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Effect on germination parameters and early growth in barley (Hordeum vulgare L.) seeds exposed to chitosan and silicon dioxide (SiO
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nanoparticles
Faride Behboudia, Zeinolabedin Tahmasebi Sarvestanib*, Mohamad Zaman Kassaeec, Seyed Ali Mohamad Modares Sanavid, Ali Sorooshzadehe
a:PhD Student, Department of Agronomy, Faculty of Agriculture, Tarbiat Modares University, Iran;
*b:Associate Professor, Department of Agronomy, Faculty of Agriculture, Tarbiat Modares University, Iran;
c:Professor, Department of Chemistry, Collage of Sciences, Tarbiat Modares University, Iran;
d:Professor, Department of Agronomy, Faculty of Agriculture, Tarbiat Modares University, Iran;
e:Associate Professor, Department of Agronomy, Faculty of Agriculture, Tarbiat Modares University, Iran
*Corresponding author:[email protected]
ABSTRACT
Plants need to be included to develop a comprehensive toxicity profile for nanoparticles (NPs). In this experiment, the effects of chitosan silicon dioxide (SiO2)NPs with concentrations 0 (control), 30, 60 and 90 ppm was assessed on barley seeds. For starting the experiment, the barley seeds were separately placed in petri dishes. Then, 5 mL of the both NPs concentration were added to them. Results showed that the treated seeds with different concentration of NPs showed significant effects on barley germination and growth parameters. Applications of both NPs in 90 ppm, especially SiO2 NPs, displayed adverse effects on the studied traits; whereas, using both NPs with 30 or 60 ppm had not significant effects on majority of the studied traits in barley plants. In general, the usage of these NPs in high concentration (90 ppm) could curb the undesired growth and germination of barley plants.
Key words: Vigor Index, Chitosan, Seedling length, Toxicity, SiO2
Introduction
Increasing production and use of nanoparticles (NPs) have raised concerns about their possible impacts on environmental and human health (Hood, 2004). As NPs are being used on large scale, they might have entered in ecosystems. Such NPs on entering in ecosystem might have affected seed germination process, plant growth and metabolism. Hence, plants should be tested to establish their response to NPs stress. NPs may be more toxic than the corresponding bulk materials, and they have the potential ability of passing the cell membrane of living organisms because of their general size between 1 to 100 nm (Zhu et al., 2006). Use of SiO2 nanoparticles (NPs) and associative polymers has increased during the last decade (Lin et al., 2006). These NPs are widely used in construction, catalysis, paints and pharmaceutical applications (Wang et al., 2005). Study on the influence of metal NPs (Si, Pd, Au, and Cu) on germination of lettuce seeds indicated that NPs (Pd, Au at low concentrations; Si, Cu at higher concentrations, and combination of Au and Cu) had a positive effect on seed germination, shoot to root ratio and growth of the seedling (Shah and Belozerova, 2009). Lu et al (2002) reported that application of SiO2 and TiO2 nanoparticles mixture on soybean (Glycine max L.) could increase nitrate reductase, enhance water and fertilizer absorbing capacity, stimulate antioxidant defense system and also hasten seed germination and seedling growth.
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On the other hand, Chitosan, a biodegradable and biocompatible polymer, is a modified natural carbohydrate and the second most abundant polysaccharide in nature. It can be synthesized by the partial N-deacetylation of chitin, a natural biopolymer derived from crustacean shells such as crabs, shrimps and lobsters (IIIum, 1998). Chitosan has promoted growth of various crops such as wheat (Triticum spp.), rice (Oriza spp.), maize (Zea mays L.), peanuts (Arachis hypogaea L.) and carrots (Daucus carota L.) (El Hadrami et al., 2010).
). It promotes plant growth through increasing the availability and uptake of water and essential nutrients through adjusting cell osmotic pressure (Guan et al., 2009).
The aim of present study was to evaluate the effect of chitosan and SiO2 NPs on germination and growth parameters of barley plants.
2. Materials and methods
2.1 Growth condition
Experiments were carried out to assess the effects of chitosan and SiO2 NPs on barley germination in a completely randomized design with seven treatments. The experimental treatments included both NPs with concentrations 0 (control), 30, 60 and 90 ppm. The experiment was performed in a germinator with an average temperature of 21 ±1ºC at the College of Agriculture, Tarbiat Modares University, Tehran, Iran (June, 2015).
2.3. NPs
Size of both NPs were > 80 nm that were determined through Field Emission- Scanning Electron Microscope (FE-SEM).
Fig 1. Field Emission-Scanning Electron Microscope (FE-SEM) images of SiO2 NPs (left) and chitosan NPs (right).
2.4. SiO2 and chitosan NPs solution
The SiO2 and chitosan solution were prepared by (Adhikari et al., 2013) and (Li et al., 2008), respectively.
2.6. Seed treatment
The barley seeds were immersed in a 5% sodium hypochlorite solution for 10 min to ensure surface sterility. Then seeds after being rinsed three times with distilled water were
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shade-dried for 1 hour. The 25 seeds were placed in a Petri dish (100 mm x15 mm) with one piece of sterilized filter paper (Whatman No. 1) and 5 mL of distilled water, SiO2 and chitosan NPs solution were added (ISTA, 2009). Then they were covered and placed in a germinator at 21 ± 1◦C for seven days.
2.7. Measurements
Germination tests were performed according to the rule issued by the International Seed Testing Association (ISTA, 2009). Number of germinated seeds was noted daily for 7 days. Seeds were considered germinated when the radicle showed at least 2 mm in length. On 14th day (after germination), morphological parameters like root, shoot and seedling length were measured. Subsequently, they were oven-dried at 70 °C overnight to estimate their dry weight with a sensitive scale. Data were statistically analyzed using two way analysis of variance (SAS). The significance of differences among treatment means were compared by Duncan’s multiple range tests at P<0.05.
3. Result and discussion
3.1. Germination parameters
Analysis of variance showed that the treatments significantly affected Germination Percentage (GP), Germination Rate (GR), Germination Value (GV), Mean Germination Time (MGT), Pick Value (PV), Mean Daily Germination (MDG), vigor index I and vigor index II (Table 1).
Table 1. Analysis of variance for some related germination parameters of barley
Mean square
Source df GP GR GV PV MDG MGT Vigor index I Vigor index II
Treatment 6 162.09** 10.07 ** 84.64 ** 0.141 ** 14.68 * 0.09 ** 609668.72 ** 1023379.76 **
Error 14 33.90 0.89 1.44 0.007 3.86 0.01 9907.65 66993.97
CV (%) - 6.60 8.62 14.10 15.80 9.02 5.21 9.00 18.58
ns Non- significant, ** and * significant at the 1% , 5% probability levels, respectively
The treated barley seeds with high concentration of both NPs (90 ppm) significantly decreased GR, GV, PV and vigor index II as compared to the control (0 ppm) (Table 2).
Using both NPs with 30 up 90 ppm were significantly decreased seed vigor index II as compared to the control. Application of chitosan NPs had not significant effect on GP and MDG. On the other hand, as a general rule, lower MGT represents a faster germination speed. Exposure of seeds to 60 and 90 ppm chitosan and SiO2 NPs were demonstrated the highest MGT as compared to the control, respectively. In most cases, the influence of SiO2
NPs was more than the other NPs. Effect of SiO2 and chitosan NPs on barley seeds germination was shown in figure 2.
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and chitosan NPs on barley germination parameters Effects of SiO2
Table 2.
Vigor index II Vigor
index I MGT
MDG PV
GV GR
GP Treatments
1913.0 a 1763.17 a
2.08 cd 23.50 a
0.63 ab 14.73 a
12.63 a 94.00 a
Control (No-NPs)
1654.3 a 1252.71 b
2.17 bcd 22.66 a
0.50 b 11.23 b
11.66 ab 90.66 a
30 SiO2 NPs
1519.7 a 1319.56 b
2.12 cd 22.33 a
0.33 c 7.38 c
11.39 ab 89.33 a
60 SiO2 NPs
905.2 b 646.44 d
2.51 a 17.00 b
0.25 c 3.78 de
7.33 c 72.00 b
90 SiO2 NPs
1890.7 a 1335.51 b
1.95 d 23.00 a
0.66 a 15.30 a
12.55 a 92.00 a
30 Chitosan NPs
1560.5 a 977.69 c
2.34 ab 21.33 a
0.25 c 5.41 cd
10.00 b 88.66 a
60 Chitosan NPs
307.5 c 442.67 e
2.19 bc 22.66 a
0.08 d 1.88 e
11.05 ab 90.66 a
90 Chitosan NPs
Means by the uncommon letter in each row and column are significantly different according to Duncan tests (p<0.05).
Fig 2. Images of SiO2 and chitosan NPs effects on barley seeds germination.
The key to increased seed germination is the penetration of NPs into the seed. So, in high concentration of NPs, possibly NPs did not fully penetrate the seed coat and endosperm and thus had limited effects on the embryos. Therefore, NPs, might find the seed coat and endosperm to be effective barriers, especially when they are agglomerated (Duke and Kakefuda, 1981). Haghighi et al (2012) found that 2mM of Nano-Silicon decreased GP and seed vigor index; whereas, GR and MGT increased compared to the control. Suvannasara and Boonlertnirun (2013) stated that the chitosan types in high concentration decreased seed germination of rice.
3.2. Length and Weight
Analysis of variance showed that the effect of treatments were significant for length and dry weight (Table 3).
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Table 3. Analysis of variance for some related growth parameters of barley
Mean square
Source df Length Dry weight
Seedling Coleoptile Shoot Root Seedling Shoot Coleoptile Root Treatment 6 55.83 ** 0.65 ** 8.25 ** 24.95 ** 105.98 ** 36.09 ** 5.04 ** 23.38 **
Error 14 1.46 0.08 0.14 0.97 2.36 1.22 0.43 0.90
%(
CV ( - 9.53 10.09 7.60 12.79 9.69 10.88 15.28 15.83
ns Non- significant, ** and * significant at the 1% , 5% probability levels, respectively
Exposure seeds to both NPs were significantly decreased seedling, coleoptile, shoot and root length as compared to the control (Table 4). Also, the usage of both NPs with high concentration (90 ppm) were significantly decreased dry weight of seedling, coleoptile, shoot and root as compared to the control. In contrast, seeds exposure to 30 ppm chitosan NPs were improved dry weight of seedling, shoot and root as compared to the other treatments.
Table 4. Effects of SiO2 and chitosan NPs on barley growth parameters
Dry Weight (mg) Length (cm)
Root Shoot
Coleoptile Seedling
Root Shoot
Coleoptile Seedling
8.44 ab 11.56 ab
5.16 ab 20.37 a
11.61 a 7.12 a
2.75 b 18.73 a
Control (No-NPs)
6.00 c 12.47 ab
4.59 abc 18.61 ab
8.82 bc 4.91 c
3.05 ab 13.74 b
30 SiO2 NPs
6.87 bc 10.79 b
4.25 bc 16.33 bc
9.40 b 5.36 bc
3.20 ab 14.77 b
60 SiO2 NPs
5.50 c 8.62 c
3.53 c 14.12 c
7.06 cd 3.59 d
2.76 b 10.65 c
90 SiO2 NPs
8.86 a 11.75 ab
5.12 ab 20.50 a
8.69 bc 5.85 b
3.57 a 14.54 b
30 Chitosan NPs
6.00 c 13.04 a
5.75 a 17.71 ab
5.54 d 5.97 b
2.96 b 11.51 c
60 Chitosan NPs
0.37 d 3.00 d
1.87 d 3.37 d
2.76 e 2.13 e
2.06 c 4.90 d
90 Chitosan NPs
Means by the uncommon letter in each row and column are significantly different according to Duncan tests (p<0.05).
Different morphological effects, depending on the nanomaterial type, particle size and concentration (Rico et al., 2011). In this study, the positive influence of chitosan NPs on barley growth was more than SiO2 NPs. Also, the toxic effects of NPs in high concentrations were more pronounced in the roots, probably due to the seed coatings, which can act as a protector for the embryo, but cannot totally guard the whole seed. So, radicles, after penetrating the seed coatings, could contact the NPs directly (Sresty and Rao, 1999). The presence of NPs on the root surface could alter the surface chemistry of the root and clog the
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root openings and both hydraulic and nutrient uptake in roots is inhibited. Therefore, plant growth is negatively affected because NPs. These results seemed consistent with reports by (Siddiqui and Al-Whaibi MH, 2014) and (Mahdavi and Rahimi, 2013).
Conclusion
1. In this paper, negative effects of NPs was observed in barley seeds exposure to NPs.
Using NPs especially SiO2 NPs at high dosage could inhibit seed germination and growth parameters of barley plants.
2. The effect of NPs on plants is complex and many factors can affect plant growth and germination.
3. In general, the usage of these NPs in the applied amounts could curb the undesired growth and germination of this species.
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