Abstract
Cold atmospheric pressure plasma (CAP) has garnered significant attention in recent years for its potential applications in biomedical, environmental, and agricultural fields. Cold plasma treatment exhibits a variety of effects in agricultural applications, including impacts on seed germination and seedling growth; however, further research is required. Soybean serves as a fundamental source of nutrients for both animals and humans. Soybean seeds possess impermeable and thick testae, which results in prolonged germination times and suboptimal germination rates. The soybeans exhibit low uniformity. As a result, poor crop establishment and yield reduction are inevitable outcomes. Therefore, the purpose of this study was to examine the effects of Iranian soybean cultivars, such as Sari, Saba, Arian, Katoul, and Williams, on seedling growth properties, seed germination, and antioxidant enzyme activity, using argon at time intervals of 30, 60, 180, 300, and 420 s. Cold plasma treatment significantly enhanced germination potential from 1.18 to 66.97%, germination index from 0.50 to 60.09%, germination rate from 1.78 to 32.17%, seedling length from 2.70 cm to 78.13 cm, root length from 2.87 cm to 56.13 cm, and seedling dry weight from 1.80 g to 36.63 g. Additionally, CAT activity increased from 0.88- to 4.40-fold, SOD activity from 0.86- to 5.89-fold, and APX activities from 0.40- to 4.01-fold compared to the control treatment. The findings indicated that the samples exhibited optimal results at treatment durations of 60 and 180 s. The influence of plasma on the antioxidant responses of seedlings, seed germination, and growth characteristics was contingent upon the duration of treatment. Cold plasma, when applied for an appropriate duration, may enhance soybean seedling growth characteristics and seed germination.
Supplementary Information
The online version contains supplementary material available at 10.1186/s12896-024-00921-x.
Keywords: DBD cold plasma, Catalase (CAT), Superoxide dismutase (SOD), Ascorbate peroxidase (APx), Optical emission spectroscopy
Introduction
Soybean is an economically import oilseed crop which is mainly cultivated for its high protein (40%), oil content (20%) and other valuable nutritional materials. This crop belongs to Leguminosae family which has the ability of reinforcement of soil due to nitrogen biologically fixation following by productivity and stability enhancement of soil texture and reduction of chemical fertilizer usage for next crops in rotation [1]. In the world, the various consumptions of soybean are referred to food (for human and livestock), medical usages and industrial products which cause the increase need for its production. The top five superior countries with the highest rate of soybean production consists of United states, Brazil, Argentina, China and India with total production area under cultivation of 129, 523,964 ha and production rate of 353,370,766 t [2].
Soybean seeds have thick and impermeable testae, resulting in poor germination, long germination time and low germination uniformity, which subsequently leads to poor establishment and yield reduction [3]. Promoting seed germination is the most direct way to improve soybean production. Methods for promoting seed germination are physical methods (magnetic treatment, sunlight, ultraviolet light and hot water soaking) and chemical methods (chemicals, fungicides and hormones) [4–6]. Although these methods can promote germination to a certain extent, they are time consuming, labor-intensive and produce chemical residues [7].
Plasma technology is nonthermal technique applied to foods in recent years [8]. Plasma treatment of seeds is a new approach that is being proposed to assist germination and survival [9]. Plasma is formed by a discharge in a gas, and in the case of an air plasma consists of ions, energetic electrons, neutral species, reactive oxygen species (ROS) and reactive nitrogen species (RNS) and produces electromagnetic radiation such as UV. Preliminary investigations have confirmed that plasma pre-treatment of seeds of important agricultural crops is an effective tool for improvement of germination, shoot, and root growth [10–12]. The plasma treatments provide good fungicidal and bactericidal effects, and increased water permeability through surface coat etching and stimulation of germination and seedlings growth [11, 13].
Various types of plasma sources have been created in the last 20 years such as dielectric barrier discharge (DBD), jet plasma, microwave discharge, radiofrequency (RF) discharge, and corona discharge. The seed germination was shown to vary depending on the plasma sources, seed type, treatment period, feed gas, and humidity [14]. Dielectric barrier discharge (DBD) is a new technique of cold plasma generation that has excellent treatment flexibility, and low power requirements, therefore, a great deal of biological goals can be achieved through this technique [15–17].
Numerous examinations have confirmed the positive effect of cold plasma on plant growth performance. The mechanism of this approach is to activate physiological functions in plasma-treated seeds. One study showed that cold plasma increased the expression of essential enzymes such as peroxidase, polyphenol oxidase, and phenylalanine ammonia-lyase from tomato seeds [18].
In another study, the sunfower seeds treated for 7 min before sowing increased germination, leaf weight, and seedling growth. In addition, gibberellin and abscisic acid production was modifed to increase germination [19]. In addition, the study by Renáta Švubová et al. (2021) showed that cold plasma increased the activity of succinate dehydrogenase in soybean, proving the switching of the germinating seed metabolism from anoxygenic to oxygenic. The results also showed that SOD, and GSH activity increased compared to the control group [20].
This study is motivated by the significance of global plant production, particularly in the context of climate change and the increasing food demands of human populations. Evaluating eco-agricultural technologies, including cold plasma, is crucial for enhancing seed quality, promoting initial plant growth and development, and ultimately increasing yield in crop production. Although some researches are performed in the area of cold plasma and plant seedlings, no research about the impact of plasma on germination of Iranian soybean cultivars have been examined so far. Hence, this study was performed to inspect the effect of cold plasma treatment on seedling growth, seed germination, and antioxidant enzyme activity of important soybean cultivars in Iran including Sari, Saba, Arian, Katoul, and Williams under time intervals of 30, 60, 180, 300, and 420 s of DBD using argon.
Materials and methods
Plant materials
The soybean seeds utilized in this study were sourced from Karaj Seed and the Seedling Registration and Certification Research Institute. This study utilized seeds from five distinct cultivars: Sari, Saba, Arian, Katoul, and Williams. Each cultivar comprised two categories: Registered seeds and Certified seeds. Registered seeds are the offspring of foundation seeds cultivated by specific farmers. The mean germination rate (MGR) of various soybean cultivars was 80%. The seeds were cultivated to maintain genetic identity and purity. Certified seeds are defined as seeds that originate from a recognized variety and are produced under stringent seed certification standards to ensure varietal purity. The soybean cultivars suitable for cultivation in Iran are listed in Table 1. The size and weight of soybean seeds varied significantly among different cultivars (Table 1). Katoul cultivars represent the primary soybean cultivars in Golestan province, Sari, and are also the predominant cultivar in Mazandaran province. In Ardabil province (Moghan), the key soybean cultivars include Williams, Saba, and Arian. Golestan and Ardabil provinces rank first and second, respectively, in cultivation area; however, in terms of grain production, Ardabil ranks first and Golestan ranks second.
Table 1.
Characteristics of studied soybean cultivars
| Cultivar | Class (within cultivar) | The size of each seed (cm) | Weight of each seed (gr) |
|---|---|---|---|
| Sari | Registered | 4.55 | 0.196 |
| Certified | 4.25 | 0.175 | |
| Saba | Registered | 5.25 | 0.165 |
| Certified | 5.05 | 0.146 | |
| Arian | Registered | 5.35 | 0.249 |
| Certified | 5.15 | 0.219 | |
| Katoul | Registered | 4.95 | 0.198 |
| Certified | 4.80 | 0.154 | |
| Williams | Registered | 4.75 | 0.161 |
| Certified | 4.60 | 0.132 |
Plasma treatment
The dielectric barrier discharge (DBD) plasma, developed by High Tech Company (0045, Tehran, Iran), utilized two flat aluminum electrodes measuring 45 cm in length, 6.5 cm in width, and 0.02 cm in thickness. A mica-derived insulating sheet is placed between the electrodes to restrict the current flow to 1 µA. The dimensions of this sheet are 60 cm in length, 12 cm in width, and 0.01 cm in thickness. A plexiglass shield measuring 0.03 cm in width was installed between the electrodes. The value in the current study was measured using a thermocouple in electrodes, with the temperature recorded at approximately 40 °C. The cold plasma system was enclosed within an acrylic chamber equipped with gas inlet and outlet ports. The distance between the tip of the plasma plume and the seeds was maintained at 20 mm during plasma treatments. The existing supply was linked to an alternating current with a primary voltage of 220 V. The applied voltage and frequency on the plasma plume were 5 kV and 8 kHz, respectively, with a power output of 100 W. The feed gas utilized was argon, with a flow rate of 2 L/min. The seeds were subjected to cold plasma treatment for durations of 30, 60, 180, 300, and 420 s at the Physics Research Center of Azad University, Research and Sciences Branch, Tehran (Fig. 1).
Fig. 1.
Schematic of DBD treatment chamber
Germination and growth parameters
A total of 25 seeds were subjected to cold plasma treatment for time intervals of 30, 60, 180, 300, and 420 s on sterile petri dishes. The seeds were subsequently placed in a germinator maintained at a constant temperature of 25 °C for a duration of seven days. The germinated seeds were counted and documented daily throughout the experiment. The criterion for seed germination was approximately 3 mm in root length. During the experiment, 5 ml of distilled water was added daily to the Petri dishes to maintain humidity for germination. The seeds were incubated in a 25 °C light incubator, and the germination percentage was recorded daily for seven days. Germination of the seeds should occur when the length of the radicals reaches half the length of the seed [21] Root and seedling lengths were measured on the seventh day, and to assess dry weight, ten seedlings were randomly selected from each repetition and dried in an oven for 48 h at 70 °C. Seedlings were placed in an oven at 70 °C for 48 h to determine dry weight. The germination index (GI), germination potential (GP), and germination rate (GR) were assessed following the methodology outlined by Ling et al. (2014) [22].
Germination potential (%) = (Number of seeds germinated in 3 days /total number of seeds) × 100
Germination rate (%) = (Number of seeds germinated in 7 days /total number of seeds) × 100
Germination index = Σ (Gt/Dt)
Where, Dt shows germination days, and Gt shows the number of germinated seeds on the t day.
Enzyme extraction
The enzyme extraction was conducted following the methodology of Kar and Mishra (1976), with certain modifications [23]. Initially, 200 mg of fresh leaf tissue was weighed, cut into small pieces, and subsequently ground into a powder with a Chinese mortar with the aid of liquid nitrogen. Subsequently, 3 ml of 100 mM potassium phosphate buffer at pH 6.8 was added and thoroughly mixed. The sample was transferred to a 5 ml microtube and centrifuged at 17,000 g for 15 min at 4 °C. The supernatant was subsequently transferred to 2 ml microtubes for protein extraction and stored at -20 °C.
Enzymatic assays
Catalase (CAT)
The activity of CAT (EC 1.11.1.6) was assessed in a reaction mixture containing 50 mM K-PO4 buffer (pH 7.0), distilled water, 15 mM hydrogen peroxide (H2O2), and 0.1 ml enzyme extract solution (extinction coefficient of 40 mM-1cm-1, and shown as µM g-1). The absorbance was measured at 240 nm for 60 s [24].
Superoxide dismutase (SOD)
The activity of the superoxide dismutase (SOD) enzyme (EC 1.15.1.1) was assessed using a colorimetric method, including the measurement of formazan crystals. This was done by mixing 50 mM potassium-phosphate (K-PO4) buffer solution (pH 7.8), 0.1 mM ethylenediaminetetraacetic acid (EDTA), 82.5 µL nitroblue tetrazolium (NBT), 2.2 µM riboflavin, 14.3 mM methionine, and the 200 µL of enzyme extract. The absorbance of the reaction mixture was then measured at 560 nm. Then, the negative control sample containing the above solution except the cell-free extract was placed under the fluorescent light as a control for comparison and final calculation, reporting results in U/min/mg/protein [25].
Ascorbate peroxidase (APX)
The activity of the Ascorbate peroxidase (APX) enzyme (EC 1.11.1.11) was measured based on the Asado and Nakano methodology [26]. This was done by mixing 50 mM potassium-phosphate (K-PO4) buffer solution (pH 7.0), 0.1 mM EDTA, 0.5 mM Ascorbate, 0.1 mM H2O2, 10 µL enzyme extracts. The absorbance was measured at 290 nm for 60 s.
Optical emission spectroscopy
Optical emission spectroscopy (OES, Ocean Optics, HR4000CG-UV-NIR, USA) was employed to analyze the UV–visible emission spectrum linked to the generated free radicals. Voltage measurements were obtained using a high voltage probe (P6015A Tektronix HV), while current was measured with a digital oscilloscope (Tektronix MSO4032) paired with a TCP202 Tektronix current probe (5 kV) [11].
SEM (scanning Electron microscope)
Plasma-treated seeds and control seeds were examined using a scanning electron microscope (FEI S50, FEI Technologies Inc., Oregon, United States) to analyze alterations in seed structure, surface topology, and overall surface characteristics. SEM analysis was conducted on both non-treated and plasma-treated seeds, with treatment durations of 60 and 240 s. Images were captured at microscopic magnifications of 80X, 2500X, and 5000X using an accelerating voltage of 20 kV and secondary electron scanning mode, with distances of 1 mm, 50, and 20 μm [11].
Statistical analysis
The statistical analysis of the data was conducted using SPSS 16.0 software (SPSS Inc., Chicago, Illinois, USA). The data were reported as the mean ± standard error of the mean (SEM). The data underwent statistical analysis using a one-way analysis of variance (ANOVA). Given the exploratory character of the investigation, statistical significance was determined by considering probability values (P) that were less than or equal to 0.05.
Results
Seed germination and seedling properties
The results of the studied cultivars exhibited significant differences in seed germination and seedling characteristics, highlighting considerable genetic diversity among the cultivars. Plasma treatment demonstrated significant differences across all traits. The interaction effect of cultivar × time and class × time was significant, as it demonstrated the varying responses of cultivars and classes at different durations of plasma treatment (Table 2).
Table 2.
Results of analysis of variance for seed germination and seedling growth characteristics in soybean
| Means of Square | |||||||
|---|---|---|---|---|---|---|---|
| S.O.V | df | Germination percentage | Germination Rate | Germination index | Root length | Seedling length | Seedling dry weight |
| Time | 5 | 330.708** | 255.667** | 278.876** | 72.995** | 707.616** | 0.253** |
| Cultivar | 4 | 501.853** | 516.071** | 809.065** | 106.986** | 528.487** | 1.455** |
| Cluster | 1 | 700.057** | 209.162** | 1229.971** | 458.587** | 269.765** | 0.350** |
| Time × Cultivar | 20 | 36.085** | 35.910** | 47.429** | 3.42 | 9.214** | 0.042** |
| Cultivar × Cluster | 4 | 182.91** | 97.278** | 228.877** | 41.671** | 80.426** | 0.043* |
| Time × Cluster | 5 | 85.203** | 30.441** | 65.252** | 5.169** | 39.672** | 0.021ns |
| Time × Cultivar × Cluster | 20 | 34.985** | 50.799** | 28.567** | 4.708** | 17.241** | 0.031** |
| Error | 180 | 13.394 | 12.043 | 13.259 | 1.333 | 3.679 | 0.015 |
| CV (%) | - | 3.94 | 3.73 | 6.23 | 7.27 | 5.42 | 8.74 |
ns, **,*: Not significant, significant at 1% and 5% levels
Seed germination properties
Germination potential (GP)
The results demonstrated a significant improvement in GP after plasma treatment (P ≤ 0.05). The findings revealed that the maximum and minimum GP were 98.13 ± 0.66% and 54.50 ± 2.22%, respectively, as reported in the Arian-Certified report for the control treatment (Table 3). The results indicated that Williams and Sari exhibited the highest percentage of GP associated with plasma treatment at the 30th, 60th, and 180th s. The control group in Saba and Arian cultivars exhibited the lowest germination potential, as shown in Table 3. The GP for all control cultivars increased by 1.18–66.97% after 7 days of plasma treatment. The highest GP for Arian-Certified plasma treatments occurred at 60 s, while the lowest increase in germination potential compared to all control cultivars was noted for Katoul-Certified at 420 s.
Table 3.
Effect of cold plasma on seed germination properties in soybean
| Cultivar | Class (within cultivar) | Plasma treatment | Germination potential (%) | Germination Rate (seed day − 1) | Germination index |
|---|---|---|---|---|---|
| Sari | Registered | 0 | 85.50 ± 1.26f-m | 92.75 ± 0.63a-k | 57.12 ± 1.18 g-p |
| 30 | 96.25 ± 0.85ab | 98.13 ± 0.43a | 66.19 ± 1.03a | ||
| 60 | 96.00 ± 1.22abc | 98.00 ± 0.61a | 63.94 ± 1.22a-d | ||
| 180 | 98.13 ± 0.66a | 98.13 ± 0.66a | 64.31 ± 1.25abc | ||
| 300 | 93.75 ± 0.75a-e | 94.88 ± 0.37a-h | 62.85 ± 0.69a-f | ||
| 420 | 95.25 ± 1.80a-d | 97.13 ± 1.39a-d | 63.51 ± 1.49a-d | ||
| Certified | 0 | 88.25 ± 1.11d-l | 94.13 ± 0.55a-h | 58.34 ± 1.06d-m | |
| 30 | 94.50 ± 0.29a-d | 97.25 ± 0.14abc | 62.44 ± 0.29a-g | ||
| 60 | 95.75 ± 0.48abc | 97.88 ± 0.24a | 63.69 ± 0.48a-d | ||
| 180 | 95.75 ± 1.49abc | 97.38 ± 0.97abc | 63.69 ± 0.48a-d | ||
| 300 | 95.50 ± 1.50a-d | 97.25 ± 1.11abc | 63.69 ± 0.48a-d | ||
| 420 | 90.00 ± 1.29b-i | 93.25 ± 1.44a-j | 63.69 ± 0.48a-d | ||
| Saba | Registered | 0 | 72.75 ± 3.66p | 82.38 ± 2.38n | 63.69 ± 0.48a-d |
| 30 | 85.75 ± 3.50f-m | 88.88 ± 3.33 g-m | 55.40 ± 2.81j-r | ||
| 60 | 88.75 ± 1.93c-k | 92.38 ± 1.76a-l | 57.48 ± 1.84f-o | ||
| 180 | 91.00 ± 1.29a-h | 94.00 ± 1.47a-h | 59.41 ± 1.15c-l | ||
| 300 | 81.75 ± 1.44k-o | 87.88 ± 1.71i-n | 51.91 ± 1.15o-t | ||
| 420 | 84.50 ± 1.76 h-n | 90.25 ± 2.03e-l | 49.99 ± 0.62rst | ||
| Certified | 0 | 79.50 ± 1.76mno | 86.75 ± 2.11k-n | 50.61 ± 2.10q-t | |
| 30 | 86.50 ± 1.55f-m | 92.25 ± 1.76a-l | 55.67 ± 1.15i-q | ||
| 60 | 85.50 ± 1.19f-m | 91.25 ± 0.92c-l | 54.88 ± 1.46j-r | ||
| 180 | 84.75 ± 1.80 g-n | 89.88 ± 1.83f-m | 54.18 ± 1.62k-s | ||
| 300 | 88.25 ± 3.17d-l | 94.63 ± 2.78a-h | 56.53 ± 3.09i-p | ||
| 420 | 87.25 ± 3.12e-l | 90.38 ± 2.55e-l | 56.84 ± 2.20 h-p | ||
| Arian | Registered | 0 | 84.50 ± 0.65 h-n | 90.25 ± 0.66e-l | 54.05 ± 0.66 L-s |
| 30 | 90.75 ± 2.14a-h | 93.88 ± 1.51a-i | 59.90 ± 1.88b-j | ||
| 60 | 92.00 ± 2.12a-g | 96.00 ± 1.06a-f | 62.31 ± 1.09a-h | ||
| 180 | 92.25 ± 1.65a-f | 94.63 ± 1.25a-h | 61.15 ± 1.42a-i | ||
| 300 | 85.50 ± 4.33f-m | 87.63 ± 3.13j-n | 56.00 ± 3.27i-q | ||
| 420 | 81.50 ± 3.38k-o | 84.25 ± 3.90mn | 52.73 ± 2.01 m-t | ||
| Certified | 0 | 54.50 ± 2.22q | 71.88 ± 2.05 o | 37.01 ± 1.31v | |
| 30 | 84.75 ± 0.75 g-n | 91.00 ± 0.84d-l | 48.52 ± 3.64t | ||
| 60 | 91.00 ± 2.71a-h | 95.00 ± 1.85a-g | 59.26 ± 2.39c-l | ||
| 180 | 89.75 ± 1.55b-j | 91.25 ± 3.44c-l | 57.81 ± 0.66e-n | ||
| 300 | 86.50 ± 4.09f-m | 91.25 ± 3.44c-l | 52.80 ± 2.07 m-t | ||
| 420 | 82.50 ± 3.62j-o | 88.75 ± 3.26 h-m | 52.61 ± 2.43n-t | ||
| Katoul | Registered | 0 | 81.25 ± 0.75 L-o | 90.25 ± 0.43e-l | 54.89 ± 0.44j-r |
| 30 | 94.75 ± 1.65a-d | 97.38 ± 0.83abc | 63.44 ± 1.51a-d | ||
| 60 | 96.25 ± 0.95ab | 98.13 ± 0.47a | 63.44 ± 0.29a-d | ||
| 180 | 93.75 ± 1.93a-e | 96.38 ± 1.46a-e | 62.08 ± 1.63a-h | ||
| 300 | 90.25 ± 5.19b-i | 94.63 ± 2.55a-h | 63.76 ± 0.81a-d | ||
| 420 | 94.00 ± 2.45a-e | 96.00 ± 1.78a-f | 63.76 ± 1.37a-d | ||
| Certified | 0 | 85.50 ± 1.26f-m | 92.75 ± 0.63a-k | 43.63 ± 1.03u | |
| 30 | 83.00 ± 2.92i-o | 91.50 ± 1.46b-l | 51.76 ± 2.21p-t | ||
| 60 | 83.00 ± 2.92i-o | 91.50 ± 1.46b-l | 54.21 ± 1.69k-s | ||
| 180 | 76.50 ± 2.33op | 82.75 ± 2.38n | 48.89 ± 1.31st | ||
| 300 | 78.00 ± 2.16nop | 86.50 ± 2.25lmn | 48.91 ± 1.29st | ||
| 420 | 85.50 ± 1.26f-m | 92.75 ± 0.63a-k | 52.49 ± 1.72n-t | ||
| Williams | Registered | 0 | 83.25 ± 1.38i-o | 89.63 ± 0.69 g-m | 59.73 ± 4.74b-k |
| 30 | 96.00 ± 1.29abc | 97.50 ± 0.91ab | 64.63 ± 1.06abc | ||
| 60 | 96.00 ± 1.87abc | 97.50 ± 1.40ab | 64.26 ± 1.59abc | ||
| 180 | 96.75 ± 0.48ab | 97.88 ± 0.66a | 65.01 ± 0.41abc | ||
| 300 | 96.75 ± 1.31ab | 98.38 ± 0.66a | 65.31 ± 0.90ab | ||
| 420 | 95.25 ± 2.59a-d | 96.13 ± 2.75a-e | 64.76 ± 1.27abc | ||
| Certified | 0 | 85.50 ± 1.26f-m | 92.75 ± 0.63a-k | 54.25 ± 1.73k-s | |
| 30 | 95.50 ± 1.04a-d | 97.75 ± 0.52a | 63.76 ± 1.11a-d | ||
| 60 | 96.75 ± 0.75ab | 98.38 ± 0.38a | 64.19 ± 0.63abc | ||
| 180 | 96.50 ± 0.87ab | 97.75 ± 0.92a | 64.76 ± 0.56abc | ||
| 300 | 94.75 ± 1.93a-d | 96.88 ± 1.46a-d | 63.34 ± 1.46a-e | ||
| 420 | 93.75 ± 1.31a-e | 95.88 ± 1.16a-f | 63.24 ± 1.00a-e |
The data were expressed as the mean ± standard error of mean (Se). Different letters within a column indicate significant differences as determined by the Duncan test (P ≤ 0.01) and (P ≤ 0.05)
Germination Rate (GR)
The results demonstrated a significant improvement in GR after plasma treatment (P ≤ 0.05). The results indicated that the highest germination rate was 98.13 ± 0.47%, while the lowest was 71.88 ± 2.05%, both associated with Arian-Certified in the control treatment (Table 3). The lowest and highest percentages of increase in germination speed compared to all control cultivars were 1.78% and 32.17%, respectively, associated with Katoul-Certified and Arian-Certified treatments at the 420th and 60th s. Sari-Certified (0.93) exhibited the slowest germination speed, requiring 420 s for treatment, while Arian-Registered demonstrated the fastest rate at 6.65% treatment efficiency.
Germination index (GI)
The findings indicated a notable enhancement in GI following plasma treatment (P ≤ 0.05). The highest GI index, exhibiting a significant difference, was associated with plasma treatment durations of 60 and 180 s for Williams and Sari cultivars. The control treatment in the Arian report exhibited the lowest GI, which was statistically significant. The cultivars exhibited a germination index ranging from 66.19 ± 1.03% to 37.01 ± 1.31%, with the latter value associated with Arian-Certified in the control treatment (Table 3). The seeds of Registered Arian and Certified Arian exhibited the lowest and highest percentage increases in the GI, at 3.6% and 60.09%, respectively, following 300- and 60-s treatments compared to the control. The lowest decrease in germination index, recorded at 2.44%, was observed in Arian after 420 s of plasma treatment, relative to the control group.
Seedling growth properties
Root length
The findings indicated that the Katoul cultivar exhibited the greatest root length, with a statistically significant difference observed in plasma treatment durations of 60 and 180 s. The control group in the Sari cultivar exhibited the lowest root length, demonstrating a significant difference. The minimum and maximum root lengths measured were 10.20 ± 0.14 cm and 21.17 ± 0.17 cm, respectively (Table 4). The lowest and highest percentages of root length increase relative to all control cultivars were 2.87% and 56.13%, attributed to Certified-Williams and Katoul-Registered, respectively, under cold plasma treatment for 420 and 180 s. Given that five cultivars were employed across two distinct classes of soybean seeds, it appears improbable that the variation in root length range can be attributed to genetic differences among the seeds.
Table 4.
Effect of cold plasma on seeding properties in soybean
| Cultivar | Class (within cultivar) | Plasma Treatment | Root length (cm) | Seedling length (cm) | Seedling dry weight (gr) |
|---|---|---|---|---|---|
| Sari | Registered | 0 | 10.20 ± 0.14z | 23.52 ± 0.45y | 1.07 ± 0.08y |
| 30 | 14.67 ± 0.13o-s | 34.35 ± 75n-u | 1.17 ± 0.03u-y | ||
| 60 | 13.92 ± 0.35q-v | 34.17 ± 0.52p-u | 1.16 ± 0.03u-y | ||
| 180 | 14.87 ± 0.38o-r | 34.57 ± 0.61 m-u | 1.35 ± 0.02k-u | ||
| 300 | 13.75 ± 0.86q-v | 30.75 ± 0.63vw | 1.08 ± 0.04xy | ||
| 420 | 11.47 ± 0.54y | 27.25 ± 0.37x | 1.13 ± 0.07v-y | ||
| Certified | 0 | 12.1 ± 0.12w-y | 22.57 ± 0.5y | 1.27 ± 0.04p-y | |
| 30 | 14.02 ± 0.25p-v | 34.22 ± 0.22q-u | 1.22 ± 0.03r-y | ||
| 60 | 15.17 ± 0.22 n-q | 36.37 ± 0.46k-p | 1.36 ± 0.04k-u | ||
| 180 | 14.97 ± 0.22 o-r | 35.10 ± 0.31 m-t | 1.22 ± 0.03r-y | ||
| 300 | 14.00 ± 0.32q-v | 34.72 ± 0.39 m-u | 1.22 ± 0.03r-y | ||
| 420 | 13.67 ± 0.28r-n | 33.17 ± 0.19s-v | 1.22 ± 0.03r-y | ||
| Saba | Registered | 0 | 14.87 ± 0.94o-r | 22.97 ± 0.69y | 1.22 ± 0.03r-y |
| 30 | 16.82 ± 0.26i-l | 36.00 ± 0.37 L-r | 1.24 ± 0.05q-y | ||
| 60 | 21.17 ± 0.17a | 40.92 ± 0.75c-f | 1.32 ± 0.02 m-w | ||
| 180 | 20.60 ± 0.34ab | 40.15 ± 0.35d-g | 1.35 ± 0.05 L-u | ||
| 300 | 20.52 ± 0.62ab | 39.57 ± 0.39d-i | 1.16 ± 0.09u-y | ||
| 420 | 20.15 ± 0.65abc | 39.47 ± 0.69d-i | 1.28 ± 0.06o-x | ||
| Certified | 0 | 11.87 ± 0.15xy | 32.50 ± 0.81t-w | 1.23 ± 0.04q-y | |
| 30 | 13.30 ± 0.17s-w | 32.22 ± 0.14r-v | 1.20 ± 0.08t-y | ||
| 60 | 14.87 ± 0.21o-r | 34.60 ± 0.42 m-u | 1.31 ± 0.04 m-w | ||
| 180 | 13.87 ± 0.21q-v | 34.82 ± 0.16 m-t | 1.29 ± 0.03n-w | ||
| 300 | 13.62 ± 0.17r-w | 34.32 ± 59n-u | 1.21 ± 0.02s-y | ||
| 420 | 13.30 ± 0.18s-w | 33.32 ± 0.42q-v | 1.11 ± 0.05wxy | ||
| Arian | Registered | 0 | 12.60 ± 0.29v-y | 22.05 ± 0.37y | 1.28 ± 0.050-x |
| 30 | 15.75 ± 0.60 L-o | 34.50 ± 1.04 m-u | 1.62 ± 0.10b-h | ||
| 60 | 14.25 ± 0.09p-u | 32.76 ± 0.29t-v | 1.54 ± 0.04c-l | ||
| 180 | 14.70 ± 0.12o-s | 31.97 ± 0.23 m-u | 1.35 ± 0.07 L-u | ||
| 300 | 14.03 ± 0.09 p-t | 30.77 ± 0.07vw | 1.39 ± 0.08i-t | ||
| 420 | 14.00 ± 0.03q-v | 29.97 ± 0.14w | 1.30 ± 0.06 m-w | ||
| Certified | 0 | 13.7 ± 0.46r-v | 22.00 ± 0.19y | 1.31 ± 0.03 m-w | |
| 30 | 18.52 ± 0.25d-h | 36.70 ± 0.22j-p | 1.65 ± 0.09a-f | ||
| 60 | 18.97 ± 0.13c-f | 37.10 ± 0.14 h-n | 1.62 ± 0.11b-g | ||
| 180 | 19.75 ± 0.20bcd | 37.87 ± 0.12 g-l | 1.67 ± 0.09a-e | ||
| 300 | 18.22 ± 0.21e-h | 36.00 ± 0.24 L-r | 1.58 ± 0.10c-i | ||
| 420 | 17.77 ± 0.35fk | 35.90 ± 0.18 L-s | 1.60 ± 0.08b-h | ||
| Katoul | Registered | 0 | 12.65 ± 0.66v-y | 30.82 ± 0.61vw | 1.31 ± 0.03 m-w |
| 30 | 16.42 ± 0.46k-n | 40.55 ± 0.85c-g | 1.47 ± 0.11f-o | ||
| 60 | 18.02 ± 0.67e-i | 41.95 ± 0.51b-e | 1.61 ± 0.05b-h | ||
| 180 | 19.75 ± 0.92bcd | 43.94 ± 0.94abc | 1.73 ± 0.03abc | ||
| 300 | 18.37 ± 1.13d-h | 42.15 ± 0.52bcd | 1.79 ± 0.08ab | ||
| 420 | 17.62 ± 0.61 g-k | 39.35 ± 0.47e-i | 1.50 ± 0.11d-m | ||
| Certified | 0 | 16.50 ± 0.37 j-n | 30.77 ± 0.42vw | 1.38 ± 0.03j-t | |
| 30 | 18.85 ± 0.89c-h | 39.82 ± 0.52d-h | 52.35 ± 0.04a | ||
| 60 | 18.80 ± 0.10c-h | 39.22 ± 0.15f-i | 1.74 ± 0.04abc | ||
| 180 | 19.30 ± 0.44bcd | 40.17 ± 0.97d-g | 1.69 ± 0.04a-d | ||
| 300 | 18.68 ± 0.50d-h | 38.35 ± 0.58f-l | 1.73 ± 0.07abc | ||
| 420 | 17.47 ± 0.47 h-k | 36.02 ± 0.29 L-q | 1.69 ± 0.03a-d | ||
| Williams | Foundation | 0 | 16.00 ± 0.34 L-o | 31.42 ± 1.02vw | 1.31 ± 0.06 m-w |
| 30 | 16.80 ± 0.16i-m | 44.03 ± 0.18ab | 1.49 ± 0.04e-n | ||
| 60 | 18.65 ± 0.31d-h | 44.50 ± 0.30ab | 1.61 ± 0.04b-h | ||
| 180 | 19.1 ± 0.37c-f | 45.20 ± 0.82a | 1.42 ± 0.10 g-r | ||
| 300 | 18.45 ± 0.53d-h | 42.90 ± 0.92abc | 1.47 ± 0.04f-p | ||
| 420 | 17.85 ± 0.42f-j | 40.60 ± 1.23c-g | 1.57 ± 0.06c-j | ||
| Certified | 0 | 12.82 ± 0.43u-y | 30.62 ± 0.39vw | 1.23 ± 0.03q-y | |
| 30 | 15.45 ± 020 m-p | 39.05 ± 0.40f-k | 1.55 ± 0.03c-k | ||
| 60 | 14.2 ± 0.21p-u | 37.25 ± 0.16 h-m | 1.43 ± 0.06 g-q | ||
| 180 | 13.80 ± 0.22q-v | 37.00 ± 0.15i-o | 1.58 ± 0.04c-i | ||
| 300 | 13.32 ± 0.15s-w | 37.27 ± 0.16 h-m | 1.46 ± 0.05f-p | ||
| 420 | 13.90 ± 0.16t-x | 37.10 ± 0.17 h-n | 1.41 ± 0.05 h-s |
Seedling length
The findings indicated that the Williams and Katoul cultivars exhibited the greatest seedling length, with a statistically significant difference, following plasma treatment at 30, 60, and 180 s. The control treatment exhibited the lowest seedling length with a statistically significant difference in both Sari and Arian cultivars. The minimum and maximum seedling lengths were reported as 22.00 ± 0.19 cm and 45.20 ± 0.82 cm, respectively (Table 5). The lowest and highest percentages of seedling length increases, 2.7% and 78.13% respectively, compared to the control group, were observed in Saba-Certified and Saba-Registered at 30 and 60 s of treatment.
Table 5.
Results of analysis of variance for catalase enzyme (CAT), superoxide dismutase (SOD), and ascorbate peroxidase (APX)
| Means of Square | ||||
|---|---|---|---|---|
| S.O.V | df | CAT | SOD | APX |
| Time | 5 | 33827.191** | 24030.046** | 28339.375** |
| Cultivar | 4 | 8173.209** | 3072.797** | 851.359** |
| Cluster | 1 | 3563.15** | 1841.181** | 6188.065** |
| Time × Cultivar | 20 | 370.274** | 304.585** | 173.278** |
| Cultivar × Cluster | 4 | 2564.238** | 233.108** | 314.454** |
| Time × Cluster | 5 | 1142.240** | 208.07** | 145.001** |
| Time × Cultivar × Cluster | 20 | 432.366** | 21.678** | 92.415** |
| Error | 180 | 14.452 | 15.216 | 7.632 |
| CV (%) | - | 6.25 | 8.58 | 4.99 |
**: significant at 1% probability level
Seedling dry weight
The results indicated that the highest seedling dry weight was observed in the 30, 60, and 180 s plasma treatments for the Williams and Katoul cultivars. The control treatment exhibited the lowest seedling dry weight in both Sari and Saba cultivars. The minimum and maximum seedling dry weights recorded were 1.07 ± 0.08 g and 1.83 ± 0.04 g, respectively (Table 4). The lowest and highest percentages of seedling dry weight increase were observed in the 30- and 300-s treatments, respectively, compared to the control group (1.4% and 36.6%). These increases were associated with the Registered-Sari and Registered-Katoul classes (Table 4). The Saba-Certified exhibited the lowest and highest percentages of seedling dry weight loss at 1.47% and 9.61%, respectively, observed during the 300 and 420 s of treatment.
The activity of antioxidant enzymes
Catalase enzyme (CAT)
The results indicated that the highest CAT activity was observed in the Katoul cultivar at the 60 s treatment, whereas the lowest values were recorded in the Saba- Certified, Arian- Registered, and Arian- Certified cultivars following 300 and 420 s of plasma treatment. The range of CAT activities varied from − 0.88 to 4.40 times greater than the control condition. The findings indicated that the CAT varied from 20.19 ± 0.76 to 148.97 ± 0.51 µM g-1 FW min− 1 (Table 6). The results indicate that Sari-Registered exhibited the highest CAT activity at the 60 s mark of treatment compared to the control group. Conversely, the lowest values were observed in Saba-Certified, Arian-Registered, and Arian-Certified at the 0, 300, and 420 s intervals of plasma treatment, respectively. The antioxidant activity of catalase varied from − 0.88 to 4.40 times greater than that of all control cultivars.
Table 6.
Effect of cold plasma on the activity of antioxidant enzymes in soybean
| Cultivar | Class (within cultivar) | Plasma Treatment | CAT activity (µM g-1 FW min-1) |
SOD activity (µM g-1 FW min-1) |
APX activity (µM g-1 FW min-1) |
|---|---|---|---|---|---|
| Sari | Registered | 0 | 33.82 ± 0.7v | 17.99 ± 0.49tuv | 36.42 ± 1.02np |
| 30 | 130 ± 1.1c | 72.86 ± 0.94ef | 96.03 ± 0.7ab | ||
| 60 | 148.97 ± 0.51a | 77.48 ± 0.58cde | 97.48 ± 0.81a | ||
| 180 | 133.72 ± 1.06b | 73.47 ± 0.78def | 95.31 ± 0.80ab | ||
| 300 | 34.6 ± 0.78v | 16.4 ± 0.48uv | 36.45 ± 0.84np | ||
| 420 | 31.89 ± 0.62 ± | 15.40 ± 0.42vw | 33.47 ± 0.40o-r | ||
| Certified | 0 | 39.07 ± 0.44u | 10.65 ± 0.41wx | 27.15 ± 0.94uvw | |
| 30 | 75.45 ± 0.27klm | 56.90 ± 0.28 lm | 76.98 ± 0.69e-j | ||
| 60 | 79.69 ± 0.69j | 62.76 ± 0.37 h-k | 79.81 ± 0.47e | ||
| 180 | 74.14 ± 0.16 lm | 57 ± 1.15 lm | 77.87 ± 0.62e-j | ||
| 300 | 39.75 ± 1.44tu | 10.75 ± 0.67wx | 26.58 ± 0.63vw | ||
| 420 | 39.53 ± 1.27tu | 9.11 ± 1.61x | 24.98 ± 0.56w | ||
| Saba | Registered | 0 | 39.24 ± 0.41u | 20.00 ± 0.67s-v | 54.8 ± 0.89 m |
| 30 | 72.45 ± 0.84mn | 58.19 ± 1.03jkl | 94.78 ± 0.54ab | ||
| 60 | 78.15 ± 0.50jk | 60.03 ± 0.86i-l | 93.97 ± 0.59b | ||
| 180 | 76.89 ± 0.17jkl | 58.46 ± 0.29jkl | 95.16 ± 0.65ab | ||
| 300 | 38.71 ± 1.17u | 22.58 ± 0.87r-t | 21.93 ± 1.12x | ||
| 420 | 38.83 ± 0.73u | 21.80 ± 0.78r-u | 25.30 ± 0.55w | ||
| Certified | 0 | 22.32 ± 0.32w | 24 ± 0.47rs | 33.84 ± 0.62o-q | |
| 30 | 61.87 ± 0.5q | 51.38 ± 1.10 m | 74.77 ± 0.55f-k | ||
| 60 | 67.10 ± 0.52op | 57.27 ± 0.72kl | 77.31 ± 0.36e-l | ||
| 180 | 62.55 ± 0.5q | 51.53 ± 0.83 m | 73.31 ± 1.15jk | ||
| 300 | 22.78 ± 1.47w | 27.20 ± 0.77r | 32.94 ± 1.29p-s | ||
| 420 | 21.86 ± 1.85w | 25.32 ± 0.81r-u | 21.53 ± 1.17x | ||
| Arian | Registered | 0 | 22.93 ± 0.97w | 17.06 ± 0.72uv | 35.61 ± 0.79np |
| 30 | 74.66 ± 1.94 lm | 63.58 ± 0.977 h-j | 74.41 ± 0.85f-k | ||
| 60 | 78.13 ± 1.08jk | 68.17 ± 0.62fgh | 75.89 ± 0.48f-j | ||
| 180 | 74.61 ± 0.94 lm | 62.98 ± 0.87 h-k | 74.18 ± 0.43 h-k | ||
| 300 | 23.23 ± 1.40w | 20.96 ± 1.02s-v | 35.87 ± 0.49np | ||
| 420 | 20.19 ± 0.76w | 21.09 ± 0.64s-v | 31.48 ± 0.90q-t | ||
| Certified | 0 | 21.12 ± 0.67w | 16.4 ± 0.83uv | 29.79 ± 1.45s-v | |
| 30 | 64.77 ± 0.71pq | 41.84 ± 0.25n | 72.13 ± 0.85k | ||
| 60 | 69.94 ± 0.5no | 64.43 ± 1.4 g-i | 77.94 ± 0.5ef | ||
| 180 | 64.74 ± 1.52pq | 54.25 ± 0.39 lm | 73.49 ± 0.45jk | ||
| 300 | 22.28 ± 0.92w | 18.21 ± 0.36tuv | 30.19 ± 1.49p-u | ||
| 420 | 21.12 ± 1.2w | 18.02 ± 0.75tuv | 31.48 ± 0.9q-t | ||
| Katoul | Registered | 0 | 51.90 ± 0.9r | 20.33 ± 0.70s-v | 36.63 ± 0.42no |
| 30 | 114.61 ± 1.62e | 80.6 ± 1.09bc | 74.30 ± 1.083 g-k | ||
| 60 | 118.05 ± 1.09d | 87.91 ± 1.32a | 77.73 ± 0.32e-h | ||
| 180 | 114.45 ± 1.87e | 82.30 ± 0.74bc | 74.83 ± 0.76f-k | ||
| 300 | 54.03 ± 0.86r | 25.75 ± 0.93rs | 37.01 ± 2.13no | ||
| 420 | 53.10 ± 0.48r | 24.39 ± 1.16rs | 38.02 ± 1.07n | ||
| Certified | 0 | 44.28 ± 0.99s | 21.33 ± 0.71r-v | 16.31 ± 0.82y | |
| 30 | 95.57 ± 0.59 g | 77.44 ± 0.45cde | 62.77 ± 0.52 L | ||
| 60 | 99.39 ± 1.0f | 85.2 ± 0.43ab | 65.52 ± 0.40 L | ||
| 180 | 95.37 ± 0.65 g | 79.64 ± 1.35bc | 63.40 ± 0.75 L | ||
| 300 | 45.08 ± 1.27s | 25.23 ± 0.79rs | 19.64 ± 1.38x | ||
| 420 | 42.47 ± 0.77st | 24.49 ± 1.03rs | 19.29 ± 1.39xy | ||
| Williams | Registered | 0 | 33.26 ± 1.36v | 35.26 ± 0.68np | 35 ± 26 ± np |
| 30 | 84.24 ± 0.65kl | 86.03 ± 0.32d | 86.03 ± 0.32d | ||
| 60 | 90.14 ± 0.93j | 89.65 ± 0.29c | 89.26 ± 0.29c | ||
| 180 | 86.02 ± 1.16kl | 87.93 ± 0.46 cd | 87.93 ± 0.46 cd | ||
| 300 | 34.33 ± 0.85v | 36.15 ± 1.42np | 36.15 ± 1.42np | ||
| 420 | 32.74 ± 0.72v | 35.04 ± 1.03np | 35.04 ± 1.03np | ||
| Certified | 0 | 34.70 ± 0.69v | 32.79 ± 1.33q | 28.9 ± 0.21tuw | |
| 30 | 75.93 ± 1.21i | 67.22 ± 1.73gh | 73.96 ± 0.51ijk | ||
| 60 | 79.54 ± 1.2 h | 69.87 ± 1.07 fg | 75.78 ± 0.84f-j | ||
| 180 | 76.13 ± 1.83i | 66.87 ± 0.35gh | 73.4 ± 0.18jk | ||
| 300 | 34.63 ± 0.70v | 35.12 ± 1.34opq | 26.69 ± 2.32vw | ||
| 420 | 33.44 ± 0.92v | 34.14 ± 0.61pq | 29.83 ± 1.07 |
The data were expressed as the mean ± standard error of mean (Se). Different letters within a column indicate significant differences as determined by the Duncan test (P ≤ 0.01)
Superoxide dismutase (SOD)
The findings indicated that the highest SOD activity was associated with the 60 s treatment of the Williams cultivar. Conversely, the lowest SOD activity, with a significant difference, was observed in the Arian and Saba cultivars during the 420 s treatment and control groups. The SOD activity varied from 9.11 ± 1.16 to 87.91 ± 1.32 µMg− 1 FW min− 1 across all samples (Table 6). Additionally, the highest SOD activity was recorded in Katoul-Registered, Katoul-Certified, and Williams-Registered under the 60 s treatment compared to control cultivars, while the Sari-Certified treatments at 0, 300, and 420 s exhibited the lowest SOD activity. An increase in SOD activity was noted, ranging from 0.86 to 5.89 times greater than that of all control cultivars.
Ascorbate peroxidase (APX)
The findings indicated that the Sari and Katoul cultivars exhibited the highest and lowest APX values, respectively, observed at 60 and 420 s of treatment. The APX values varied from 16.31 ± 0.82 to 97.84 ± 0.81 µMg− 1 FW min− 1 across all samples (Table 6). The highest levels of APX were observed in Sari-Registered and Saba-Registered across the treatments of 30, 60, and 180 s. The lowest antioxidant activity of ascorbate peroxidase was observed in Katoul-Certified during the control at 420 s of treatment. The antioxidant activity of APX was reported to be approximately 0.40 to 4.01 times greater than that of all control cultivars.
Optical emission spectroscopy
The results indicated that the species identified in the optical emission spectrum included OH (309 nm), N2I (380 nm), N2II (405.79 nm), oxygen atoms (755 and 842 nm), and argon (from 696 to 861.8 nm), as represented by the horizontal bar in Fig. 2. The results indicated that the N2 generated by Plasma significantly enhanced biological processes in seeds.
Fig. 2.
Optical emission spectra of DBD plasma
Scanning Electron microscope (SEM)
The results indicated that the maximum and minimum morphological characteristics of the seed surface were observed at 60 and 420 s, respectively. Results indicated that there were significant differences in the levels of seeds treated for 420 s when compared to those treated for 60 s and the untreated seeds. The seed structure exhibited minor alterations at 60 s relative to the control group. Significant changes in the level of treated seeds are observed in SEM images at 420 s. The results indicated that plasma significantly altered the morphological properties during the 420 s treatments (Fig. 3). The growth and germination parameters appeared to be influenced by the duration of plasma treatment and the type of plasma discharge. A reduced duration correlates with enhanced outcomes for seed germination.
Fig. 3.
SEM images of the seeds surface at different scales, before treatment (a-c), after 60 s plasma treatment (d-f), and after 420 s plasma treatment (g-i)
Discussion
This study demonstrated that cold plasma significantly influenced soybean seed germination. The findings indicated that cold plasma treatment enhanced antioxidant activity and promoted soybean seed germination. The cold plasma technique has the potential to modify surface roughness and chemical structure, resulting in significant alterations to the seed surface that may influence water uptake and enhance germination (Fig. 2). The current research demonstrated that cold plasma treatment significantly increased seed weight, consistent with the findings of Šerá et al., 2021 [27], Guo et al., 2018 [28], and Ling et al., 2014 [7].
Plasma treatment durations of 60 s and 180 s enhanced germination potential, germination index, and germination rate relative to the control group. Ling et al. (2014) observed a significant increase in germination potential, with a rise of 14.7% relative to the control treatment, following 15 s of 80 W plasma treatment [29]. Li et al. (2017) demonstrated that the application of dielectric-barrier plasma in an air atmosphere for 7 min resulted in a 26.7% increase in germination potential in wheat following 7 min of DBD plasma treatment [30]. Fereydoni and Alizadeh (2022) found that the germination percentage potential of chickpea cultivars increased by 2% following 30 s of plasma treatment and by 11% after 60 s, relative to the control treatment [31]. Additionally, a study conducted by Pizá et al. (2018) investigated the exposure of soybeans to dielectric-barrier plasma at atmospheric pressure and a frequency of 50 Hz, utilizing nitrogen and oxygen gas sources for durations of 1, 2, and 3 min. A significant stimulating effect on germination potential was observed. The findings of these studies were consistent with the current research [16]. This study observed a maximum percentage increase in germination potential of 66.97% at treatment durations of 30, 60, and 180 s, in comparison to all control treatment figures, indicating a notable enhancement.
Švubová et al. (2021) reported germination inhibition following nitrogen plasma treatment for durations of 90 and 120 s [20]. In other treatments (30 and 60 s), a higher germination percentage was noted compared to the control [32], aligning with the findings of the current study. This study indicated that plasma treatment durations of 30, 60, and 180 s across all examined cultivars resulted in the highest germination potential. The germination potential decreased by 3.5% relative to the control treatment associated with Arian - Plasma treatment was registered, with a reported duration of 420 s, attributed to its destructive effects.
The impact of plasma irradiation duration on grain morphology was examined over 2- and 5-min intervals in a separate study. The results indicated an increase in seed pattern intensity, likely due to the removal of upper cuticle layers that may have been coated with wax during the 2 min plasma treatment. Extended exposure of seeds to plasma influenced the internal structure of the cuticle, resulting in damage or rupture in certain areas of the cuticle. Also, based on the results, a 5-min plasma treatment exhibited a destructive effect [33].
The SEM images obtained in this study indicated significant alterations in the surface of seeds subjected to cold plasma treatment for 60 s, resulting in the formation of cracks and porosity. These changes significantly influenced the water penetration of the seeds when compared to the control treatment. This results in enhanced seed germination. The results of the plasma treatment at 420 s demonstrated significant alterations in seed morphology, specifically evidenced by cracking in portions of the seed coat. Plasma treatment lasting less than 420 s significantly affected germination, growth, and enzyme activity traits. Increased plasma irradiation duration results in a higher formation of reactive nitrogen species (RNS) and reactive oxygen species (ROS) [34–37].
Extending the exposure of seeds to plasma may yield diverse outcomes. More significant alterations in the seed coat, particularly in the regions where it is detached from the peripheral sides, are observed. In certain cases, particularly at an exposure time of 420 s, seed splitting and significant changes in seed coat morphology are noted. Plasma interacts with biological material in various ways [38]. Excessive exposure likely results in structural degradation, volatilization, and cellular etching. Consequently, an increase in exposure time correlates with a greater overall effect. The alterations in the surface structure of seeds are likely attributable to the effects of gases produced, such as NO2, O3, and NO, as prolonged plasma irradiation may induce nitrogen stress and excessive oxidative conditions [39]. This may explain the reduced germination energy and capacity observed after the extended exposure of seeds to plasma.
Li et al. (2017) demonstrated that the application of dielectric barrier plasma in an air atmosphere for 7 min resulted in a 9.1% increase in wheat germination rates [40]. Švubová et al. (2021) subjected soybean seeds to plasma treatment in oxygen, nitrogen, and ambient atmospheres for durations of 30, 60, 90, and 120 s. The rate of seed germination increased by 20% in all treatments except the nitrogen plasma treatment at 90 and 120 s. This is likely because the plasma improved the activity of enzymes that are important in the early stages of germination [20]. The current research demonstrated that the germination rate increased by up to 32.17% in the 60-s treatment, representing the highest rate among all control cultivars. This represents a notable increase relative to the outcomes observed in other experiments. Additionally, the electron microscope images in the studies by Hosseini et al. (2018) demonstrated that plasma treatment induced micrometric fractures in the hard coating of artichoke, which ultimately enhanced the germination rate by increasing water absorption and facilitating the breaking of the seed shell [41]. Gómez-Ramírez et al. (2017) posited that the observed increase in germination rate may be attributed to plasma etching on the seed coat, which disrupts the macromolecules present on the seed surface. This disruption facilitates the release of nitrate functional groups from the seed coat into the seed [42].
The mechanism for enhancing germination rates is associated with the seed surface structure, the capacity to rupture the seed coat, water absorption, and the release of chemically active species, such as reactive oxygen species (ROS), generated by atmospheric cold plasma through the seed shell. The absorption of water by the seed facilitates the movement of gibberellic acid hormones to the aleurone layer of the endosperm, resulting in enhanced expression of the alpha-amylase enzyme. The resulting alpha-amylase hydrolyzes nutrients stored in the endosperm, including starch, into sugars. The sugar generated as an energy source enhances the growth of the embryo during the germination phase [43]. Li et al. (2017) observed that exposure of dielectric barrier plasma to air for 7 min resulted in a 16.9% increase in the germination index of wheat [40]. Guo et al. (2018) observed a 13.9% enhancement in the germination index of wheat plants subjected to an 11 kV treatment for 4 min, relative to the control group [28]. The results of the present study indicated that the highest percentage increase in the germination index among various cultivars reached 60.09% after 60 s of treatment, in comparison to the control figures, representing a significant enhancement.
The oxidation processes of reactive plasma species enhance water absorption capacity by improving the wettability of the seed coat. They may also be associated with gas exchange and electrolyte leakage in the seed. Cold plasma may effectively alter the dormancy of rigid seeds by influencing seed permeability and initiating subsequent processes. Cold plasma positively influences seed germination and growth, subsequently enhancing seedling traits. It can reduce the stiffness associated with mechanical dormancy in dark legume species, including blue lupine, alfalfa, green peas, Mimosa sp., and Trifolium sp. Cold plasma can be employed to disinfect the surfaces of plant seeds and leguminous products. Legumes exhibit a high tolerance to the specified physicochemical treatment, and the mild tension generated by plasma has a beneficial effect on them. Changes in physiological factors can enhance both the quantity of plants in the field and their performance [44].
Multiple studies have reported a significant increase in root length following cold plasma treatment in soybean, wheat, sesame, and chickpea, consistent with the findings of this study [45–47]. The current investigation revealed a significant increase in average root length across all cold plasma treatments when compared to the control measurements. Ling et al. (2014) observed that a 15-s treatment of 80 W cold plasma resulted in a notable increase in soybean seedling size by 13.8% relative to the control group [29]. Studies have documented a notable increase in seedling length following cold plasma treatment in wheat [30, 48, 49], Catharanthus roseus [50], and sesame [51], which aligns with the findings of the current study. Filatova et al. (2014) indicated that molecular nitrogen generated by plasma enhances root length, seedling length, and enzyme activity. Studies have demonstrated that plasma significantly boosts the oxidation of chemical compounds in seed shells by producing oxygen molecules, which in turn stimulates biochemical processes that aid in seed germination [52]. This study observed the highest percentage increase in seedling length, reaching 78.13% compared to the control at the 60-s treatment mark. Ling et al. (2014) observed that, relative to the control group in soybeans, an 80 W cold plasma treatment for 15 s resulted in a significant increase in shoot dry weight by 21.9% [7]. Li et al. (2017) observed that exposure to dielectric-barrier plasma in an air atmosphere for 7 min improved both the fresh weight and dry weight of wheat seedlings [40]. The findings indicated that the 300-s treatment resulted in the highest percent increase in dry-weight seedlings, measuring 36.6% relative to the control group. Zhang et al. (2018) observed an enhancement in peroxidase and catalase activity following DBD-argon plasma treatment of soybean seeds [53].
The findings of Pizá et al. (2018) indicated that plasma treatment of soybeans using various gas sources (nitrogen and oxygen) for durations of 1, 2, and 3 min resulted in a 29% reduction in catalase activity in healthy seeds, while superoxide dismutase activity showed no significant difference between treated and control seeds. The findings indicated that the low levels of catalase activity, glutathione content, and lipid peroxidation in plasma-treated seeds suggest that hydrogen peroxide (H2O2) generated during plasma treatment did not stimulate the antioxidant defense mechanisms nor inflict damage on the cell membrane. Hydrogen peroxide that is produced can function as a signaling molecule in the induction of germination [16]. In 2021, Švubová et al. did a study on cumin seeds that were treated with cold plasma for 5 and 10 min. They found that there were no significant differences in the levels of catalase enzymes and superoxide dismutase between control plants and those that were treated with plasma for 5 min. However, the enzymes ascorbate peroxidase and glutathione reductase exhibited a significant reduction in comparison to the control group. A 10-min plasma treatment resulted in an increase in superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase enzymes compared to the control [20]. A study by Hosseini et al. (2018) investigated the effects of cold plasma on soya utilized various gas sources (nitrogen, air, and oxygen) for durations of 30, 60, 90, and 120 s. They showed that the activity of superoxide dismutase increased following all plasma treatments, with the exception of nitrogen plasma at 90 and 120 s, when compared to the control group [41]. Rasooli et al. (2021) demonstrated that a 5-min plasma treatment for seed pretreatment enhanced nutrient uptake and increased the germination index, seedling height, root length, and total dry weight of cumin seeds. The results also showed that priming seeds with plasma for 10 min increased the activities of SOD, CAT, APX, and glutathione reductase enzymes compared to the control group, which is similar to the results of this study [54]. This study demonstrated that cold plasma treatment for durations of 60 and 180 s significantly influenced seed germination characteristics and seedling development. Furthermore, the plasma treatment significantly influenced the activity of SOD, ranging from 0.86 to 5.89, in contrast to CAT, which ranged from 0.88 to 4.40, and APX, which ranged from 0.40 to 4.01, at 60 s. In the 420-s treatment, the activity of all three enzymes studied exhibited a significant decrease relative to the control group.
Extended exposure duration of CP leads to progressive seed damage. This suggests that the agricultural sector should consider CP treatment as an innovative strategy. CP serves as an effective treatment for seed stimulation and also enhances the plant’s defense system against diseases. Recently, plant pathologists have encountered challenges in creating alternative, chemical-free methods for the control of postharvest fungi at a commercial scale. CP represents a promising novel postharvest treatment, exhibiting no known adverse effects on fresh crops or the environment while effectively controlling bacterial infections that pose food safety risks [55].
Conclusion
The results of the present study indicate that cold plasma seed priming enhanced GP, GR, GI, and various growth characteristics, including root length, seedling length, and seedling dry weight, in comparison to the control sample. Furthermore, it positively influenced the activity of CAT, SOD, and AXP enzymes. This study yielded three novel findings: Different cultivars and classes within cultivars exhibited notable responses to the cold plasma pretreatment. The variation in plasma treatment duration resulted in notable differences in seedling growth characteristics and seed germination rates. The treatments lasting 60 and 180 s demonstrated the most favorable impact on the examined traits. Cold plasma, when applied within an optimal time frame, is an effective method for enhancing seed germination and seedling development in agricultural crops. Priming soy seeds with cold plasma may represent an effective strategy for mitigating environmental stress in soybean crops.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Acknowledgements
This work was supported by Seed and Plant Certification and Registration Institute (SPCRI) and Plasma Physics Research Center of Azad University, Research Sciences Branch, Tehran.
Author contributions
Khadijeh Sayahi conceived and carried out the experiments. Amir Hossein Sari, Aidin Hamidi, Bahareh Nowruzi, Farshid Hassani analyzed the data. Amir Hossein Sari, Aidin Hamidi, Bahareh Nowruzi, Farshid Hassani wrote the manuscript. All authors contributed to manuscript revision, read and approved the submitted version.
Funding
Not applicable.
Data availability
This research was done according to Ling et al. 2014, with the modification of seed number per replicate. All data generated or analysed during this study are included in this published article (supplementary information files: S1, S2, S3, S4 and S5).
Declarations
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
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Not applicable.
Ethics approval and consent for participant
All local, national or international guidelines and legislation, including Act of Plant Varieties Registration, Control and Plant Material of Islamic Republic of Iran were adhered to in registration of studied cultivars and their seeds production and certification to the production of this study.
The seeds of soybean varieties used in this research were obtained free of charge from the Seed and Plant Certification and Registration Institute (SPCRI) in Iran.
The research was conducted by use of 5 soybean improved common commercial cultivars entitled in Iran Registered Plant Varieties National List seeds which are certified by Iranian national plant varieties registration and seed certification focal point, Seed and Plant Certification and Registration Institute (SPCRI) based on the act of plant varieties registration, control and certification of seed and plant material Islamic Republic of Iran.
The studied soybean seeds were not collected, they were obtained from the research institute.
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References
- 1.Mansouri Daneshvar MR, Ebrahimi M, Nejadsoleymani H. An overview of climate change in Iran: facts and statistics. Environ Syst Res. 2019;8(1):1–10. [Google Scholar]
- 2.Majidian P, Ghorbani HR, Farajpour M. Achieving agricultural sustainability through soybean production in Iran: potential and challenges. Heliyon 2024, 10(4). [DOI] [PMC free article] [PubMed]
- 3.Soltani A, Gholipoor M, Zeinali E. Seed reserve utilization and seedling growth of wheat as affected by drought and salinity. Environ Exp Bot. 2006;55(1–2):195–200. [Google Scholar]
- 4.Zhang ShuJie ZS, Li Ling LL, Zhang ChunLei ZC. Li GuangMing LG: ALA altered ABA content of winter oilseed rape (Brassica napus L.) seedling. 2011.
- 5.Błaszczak W, Doblado R, Frias J, Vidal-Valverde C, Sadowska J, Fornal J. Microstructural and biochemical changes in raw and germinated cowpea seeds upon high-pressure treatment. Food Res Int. 2007;40(3):415–23. [Google Scholar]
- 6.Al-Bachir M. Effect of gamma irradiation on microbial load and sensory characteristics of aniseed (Pimpinella anisum). Bioresour Technol. 2007;98(10):1871–6. [DOI] [PubMed] [Google Scholar]
- 7.Ling L, Jiafeng J, Jiangang L, Minchong S, Xin H, Hanliang S, Yuanhua D. Effects of cold plasma treatment on seed germination and seedling growth of soybean. Sci Rep. 2014;4(1):5859. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Ucar Y, Ceylan Z, Durmus M, Tomar O, Cetinkaya T. Application of cold plasma technology in the food industry and its combination with other emerging technologies. Trends Food Sci Technol. 2021;114:355–71. [Google Scholar]
- 9.Randeniya LK, de Groot GJ. Non-thermal plasma treatment of agricultural seeds for stimulation of germination, removal of surface contamination and other benefits: a review. Plasma Processes Polym. 2015;12(7):608–23. [Google Scholar]
- 10.Benabderrahim MA, Bettaieb I, Hannachi H, Rejili M, Dufour T. Cold plasma treatment boosts barley germination and seedling vigor: insights into soluble sugar, starch, and protein modifications. J Cereal Sci. 2024;116:103852. [Google Scholar]
- 11.Sayahi K, Sari AH, Hamidi A, Nowruzi B, Hassani F. Application of cold argon plasma on germination, root length, and decontamination of soybean cultivars. BMC Plant Biol. 2024;24(1):59. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Guragain RP, Kierzkowska-Pawlak H, Fronczak M, Kędzierska-Sar A, Subedi DP, Tyczkowski J. Germination improvement of fenugreek seeds with cold plasma: exploring long-lasting effects of surface modification. Sci Hort. 2024;324:112619. [Google Scholar]
- 13.Bozhanova V, Marinova P, Videva M, Nedjalkova S, Benova E. Effect of Cold plasma on the Germination and Seedling Growth of Durum Wheat genotypes. Processes. 2024;12(3):544. [Google Scholar]
- 14.Mohandoss S, Mohan H, Balasubramaniyan N, Assadi AA, Loganathan S. Pearl Millet seed surface modification and improved germination by non-thermal plasma discharge: understanding the role of reactive species. Plasma Chem Plasma Process. 2024;44(2):1031–51. [Google Scholar]
- 15.Brandenburg R. Dielectric barrier discharges: progress on plasma sources and on the understanding of regimes and single filaments. Plasma Sources Sci Technol. 2017;26(5):053001. [Google Scholar]
- 16.Pizá MCP, Prevosto L, Zilli C, Cejas E, Kelly H, Balestrasse K. Effects of non–thermal plasmas on seed-borne Diaporthe/Phomopsis complex and germination parameters of soybean seeds. Innovative food Sci Emerg Technol. 2018;49:82–91. [Google Scholar]
- 17.Pérez-Pizá MC, Prevosto L, Grijalba PE, Zilli CG, Cejas E, Mancinelli B, Balestrasse KB. Improvement of growth and yield of soybean plants through the application of non-thermal plasmas to seeds with different health status. Heliyon. 2019;5(4):e01495. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 18.Jiang J, Lu Y, Li J, Li L, He X, Shao H, Dong Y. Effect of seed treatment by cold plasma on the resistance of tomato to Ralstonia solanacearum (bacterial wilt). PLoS ONE. 2014;9(5):e97753. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 19.Mildažienė V, Aleknavičiūtė V, Žūkienė R, Paužaitė G, Naučienė Z, Filatova I, Lyushkevich V, Haimi P, Tamošiūnė I, Baniulis D. Treatment of common sunflower (Helianthus Annus L.) seeds with radio-frequency electromagnetic field and cold plasma induces changes in seed phytohormone balance, seedling development and leaf protein expression. Sci Rep. 2019;9(1):6437. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 20.Švubová R, Slováková Ľ, Holubová Ľ, Rovňanová D, Gálová E, Tomeková J. Evaluation of the impact of cold atmospheric pressure plasma on soybean seed germination. Plants. 2021;10(1):177. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21.Xiao S, Liu L, Wang H, Li D, Bai Z, Zhang Y, Sun H, Zhang K, Li C. Exogenous melatonin accelerates seed germination in cotton (Gossypium hirsutum L). PLoS ONE. 2019;14(6):e0216575. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 22.Ling L, Jiafeng J, Jiangang L, Minchong S, Xin H, Hanliang S, Yuanhua D. Effects of cold plasma treatment on seed germination and seedling growth of soybean. Sci Rep. 2014;4(1):1–7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 23.Kar M, Mishra DJP. Catalase, peroxidase, and polyphenoloxidase activities during rice leaf senescence. 1976, 57(2):315–9. [DOI] [PMC free article] [PubMed]
- 24.Heikal TM, El-Sherbiny M, Hassan SA, Arafa A, Ghanem HZ. Antioxidant effect of selenium on hepatotoxicity induced by chlorpyrifos in male rats. Int J Pharm Pharm Sci. 2012;4(4):603–9. [Google Scholar]
- 25.Halliwell B, Foyer C. Properties and physiological function of a glutathione reductase purified from spinach leaves by affinity chromatography. Planta. 1978;139:9–17. [DOI] [PubMed] [Google Scholar]
- 26.Nakano Y, Asada K. Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts. Plant Cell Physiol. 1981;22(5):867–80. [Google Scholar]
- 27.Šerá B, Scholtz V, Jirešová J, Khun J, Julák J, Šerý M. Effects of non-thermal plasma treatment on seed germination and early growth of leguminous plants—A review. Plants. 2021;10(8):1616. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 28.Guo Q, Meng Y, Qu G, Wang T, Yang F, Liang D, Hu S. Improvement of wheat seed vitality by dielectric barrier discharge plasma treatment. Bioelectromagnetics. 2018;39(2):120–31. [DOI] [PubMed] [Google Scholar]
- 29.Ling L, Jiafeng J, Jiangang L, Minchong S, Xin H, Hanliang S. Yuanhua DJSr: effects of cold plasma treatment on seed germination and seedling growth of soybean. 2014, 4(1):5859. [DOI] [PMC free article] [PubMed]
- 30.Li Y, Wang T, Meng Y, Qu G, Sun Q, Liang D, Hu SJPC. Air atmospheric dielectric barrier discharge plasma induced germination and growth enhancement of wheat seed. Process P. 2017;37:1621–34. [Google Scholar]
- 31.Fereydooni M, Alizadeh HHAJJAM. Microscopic investigation of cold plasma effect on chickpea seed germination. 2022, 12(2):231–40.
- 32.Švubová R, Slováková Ľ, Holubová Ľ, Rovňanová D, Gálová E, Tomeková JJP. Evaluation of the impact of cold atmospheric pressure plasma on soybean seed germination. 2021, 10(1):177. [DOI] [PMC free article] [PubMed]
- 33.Leti L-I, Gerber IC, Mihaila I, Galan P-M, Strajeru S, Petrescu D-E, Cimpeanu M-M, Topala I, Gorgan D-L. The modulatory effects of non-thermal plasma on seed’s morphology, germination and genetics—A review. Plants. 2022;11(16):2181. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 34.Lukes P, Dolezalova E, Sisrova I, Clupek M. Aqueous-phase chemistry and bactericidal effects from an air discharge plasma in contact with water: evidence for the formation of peroxynitrite through a pseudo-second-order post-discharge reaction of H2O2 and HNO2. Plasma Sources Sci Technol. 2014;23(1):015019. [Google Scholar]
- 35.Hensel K, Kučerová K, Tarabová B, Janda M, Machala Z, Sano K, Mihai CT, Ciorpac M, Gorgan LD, Jijie R. Effects of air transient spark discharge and Helium plasma jet on water, bacteria, cells, and biomolecules. Biointerphases. 2015;10(2):029515. [DOI] [PubMed] [Google Scholar]
- 36.Janda M, Hensel K, Tóth P, Hassan ME, Machala Z. The role of HNO2 in the generation of plasma-activated water by air transient spark discharge. Appl Sci. 2021;11(15):7053. [Google Scholar]
- 37.Barraud N, Hassett DJ, Hwang S-H, Rice SA, Kjelleberg S, Webb JS. Involvement of nitric oxide in biofilm dispersal of Pseudomonas aeruginosa. J Bacteriol. 2006;188(21):7344–53. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 38.Pawlat J, Starek A, Sujak A, Kwiatkowski M, Terebun P, Budzeń MJPP, Polymers. Effects of atmospheric pressure plasma generated in GlidArc reactor on Lavatera thuringiaca L. seeds’ germination. 2018, 15(2):1700064. [DOI] [PMC free article] [PubMed]
- 39.Janda M, Martišovitš V, Hensel K, Machala ZJPC, Processing P. Generation of antimicrobial NO x by atmospheric air transient spark discharge. 2016, 36:767–81.
- 40.Li Y, Wang T, Meng Y, Qu G, Sun Q, Liang D, Hu S. Air atmospheric dielectric barrier discharge plasma induced germination and growth enhancement of wheat seed. Plasma Chem Plasma Process. 2017;37:1621–34. [Google Scholar]
- 41.Hosseini SI, Mohsenimehr S, Hadian J, Ghorbanpour M, Shokri B. Physico-chemical induced modification of seed germination and early development in artichoke (Cynara scolymus L.) using low energy plasma technology. Phys Plasmas 2018, 25(1).
- 42.Gómez-Ramírez A, López-Santos C, Cantos M, García JL, Molina R, Cotrino J, Espinós J, González-Elipe AR. Surface chemistry and germination improvement of Quinoa seeds subjected to plasma activation. Sci Rep. 2017;7(1):5924. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 43.Muñoz-Llandes CB, Martínez-Villaluenga C, Palma-Rodríguez HM, Román-Gutiérrez AD, Castro-Rosas J, Guzmán-Ortiz FA. Effect of germination on starch. Starch: advances in modifications, technologies and applications. Springer; 2023. pp. 457–86.
- 44.Šerá B, Scholtz V, Jirešová J, Khun J, Julák J, Šerý MJP. Effects of non-thermal plasma treatment on seed germination and early growth of leguminous plants—A review. 2021, 10(8):1616. [DOI] [PMC free article] [PubMed]
- 45.Meng Y, Qu G, Wang T, Sun Q, Liang D, Hu S. Enhancement of germination and seedling growth of wheat seed using dielectric barrier discharge plasma with various gas sources. Plasma Chem Plasma Process. 2017;37:1105–19. [Google Scholar]
- 46.Goudarzi S, Ghafoorifard H, Ghasemi S, Mazandarani A. The effect of atmospheric cold plasma on the rates of germination and root length and shoot length of sesame seed. In: 2021. P1-7. In 27th Iranian nuclear conference.
- 47.Fereydoni M, Alizaheh H. Microscopic investigation of cold plasma effect on chickpea seed germination. 2020.
- 48.Guo Q, Meng Y, Qu G, Wang T, Yang F, Liang D, Hu SJB. Improvement of wheat seed vitality by dielectric barrier discharge plasma treatment. 2018, 39(2):120–31. [DOI] [PubMed]
- 49.Meng Y, Qu G, Wang T, Sun Q, Liang D, Hu SJPC, Processing P. Enhancement of germination and seedling growth of wheat seed using dielectric barrier discharge plasma with various gas sources. 2017, 37:1105–19.
- 50.Noormohammadi Z, Mohammadzadeh-Shahir M, Fahmi D, Atyabi SM, Farahani FJNBR. Induced genetic and morphological changes in Catharanthus roseus L. by cold atmospheric plasma. 2019, 6(3):302–10.
- 51.Goudarzi S, Ghafoorifard H, Ghasemi S, Mazandarani AJP, with English Abstract. The effect of atmospheric cold plasma on the rates of germination and root length and shoot length of sesame seed. 27th Iran Nuclear conf. Mar. 1–7. 2021.
- 52.Filatova I, Azharonok V, Goncharik S, Lushkevich V, Zhukovsky A, Gadzhieva G. Effect of RF plasma treatment on the germination and phytosanitary state of seeds. J Appl Spectrosc. 2014;81(2):250–6. [Google Scholar]
- 53.Zhang JJ, Kwon T, Kim SB, Jeong DK. Plasma farming: non-thermal dielectric barrier discharge plasma technology for improving the growth of soybean sprouts and chickens. Plasma. 2018;1(2):285–96. [Google Scholar]
- 54.Rasooli Z, Barzin G, Mahabadi TD, Entezari M. Stimulating effects of cold plasma seed priming on germination and seedling growth of cumin plant. South Afr J Bot. 2021;142:106–13. [Google Scholar]
- 55.Qadri R, Azam M, Khan I, Yang Y, Ejaz S, Akram MT, Khan MA. Conventional and modern technologies for the management of post-harvest diseases. Plant Disease Manage Strategies Sustainable Agric through Traditional Mod Approaches 2020:137–72.
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Supplementary Materials
Data Availability Statement
This research was done according to Ling et al. 2014, with the modification of seed number per replicate. All data generated or analysed during this study are included in this published article (supplementary information files: S1, S2, S3, S4 and S5).