment did not increase the seed germination rate, it speeded up the process by mechanisms that are mentioned above and increased the length and biomass of plantlets, thus resulting in an increased seed vigor index. Even though the root length and biomass of the tomato plants increased after nano-priming treatment, the shoot length of tomato plantlets was not affected (Table 2). The translocation of TiO₂ nanoparticle studies showed that TiO₂ nanoparticles were accumulated in the root (Figure 8). While the stem length was not affected by the TiO₂ nano-priming treatment, the probable reason for the increase in the root length is the TiO₂ accumulation. Translocation of TiO₂ nanoparticles was investigated in basil and similarly, when the TiO₂ accumulation was observed in roots, there was no accumulation in the shoots (Tan et al., 2018). Treatment of TiO₂ nanoparticles by sludge was shown not to affect the accumulation of TiO₂ in the stem, leaf, or fruit parts of the tomato plant (Bakshi et al., 2019). The size of the TiO₂ affects the accumulation of the NPs in different parts of the plant. If the NPs are bigger than 140 nm, they cannot accumulate in the root. The threshold value for accumulation in the stem is 36 nm for wheat (Larue et al., 2012). The size of the nanoparticles was approximately 70 nm in the present study (Figure 2), which may have prevented the translocation of NPs from the root through the stem. The absence of TiO₂ nanoparticles especially in the fruit of tomato plants is an advantage for using this material as a priming agent, considering that this technique counteracts the risks and concerns about TiO₂ contamination in the edible part of the tomato.

After in vitro conditions, the effect of TiO₂ nano-priming on tomato plants was observed under the greenhouse conditions. For this purpose, 10 mg L^-1 of TiO₂ was selected for greenhouse experiments according to the seed vigor index results. The stem length of the tomato plants was not affected by TiO₂ nano-priming treatment, which was expected, similar to the shoot length of the tomato plantlets.

Some physiological traits of the tomato plants were increased by TiO₂ nano-priming treatment (Figure 5). The stomatal conductance and transpiration rate of the tomato plants presented a similar trend after TiO₂ nano-priming (Figures 5a and 5b). Stomatal conductance and transpiration rates increased due to the opening of stomata (Qi et al., 2013). This may be caused by the increases in the root length and water uptake, which decreased the water need of the plant and increased the stomatal conductance and transpiration rate. However, the mechanisms behind the increases in stomatal conductance and transpiration rate after TiO₂ nano-priming treatment are not yet fully understood.

As the core reactive molecule in the photosystem I and the photosystem II, Chlorophyll a content was increased following the TiO₂ nano-priming (Figure 5c) (Blankenship, 2013). Similarly, chlorophyll b content increased with the TiO₂ treatment (Figure 5d), which increased the wavelength range and amount of light absorbed by plants during photosynthesis (Conway et al., 2015). The increases in both chlorophyll a and b molecules lead inevitably to an increase in total chlorophyll content (Figure 5e). However, relative chlorophyll content was not affected by TiO₂ nano-priming treatment. Similar results were observed in a study where, nano and bulk-TiO₂ treated spinach plants had higher chlorophyll content 37.48% and 13.34%, respectively (Yang et al., 2007). Furthermore, an increase in chlorophyll content week by week after continuous TiO₂ treatment in wheat was reported (Refique et al., 2018). However, it was highlighted that TiO₂ treatment in tomatoes did not affect the chlorophyll a and b content (Bakshi et al., 2019). The method of application is also a factor that needs to be taken into account for the chlorophyll content of tomatoes since the foliar application leads to a higher increase in the total chlorophyll content in comparison to priming (Perveen and Siddique, 2022).

The photosynthesis rate of the tomato plants increased after 56 DAP and 123 DAP with the TiO₂ nano-priming treatment (Figure 6). The photosynthesis rate is correlated with chlorophyll content. As mentioned above, the chlorophyll content of the tomato plant was increased by TiO₂ nano-priming treatment in the current study. By increasing the chlorophyll content a and b increased the photosynthesis ratio individually. Besides, TiO₂ promoted photochemical reactions and increased the photosynthesis activity in spinach (Hong et al., 2005). TiO₂ treatment also caused conformational changes in the rubisco activase enzyme and increased photosynthesis rate (Gao et al., 2006, 2008). TiO₂ nanoparticles have a role as photocatalysts and promote the redox reaction (Higashimoto, 2019). Consequently, the activation of Rubisco by TiO₂ treatment increases, thereby leading to an increase in the photosynthesis ratio (Yang et al., 2008). In the current study, the photosynthesis rate increased when the plant was in the flowering stage and before dying, which were critical periods for the plant's development.

Although TiO₂ nano-priming increased plant development and growth, the yield of tomato plants was not affected by TiO₂ nano-priming treatment (Figures 7a and 7b). There are various studies that have shown increases in the yield of tomato plants after TiO₂ treatment. For instance, the yield of tomato plants was reported to increase after 100 mg L^-1 by foliar spray application (Choi, 2021). In another study, the positive effect of TiO₂ treatment on the yield of tomato plant by foliar application was reported (Parveen and Siddique, 2022). The reason why there was no effect on yield in tomato plants in the study is thought to be due to the use of the priming method.

Conclusion: Use of traditional chemicals during plant production has hazardous effects on the environment and human health. Thus, different technologies have been developed to decrease this toxic effect and one of them is nanoparticles. However, using nanoparticles can cause phytotoxicity; therefore, the determination of the correct concentration and application technique is very important. With the in vitro experiments, it was observed that using a very small amount of TiO₂ (10 mg L^-1) as a priming agent induced plant development. Under greenhouse conditions, the TiO₂ nano-priming increased the chlorophyll content and photosynthesis rate of the tomato plants. During this process, the nanoparticles did not translocate in the plant, which was a fundamental issue regarding the use of nanoparticles in crop production. TiO₂ nano-priming application had a positive effect on tomato plant development and increased chlorophyll content and photosynthesis rate, which could be a possible alternative to traditional fertilizers. However, the impact of TiO₂ nanoparticles on plant development should be investigated in more detail.

Acknowledgment: The study was supported by Niğde Ömer Halisdemir University Research Project Unit with FEB2017/31-BAGEP numbered project. The authors would like to thank stuff of Central Research Laboratory of Niğde Ömer Halisdemir University for performing SEM, XRD and XRF analysis of the study.

Author contributions: AKY and MEC designed the study. AKY performed the experiments and collected data. AKY and MEC analyzed data. AKY wrote the manuscript and MEC reviewed and edited the manuscript.

Conflict of Interest: The authors declare that they have no conflict of interest.

REFERENCES

1. Almutairi, Z. M. (2016). Effect of nano-silicon application on the expression of salt tolerance genes in germinating tomato (Solanum lycopersicum) seedlings under salt stress. Plant Omics 9(1): 106-114. Avaible from: https://search.informit.org/doi/10.3316/informit.888088806058398
2. Almutairi, Z.M. and A. Amjad (2015). Effect of Silver Nanoparticles on Seed Germination of Crop Plants. Adv. Chem. 4: 280–285. doi.org/10.5281/zenodo.1108056
3. Anand A., A. Kumari, M. Thakur and A. Koul (2019). Hydrogen peroxide signaling integrates with phytohormones during the germination of magnetoprimed tomato seeds. Sci. Rep. 9(1): 1-11. https://doi.org/10.1038/s41598-019-45102-5
4. Azimi R., M. J. Borzelabad, H. Feizi, and A. Azimi (2014). Interaction of SiO2 nanoparticles with seed prechilling on germination and early seedling growth of tall wheatgrass (Agropyron elongatum). Pol. J. Chem. Technol. 16(3): 25-29. https://doi.org/10.2478/pjct-2014-0045
5. Bakshi M., C. Liné, D. E. Bedolla, R. J. Stein, R. Kaegi, G. Sarret, A.E. Pradas del Realg, H. Castillo-Michelh, P.C. Abhilaj and C. Larue (2019). Assessing the impacts of sewage sludge amendment containing nano-TiO₂ on tomato plants: A life cycle study. J. Hazard. Mater. 369: 191-198. https://doi.org/10.1016/j.jhazmat.2019.02.036
6. Bekhrad, H., B. Mahdavi, and A. Rahimi (2016). Effect of seed priming on growth and some physiological characteristics of sesame (Sesamum indicum ) under salinity stress condition caused by Alkali Salts. Iran. Field Crops Res. 13(4): 810-822. DOI: 10.22067/GSC.V13I4.36039
7. Blankenship, R.E. (2013). Molecular Mechanisms of Photosynthesis. John Wiley & Sons, New York, NY. DOI: 1002/9780470758472
8. Chandrasekaran, U., X. Luo, Q. Wang and K. Shu (2020). Are there unidentified factors involved in the germination of nanoprimed seeds?. Front. Plant Sci. 11: 832. DOI: 3389/fpls.2020.00832
9. Choi, H. G. (2021). Effect of TiO₂ Nanoparticles on the Yield and Photophysiological Responses of Cherry Tomatoes during the Rainy Season. Horticulturae. 7(12): 563. https://doi.org/10.3390/horticulturae7120563
10. Conway, J. R., A. L. Beaulieu, N. L. Beaulieu, S. J. Mazer and A. A. Keller (2015). Environmental stresses increase photosynthetic disruption by metal oxide nanomaterials in a soil-grown plant. ACS Nano, 9(12): 11737-11749. https://doi.org/10.1021/acsnano.5b03091
11. Dhoke, S. K., P. Mahajan, R. Kamble and A. Khanna (2013). Effect of nanoparticles suspension on the growth of mung (Vigna radiata) seedlings by foliar spray method. Nanotechnol. Dev. 3(1): e1-e1. https://doi.org/10.4081/nd.2013.e1
12. Dietz, K. J., R. Mittler and G. Noctor (2016). Recent progress in understanding the role of reactive oxygen species in plant cell signaling. Plant physiol. 171(3): 1535-1539. DOI: 1104/pp.16.00938
13. Dutta, P. (2018). Seed priming: new vistas and contemporary perspectives. In Advances in seed priming. Springer, Singapore. 3-22 p. https://doi.org/10.1007/978-981-13-0032-5_1
14. El‐Temsah, Y. S. and E. J. Joner (2012). Impact of Fe and Ag nanoparticles on seed germination and differences in bioavailability during exposure in aqueous suspension and soil. Environ. Toxicol. 27(1): 42-49. DOI: 1002/tox.20610
15. Feizi, H., S. Amirmoradi, F. Abdollahi and S. J. Pour (2013). Comparative effects of nanosized and bulk titanium dioxide concentrations on medicinal plant Salvia officinalis Annu. Rev. Res. Biol. Artic. 3: 814-824. Available from: https://journalarrb.com/index.php/ARRB/article/view/24904
16. Finch-Savage, W. E., K. C. Dent and L. J. Clark (2004). Soak conditions and temperature following sowing influence the response of maize (Zea mays) seeds to on-farm priming (pre-sowing seed soak). Field Crops Res. 90(2-3): 361-374. https://doi.org/10.1016/j.fcr.2004.04.006
17. Gao, F., C. Liu, C. Qu, L. Zheng, F. Yang, M. Su and F. Hong (2008). Was improvement of spinach growth by nano-TiO₂ treatment related to the changes of Rubisco activase?. BioMetals. 21(2): 211-217.
18. Gao, F., F. Hong, C. Liu, L. Zheng, M. Su, X. Wu, F. Yang, C. Wu and P. Yang (2006). Mechanism of nano-anatase TiO₂ on promoting photosynthetic carbon reaction of spinach. Biol. Trace Elem. Res. 111(1): 239-253.
19. Higashimoto, S. (2019). Titanium-dioxide-based visible-light-sensitive photocatalysis: Mechanistic insight and applications. Catalysts 9(2): 201-3390. DOI: 1007/s10534-007-9110-y
20. Hong, F., J. Zhou, C. Liu, F. Yang, C. Wu, Zheng and P. Yang (2005). Effect of nano-TiO₂ on photochemical reaction of chloroplasts of spinach. Biol. Trace Elem. Res. 105(1): 269-279. DOI: 10.1385/BTER:105:1-3:269
21. Kah, M., S. Beulke, K. Tiede and T. Hofmann (2013). Nanopesticides: state of knowledge, environmental fate, and exposure modeling. Crit. Rev. Environ. Sci. Technol. 43: 1823–1867. https://doi.org/10.1080/10643389.2012.671750
22. Khalaki A, M., M. Moameri, B. Asgari Lajayer and T. Astatkie (2021). Influence of nano-priming on seed germination and plant growth of forage and medicinal plants. Plant Growth Regul. 93(1): 13-28. https://doi.org/10.1007/s10725-020-00670-9
23. Laby, R. J., M. S. Kincaid, D. Kim and S. I. Gibson (2000). The Arabidopsis sugar‐insensitive mutants sis4 and sis5 are defective in abscisic acid synthesis and response. Plant J. 23(5): 587-596. DOI: 1046/j.1365-313x.2000.00833.x
24. Larue, C., J. Laurette, N. Herlin-Boime, H. Khodja, B. Fayard, A. M. Flank, F. Brisset and M. Carriere (2012). Accumulation, translocation and impact of TiO₂ nanoparticles in wheat (Triticum aestivum): influence of diameter and crystal phase. Sci. Total Environ. 431: 197-208. DOI: 0.1016/j.scitotenv.2012.04.073
25. Li, X., K. Schirmer, L. Bernard, Sigg, S. Pillai and R. Behra (2015). Silver nanoparticle toxicity and association with the alga Euglena gracilis. Environ. Sci. Nano 2(6): 594-602. DOI: 10.1039/C5EN00093A
26. Mahakham, W., A. K. Sarmah, S. Maensiri and P. Theerakulpisut (2017). Nano-priming technology for enhancing germination and starch metabolism of aged rice seeds using phytosynthesized silver nanoparticles. Sci. Rep. 7(1): 1-21. https://doi.org/10.1038/s41598-017-08669-5
27. Mahmoodzadeh, H., M. Nabavi and H. Kashefi (2013). Effect of nanoscale titanium dioxide particles on the germination and growth of canola (Brassica napus). J. Ornam. Hortic. 3(1): 25-32.
28. Man, J., Y. Yang, C. Zhang, F. Zhang, Y. Wang, M. Gu, Q. Liu and C. Wei (2013). Morphology and structural characterization of high-amylose rice starch residues hydrolyzed by porcine pancreatic α-amylase. Food Hydrocoll. 31(2): 195-203. Available from: https://api.semanticscholar.org/CorpusID:55348915
29. Moammeri, M., E. Alijafari and A. Ghorbani (2018). Effects of nano-priming and bioprimingon on growth characteristics of Onobrychis sativa Lam. under laboratory conditions. J. Rangel. 12(1): 101-111. Available from: http://rangelandsrm.ir/article-1-562-en.html
30. Oracz, K., and S. Karpiński (2016). Phytohormones signaling pathways and ROS involvement in seed germination. Front. Plant Sci. 7(1-6): 864. https://doi.org/10.3389/fpls.2016.00864
31. Parveen, A. and Z. A. Siddiqui (2022). Foliar spray and seed priming of titanium dioxide nanoparticles and their impact on the growth of tomato, defense enzymes and some bacterial and fungal diseases. Arch. Phytopathol. Pflanzenschutz 55(5): 527-548. https://doi.org/10.1080/03235408.2022.2035535
32. Peralta-Videa, J. R., L. Zhao, M. L. Lopez-Moreno, G. de la Rosa, J. Hong and J. L. Gardea-Torresdey (2011). Nanomaterials and the environment: a review for the biennium 2008–2010. J. Hazard. Mater. 186(1): 1-15.
33. Qi, M., Y. Liu and T. Li (2013). Nano-TiO₂ improve the photosynthesis of tomato leaves under mild heat stress. Biol. Trace Elem. Res. 156(1): 323-328. DOI: 1007/s12011-013-9833-2
34. Rafique, R., Z. Zahra, N. Virk, M. Shahid, E. Pinelli, T. J. Park, J. Kallerhoff and M. Arshad (2018). Dose-dependent physiological responses of Triticum aestivum to soil applied TiO₂ nanoparticles: alterations in chlorophyll content, H₂O₂ production, and genotoxicity. Agri. Ecosyst. Environ. 255: 95-101. https://doi.org/10.1016/j.agee.2017.12.010
35. Tan, W., W. Du, A. J. Darrouzet-Nardi, J. A. Hernandez-Viezcas, Y. Ye, J. R. Peralta-Videa and J. L. Gardea-Torresdey (2018). Effects of the exposure of TiO₂ nanoparticles on basil (Ocimum basilicum) for two generations. Sci. Total Environ. 636: 240-248. DOI: 1016/j.scitotenv.2018.04.263
36. Theivasanthi, T. and M. Alagar (2013). Titanium dioxide (TiO₂) nanoparticles XRD analyses: an insight. arXiv preprint arXiv:1307.1091. https://doi.org/10.48550/arXiv.1307.1091
37. Yang, F., C. Liu, F. Gao, M. Su, X. Wu, L. Zheng, F. Hong and P. Yang (2007). The improvement of spinach growth by nano-anatase TiO₂ treatment is related to nitrogen photoreduction. Biol. Trace Elem. Res. 119(1): 77-88. DOI: 1007/s12011-007-0046-4
38. Yang, F., F. Hong, W. You, C. Liu, F. Gao, C. Wu and P. Yang (2006). Influence of nano-anatase TiO₂ on the nitrogen metabolism of growing spinach. Biol. Trace Elem. Res. 110(2): 179-190. https://doi.org/10.1385/BTER:110:2:179
39. Yang, X., C. Cao, L. Erickson, K. Hohn, R. Maghirang and K. Klabunde (2008). Synthesis of visible-light-active TiO₂-based photocatalysts by carbon and nitrogen doping. J. Catal. 260(1): 128-133. https://doi.org/10.1016/j.jcat.2008.09.016