Impact of Selenium Nanoparticles on Growth, Biochemical Characteristics and Yield of Cluster Bean Cyamopsis tetragonoloba

The present study deals with the impact of selenium nanoparticles on growth, biochemical characteristics and yield of Cluster bean Cyamopsis tetragonoloba grown for a period 60 days Sodium selenite and ascorbic acid was utilized for the synthesis of Selenium nanoparticles using precipitation method. Selenium nanoparticles were characterized by using SEM, EDAX, FTIR and XRD. Pot culture studies of cluster bean in different quantity of Selenium nanoparticles such as 0,100, 200, 300, 400 and 500mg for treatment T0 (Control) T1, T2, T3, T4 and T5 and growth biochemical and yield were estimated at the end of 60 days. SEM image of selenium nanoparticles was observed as spherical in shape. EDAX spectrum recorded on purity of selenium nanoparticles. The FTIR spectrum of selenium nanoparticles was analyzed in the range of 4000-400 cm-1 spectral bands were observed. The germination percentage in T0, T1, T2, T3, T4 and T5 are 100,90,80,90,100 and 100 respectively. Among the treatments the shoot length is higher (21.8) in T1 containing 100mg of selenium nanoparticles and lower in(12.01) T5 containing 500mg of nanoparticles. Root length, fresh and dry weight and leaf area were higher in T2. The vigor index is higher T4.The chlorophyll a, b total Chlorophyll, carotenoids, anthocyanin, protein, L-proline, free amino acids and leaf nitrate were higher inT4.Among the treatments yield of cluster bean is higher in T4 and lower in T0.

INTRODUCTION Nanotechnology is highly promising and rapidly progressing discipline in research and influencing every field of science and biology. Nanotechnology is creating many new materials and devices with a vast range of application such as medicine, biomaterials and energy production. Exploring comprehensive application profile nanoparticles may revolutionize research in crop science and transform agriculture in to industry (1). Application of nanotechnology in agriculture delivery to plant technology also holds the promise of controlled release of agro chemicals and its targeted delivery of various macromolecules needs for improved plant disease resistance, efficient nutrient utilization and enhanced plant growth. Recent research on nanoparticles in a number of crop like corn, wheat, soybean, tomato and cucumber have provided evidence of enhanced seedling growth, germination, nitrogen metabolism, photosynthetic activity and protein level indicating their potential use for crop improvement. Among nanoparticles, Selenium is proved to be an essential mineral required for proper health, immunity, and reproductive functions of animals. Plants are the main source of this element, it is important to increase its plant growth. A new approach to fertilization of plants is the use of selenium nanomaterials (2).The study related to the impact of selenium nanoparticles on growth, biochemical characteristics and yield of vegetable crop cluster bean is totally wanting. Hence the present study was carried out.

Synthesis of Selenium Nanoparticles
Precipitation method is adopted for the synthesis of selenium nanoparticle. For the synthesis 0.7Mg of (700mg) sodium selenite were dissolved in 50ml of distilled water under stirring vigorously using magnetic stirrer for 20 minutes. After stirring, the precipitation was achieved by adding 50ml of ascorbic acid solution in drop wise under constant stirring. The initial pH was observed as 3 and it was increased to pH 14 .Then precipitating process was continued until the orange colour precipitate was obtained. Then this precipitate was centrifuged at 1500 rpm for 20 tetragonaloba was selected for pot culture studies based on their easy availability, relative importance in daily diet of a common man, surviving capacity, growth capabilities and economic growth.

Pot Culture Studies:
For the pot culture studies, the seeds were soaked in ground water and kept as control. Both the control and experimental seeds were allowed to grow in plastic pots (25 cm diameter, 25 cm height) containing a mixture of red soil, cow dung manure in the ratio of 1:1 The experimental pots were supplied with different quantities of selenium nanoparticles such as 0,100,200,300,400 and 500 for treatment 1 (Control) 2, 3,4,5,6 respectively. Triplicates were maintained and grown in net house for a period of 60 days. Pots were irrigated with well water. After 60 days growth and biochemical characteristics were estimated.

III.
RESULTS AND DISCUSSION As C6H8O6 was added to Na2SeO3, it is found to change colour from orange to red colour is shown in Fig.1 and this colour change indicates the synthesis of selenium nanoparticles (Se).Precipitation was observed by increasing the P H from 2.3 to 5.8 . Fig.1

: Synthesis of Selenium Nanoparticles
The SEM image (Fig.2) showing the high density chemical by synthesized Se further confirmed the development of selenium nanostructures. Obtained nanoparticles showed spherical in nature. The microscopic image showed that the Se nanoparticles did not appear as discrete particles but form much larger dendritic flocks whose size could reached micron scale size range about 9.09mm ( scale bar 5µm),9.07mm 27mm ( scale bar 10µm),9.05mm ( scale bar 20µm) for figure 1 a,b and c respectively. Scanning electron microscope (SEM) images were taken for the analysis of size and shape of SeNPs (Hitachi s-3400N) with resolution of 500 nm operated at 10 kV HV mode and detectors contain secondary electron; semiconductor BSE (Quad type)(3). The SEM images of selenium nanoparticles synthesized by different combinations were oval in shape with smooth surface. The particle size was found to be around 50-150 nm. Sonam Malhotra et al.,(2014)(4) suggested that properties forming a spherical shape nanoparticle having a size range of 20 to 30 nm as measured using particle size analyser, purity of the Nano selenium were further measured by the (SEM) Scanning Electron Microscope. Selenium nanoparticles were highly using   (Fig. 4).All diffraction peaks indexed according to the hexagonal phase of selenium characteristic peaks of impurity phase except selenium are found which revealed that good crystalline in nature of the sample. The broading of the peaks in the above XRD pattern can be attributed to the small particles size of the synthesized selenium. This proves that pure selenium nanoparticles were synthesized.Similar X-ray diffraction (XRD) patterns of selenium nanoparticles was also reported(3).

Fig.4: Analysis of Selenium Nanoparticles (XRD) Image
Fourier Transform Infrared Spectroscopy measurements were carried out to identify the possible functional group responsible for the reduction of the selenite in chemical synthesized selenium nanoparticle. The FTIR spectrum of the selenium was analyzed in the range 4000 -400cm (  Table 1). The peaks obtained were plotted as % transmit-trance in X axis and wave number (cm -1) in Y axis. Salwa and Abbas.,(2012)(6)suggested that FTIR study was carried out to confirm the coating. In dextrin coated nanoparticles, shift in peak 1417 per cm in FTIR spectrum was observed indicating H-C-OH bond. As the concentration of Dextrin increases the shift in the peak from 1417 cm-1 to 1384 cm-1 was observed.) The FTIR analysis was performed to characterize the surface chemistry of selenium nanoparticles produced by BSA and analysis of FTIR indicated protein mediated synthesis of selenium nanoparticles, the strong absorption bands at 1649 and at 1551/ cm are characteristic of amide I and C-H vibrations of CH2 groups of protein moiety respectively, with albumin as the stabilizing and capping agent surrounding the selenium nanoparticles.    (7). The shoot length of cluster bean in control is 21.7cm. Among the treatments the shoot length is higher (21.8) in T1 containing 100mg of selenium nanoparticles and lower in T5 (12.01) containing 500mg of nanoparticles. Similar result was also reported in Cluster bean treated with 100mg of Zinc (8). The root length of the cluster bean in control is 10.4cm. Among the treatments the root length is higher in T2(10.6cm) and lower in T3(7.3cm).Marisamy et al.,(2015) (9)suggested that shoot length is higher in control and lower in treatment 5(10mM).Similar root length was reported when pea nut is treated with 100mg of Zno nanoparticles (10). The fresh weight of t he cluster bean in control is 3.18g. Sanghpriya Gautam et al .,(2015) (11)reported that fresh and dry weight of S. oleracea increased at treatment T4 There was maximum increase in fresh weight (38.6 %) and dry weight (78.3 %) at T4. A reduction of fresh and dry weight was reported in Chloroxylon swietenia treated with sugar mill effluent for a period of 90 days (12). The leaf area is higher inT3, T4  -2, Issue-6, Nov-Dec-2017  http://dx.doi.org/10.22161/ijeab/2.6.19  ISSN: 2456-1878 www.ijeab.com Page | 2922 increasing quantity of zinc nanoparticles treated in Lady's finger. The chlorophyll a,b and total chlorophyll of cluster bean is presented in figure 6.The chlorophyll a is higher in T4 and lower in T1. The chlorophyll b is higher in T4 and lower in T1. Total Chlorophyll is higher in T4 and lower in T1. Similar study was also reported when Lady's finger was treated with different quantities of zinc oxide nanoparticles (13). Se treatment at the lower concentration (16μM) recorded the highest values of chlorophyll a concentration (2.68 and 1.99 mg/ g FW). Vijayarengan(2013) (8)reported that chlorophyll 'a', chlorophyll 'b', total chlorophyll and carotenoid content of cluster bean leaves increased at lower concentration. An increase in chlorophyll a and chlorophyll b contents of wheat seedlings may be attributed to selenium effect on protection of chloroplast enzymes and thus increasing the biosynthesis of photosynthetic pigments (6). High concentration of selenium induced reduction in photosynthetic pigments content. Marisamy et al.,(2015) (9)suggested that the chlotophyll a, and b are higher in control and lower in treatment 5(10mM) . Sanghpriya Gautam et al.,(2015) (11) reported that pigment contents (chlorophyll a, b) were increased maximum up to T3, whereas decreased at T4.

Fig.6: Chlorophyll a,b and Total Chlorophyll of Cluster bean
The carotenoids and anthocyanin of Cluster bean is presented in Figure 7. The carotenoids are higher in T4 and lower in T2..Salwa and Abbas.,(2012) (6) suggested that the selenium increased the contents of carotenoids and chlorophyll a,and in turn change in the photosynthetic pigments level is likely to have been connected with different effects of the selenium ions on the oxidationreduction status of leaves. The carotenoids content of Amaranthus caudatus is higher in control and lower in treatment 5(10mM) when treated with barium (9). The   (9)reported that anthocyanin is higher in control and lower in treatment 5(10mM) when Amaranthus caudatus was treated with nanoparticles.

Fig.7: Anthocyanin and Carotenoids of Cluster bean
The protein and L-proline of the cluster bean is presented in Figure 8.The protein content is higher in T4(5.5mg/g) lower in T5(3.4mg/g). Vijayarengan (2013) (8) reported that the protein content of leaves was maximum at 50 mg of zinc treated with cluster bean. Suresh kumar and Total soluble protein content was found to be 35 % in 90 days at 50% treatment when compared to control in sugarcane effluent (12). The L-proline is higher in T4(5.2mg/g) and lower in T1 (4.1mg/g). Proline concentration was significantly increased in Lady's finger treated with Zinc oxide nanoparticles when compared to untreated ones (14). Also reported that L-proline accumulates in the leaves of many tree species when subjected to stress (12).    Figure 9.The free amino acids are higher in T4 (6.8mg/g) lower in T2(5.3mg/g).). The free amino acids content in Helianthus annuus significantly increased with increasing concentration of barium (9). Suresh Kumar and Mariappan.,(2013)(12) reported a reduction in soluble protein level eventually leads to increase in free amino acids content. The leaf nitrate is higher in T4(5.9mg/g) and lower in T1(4.12mg/g). Marisamy et al.,(2015) (9)suggested that leaf nitrate content increased in control when compared to barium treatment. Suresh Kumar and Mariappan (2013) (12) reported that leaf nitrate accumulated in all the effluent treated tree species.

Fig.9: Free Amino acids and Nitrate of Cluster bean
Yield performance of cluster bean is presented in figure Table 3.Yield performance of cluster bean is higher in T4(1.41g) and lower in T2(0.69g). Sharma and Kansal(1984) (14) suggested that the yield parameters such as length, weight and number of the cluster bean showed increase over control up to 800mg and decrease gradually in the further higher concentrations. Manal et al.,(2014) (15) reported that imposition of drought stress reduced plant height, number of tillers, 1000-grain weight and grain yield of both rice cultivars. The treatment with either Se has a favorable effect on1000-grain weight which up to 9.5% and 5.2%.  T0  T1  T2  T3  T4  T5 Free amino acids(mg/g) Nitrate (mg/g)