Document Type : Original Article

Authors

1 Assistant Prof. of Plant Physiology, Department of Biology, Faculty of Science, Arak University, Arak, Iran.

2 M.Sc. Student of Plant Physiology, Department of Biology, Faculty of Science, Arak University, Arak, Iran.

Abstract

Introduction
Iron is an essential micronutrient for plant growth that plays an important role in plant metabolism. Iron deficiency is an abiotic stress that is often found in plants grown in calcareous and alkaline soils. The solubility of Fe+3 decreases dramatically with increasing pH. 30% of the arable land worldwide consists of calcareous and alkaline soils. Common iron fertilizers used to reduce deficiency syndromes contain iron(II) sulfate heptahydrate (FeSO4.7H2O) or iron chelates. Iron chelate (for example Fe-EDTA) is absorbed by plants, which however depends on soil conditions especially soil pH. Nowadays , nano-Fe fertilizer can be used as a rich source of iron for plants ,because it gradually releases Fe in a wide pH range (pH 3– 11). Nanofertilizer usage leads to increase element efficiency, reduce soil toxicity and negative effects caused by the excessive consumption of chemical fertilizers and reduce the fertilizer’ s application . This research was carried out to determine the suitable type of iron fertilizer and to evaluate the effects of different concentrations of nano-Fe fertilizer on Medicago sativa
Materials and methods
In order to investigate the effects of Fe-deficiency and different levels of Fe2O3 nanoparticles compared to Fe-EDTA on leaf growth, photosynthetic pigments and antioxidative activity of alfalfa (Medicago sativa cv.Hamadani), an experiment was conducted based on completely randomized design with three replications in Arak University during 2015. After germination of sterilized seeds of alfalfa, 1-day seedlings were cultured in plastic vases contains perlite. Plants were maintained under 25/18°C day/night temperatures with 12-hr photoperiod. Irrigation was done weekly with 100ml complete Hoagland solution (containing iron chelate (Fe-EDTA) for control plants) or 100ml Hoagland solution without iron chelate and containing different concentrations of ironoxide nanoparticles (0, 5, 10, 20 and 25µM). Plants treated with 0 µM iron nanoparticles did not receive iron in 45day period. Ironoxide nanoparticles were prepared from Pishgaman Company located in Mashhad, Iran . After the final harvest of 45-day plants, leaf fresh weights were measured. Dry weight of leaf were obtained by drying samples in an oven for 24h at 75°C until constant weight. Chlorophylla, chlorophyllb, total chlorophyll and carotenoids contents by spectrophotometry method at 663 and 645 nm were determined. Contents of proline, activities of catalase(CAT), guaiacol peroxidase(GPOX) and superoxide dismutase(SOD) and 2,2diphenyl-1-picrylhydrazyl (DPPH) scavenging activity were measured. All data were analyzed by variance analysis using SPSS16. Mean comparisons were conducted using Duncan’s test.
Results and discussion
Iron treatment had a positive effect on the growth parameters and photosynthetic pigments. The highest and lowest values of leaf growth and photosynthetic pigments were obtained at 25μM and 0µM of Fe2O3 nanoparticles, respectively. Treatment of iron nanofertilizer at different concentrations (even at 5µM) caused significant increase in leaf growth and photosynthetic pigments content compared to iron chelate. Nanoparticles have high reactivity because of more specific surface area, more density of reactive areas, or in creased reactivity of these areas on the particle surfaces. These features streamline the absorption of fertilizers , which are produced in nano scale. Iron nanofertilizer can be considered as an enriched source of bivalent iron for plant because of its high stability and gradual release of Fe in a wide pH range (pH 3–11). Iron is a necessary element for the formation of chlorophyll, therefore deficiency of iron in plants reduces chlorophyll content, so photosynthesis decreases
The highest values of proline and antioxidants activity were measured in 0μM ironoxide nanoparticles. Thus, iron concentration of 0μM is considered stressful for alfalfa. Under abiotic stresses, reactive oxygen species (ROS) content increases more than the normal condition. ROS are highly reactive and toxic and damages cell membranes. Therefore, the concentration of ROS in cell must be controlled. In such conditions, plants develop a high efficient antioxidant defense system to increase tolerance to different stress factors. Overexpression of ROS scavenging enzymes like SOD, CAT, and GPOX resulted in abiotic stress tolerance in various crop plants due to efficient ROS scavenging capacity. It has been reported that proline act as an osmolyte, a metal chelator, ROS scavenger/an antioxidative, membrane stabilizer, and a signaling molecule Thus, the proline content is a good marker for screening tolerant varieties under stress condition
 In this study , There was no statistical difference between different levels of Fe2O3 nanoparticles and Fe-EDTA based on values of proline and antioxidants activity, because stress conditions are not created in levels of Fe2O3 nanoparticles and Fe-EDTA.
Conclusions
Therefore, the suitable type of iron-fertilizer for alfalfa is iron nano-fertilizer and the concentration of 25μM Fe2O3 nanoparticles is the optimum value.

Keywords

Abe, N., Murata, T., Hirota A., 1998. Novel 1,1-diphenyl-2-picryhy-drazyl- radical scavengers, bisorbicillin and demethyltrichodimerol, from a fungus. Bioscience, Biotechnology and Biochemistry. 62, 661-662.
Apel, K., Hirt, H., 2004. Reactive oxygen species: Metabolism, oxidative stress and signal transduction. Plant Biology. 55, 373-379.
Armin, M., Akbari, S., Mashhadi, S., 2014. Effect of time and concentration of nano-Fe foliar application on yield components of wheat. International Journal of Biosciences. 4(9), 69-75.
Arnon, D.I., 1949. Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. Plant Physiology. 24, 1-15.
Ashraf, M., Foolad, M.R., 2007. Roles of glycine betaine and proline in improving plant abiotic stress resistance. Environmental and Experimental Botany. 59, 206-216.
Bakhtiari, M., Moaveni, P., Sani, B., 2015. The effect of iron nanoparticles spraying time and concentration on wheat. Biological Forum-An International Journal. 7(1), 679-683.
Bates, L.S., Waldren, R.P., Teare, I. D., 1973. Rapid determination of free proline for water stress studies. Plant and Soil. 29, 205-207.
Briat, J. F., Dubos, C., Gaymard, F., 2015.Iron nutrition, biomass production, and plant product quality. Trends in Plant Science. 20(1), 33-40.
Bybordi, A., 2012. Study effect of salinity on some physiologic and morphologic properties of two grape cultivars. Life Science Journal. 9(4), 1092-1101.
Cakmak, I., Marschner, H., 1992. Magnesium deficiency and high light intensity enhance activities of superoxide dismutase, ascorbate peroxidase, and glutathione reductase in bean leaves. Plant Physiology. 98, 1222-1227.
Chinnamuthu, C.R., Boopathi, M.P., 2009. Nanotechnology and agroecosystem. Plant Nutrition and Soil Science. 168, 558-573.
Delic, D., Stajkovic-Srbinovic, O., Radovic, J., Kuzmanovic, D., Rasulic, N., Simic, A., Knezevic-Vukcevic, N., 2013.Difference in symbiotic N2 fixation alfalfa, Medicago sativa L. cultivars and Sinorhizobium spp. strains in field conditions. Romanian Biotechnological Letters. 18(6), 8743-8750.
Dokhe, S. A., Mahajan. P., Kamble, R., Khanna, A., 2013. Effect of nanoparticles suspension on the growth of mung (Vigna radiata) seedlings by foliar spray method. Nanotechnology Development. 3(1), 1-5.
Foyer, C.H., Noctor, G., 2005. Redox homeostats and antioxidant signaling: a metabolic interface between stress perception and physiological responses. Plant Cell. 17, 1866-1875.
Hoagland, D.R., Arnon, D.I., 1950. The water-culture method for growing plants without soil. California Agricultural Experiment Station Circular. 347, 1-32.
Gohari, A., Niyaki, A.N., 2010. Effect of iron and nitrogen fertilizers on yield and yield components of peanut (Arachis hypogaea L.) in Astaneh Ashrafiyeh, Iran. American-Eurasian Journal Agriculture and Environmental Science. 9(3), 256-262.
Giannopolitis, C.N., Ries, S.K., 1997. Superoxide dismutases: I. occurrence in higher plants. Plant Physiology. 59, 309-314.
Kosova'a, K., Vita'mv'sa, P., Pra'sila, I.T., Renautb, J., 2011. Plant proteome changes under abiotic stress-contribution of proteomics studies to understanding plant stress response. Proteomics. 74(8), 1301-1322.
Lichtenthaler, H.K., Wellburn, A.R., 1983. Determinations of total carotenoids and chlorophylls a and b of leaf extract in different solvents. Biochemical Society Transactions. 11, 591-592.
Mazaherinia, M.A., Astaraei, R., Fotovat, A., Monshi, A., 2010. Effect of Nano iron oxide particles and on Fe, Mn, Zn, Cu concentrations in wheat plants.W orld Applied Science. 7(1), 156-162.
Mazlomi-Mamyandi, M. Pirzad, A. Zardoshti, M.R., 2012. Effect of Nano-iron spraying at varying growth stage of sugar beet (Beta vulgaris L.) on the size of different plant parts. International Journal of Agriculture and Crop Sciences.4 (12), 740-745.
Mirza Masoumzadeh, B., Imani, A.A., Khayamaim, S., 2012. Salinity stress effect on proline and chlorophyll rate in four beet cultivars. Scholars Research Library. 3(12), 5453-5456.
Mohamadipoor, R., Sedaghathoor, S., Mahboub-Khomami, A., 2013.Effect of application of iron fertilizer in two methods foliar and soil application on growth characteristics of Spathyphyllum illusion. European Journal of Experimental Biology. 3(1), 232-240.
Naderi, M.R., Abedi, A., 2012. Application of nanotechnology in agriculture and refinement of environmental pollutants. Journal of Nanotecnology. 11(1), 18-26.
Peyvandi, M., Parande, H., Mirza, M., 2011. Comparison of nano Fe chelate with Fe chelate effect on growth parameters and antioxidant enzymes activity of Ocimumbasilicum. New Cellular and Molecular Biotechnology Journal. 4, 89-99. [In Persian with English Summary].
Pii, Y., Penn, A., Terzano, R., Crecchio, C., Mimmo, T., Cesco, S., 2015. Plant-microorganism-soil interactions influence the Fe availability in the rhizosphere of cucumber plants. Plant Physiology and Biochemistry. 87, 45-52.
Polle, A., Otter, T., Seifert, F., 1994. Apoplastic peroxidases and lignification in needles of Norway spruce (Picea abies L.). Plant Physiology. 106, 53-56.
Prasad, T.N.V., Sreeprasad, T.S., Sajanlal, P.R., Pradeep, T., 2012. Effect of nanoscales zinc oxide on the germination, growth and yield of peanut. Journal of Plant Nutrition. 35, 905-927.
Ranjbar, M. Shams, G.A., 2009. Study of nanotechnology applications. Sabz Zist. 3, 28-34. [In Persian].
Roosta, H. R., Jalali, M., Vakili Shahrbabaki, M. A., 2015. Effect of nano-chelate, Fe-Eddha and FeSO4 on vegetative growth, physiological parameters and some nutrient elements concentration of four varieties of lettuce (Lactuca sativa L) in NFT system. Journal of Plant Nutrition. 38(14), 2176-2184.
Rose, H., Benzon, L., Rosnah, M., Rubenecia, U., Ultra, V., Lee, S.C., 2015. Nano-fertilizer affects the growth, development and chemical properties of rice. International Journal of Agronomy and Agricultural Research. 7(1), 105-117.
Rout, G.R., Das, A.B., Sahoo, S., 2014. Screening of iron toxicity in rice genotypes on the basis of morphological, physiological and biochemical analysis. Journal of Experimental Biology Agricultural Science. 2, 567-582.
Sajedi, N., Ardakani, M.R., 2008. Effect of different levels of nitrogen, iron and zinc on physiological indices and forage yield of maize (Zea mays L.) in Markazi province. Iranian Journal of Field Crops Research. 6(1), 99-110. [In Persian with English Summary].
Tan, J., Zhao, H., Hoang, J., Han, Y., Li, H., Zhao, W., 2008. Effects of exogenous nitric oxide on photosynthesis, antioxidant capacity and proline accumulation in wheat seedlings subjected to osmotic stress. Agricultural Sciences. 4, 307-313.
Wang, Y.X., Oyaizu, H., 2009. Evaluation of the phytoremediation potential of four plant species for dibenzofuran-contaminated soil. Journal of Hazardous Materials, 168, 760-764.
Vasconsuelo, A., Boland, R., 2007. Molecular aspects of the early stages of elicitation of secondary metabolites in plants. Plant Science. 172, 861-875.
Vendruscolo, E.C.G., Schuster, I., Pilegg, M., Scapim, C.A., Molinari, H.B.C., Marur, C.J., Vieira, L.G.E., 2007.Stress-induced synthesis of proline confers tolerance to water deficit in transgenic wheat. Journal of Plant Physiology. 164(10), 1367-1376.
Vigani, G., 2012. Does a similar metabolic reprogramming occur in Fe-deficient plant cell and animal tumor cells? Frontiers in Plant Science. 3, 47-51.
Yadav, S.K., 2010. Heavy metals toxicity in plants: An overview on the role of glutathione and phytochelatins in heavy metal stress tolerance of plants. South Africa Journal Botany. 76, 169-179.