Effects of cobalt oxide nanoparticles (Co3O4 NPs) on ion leakage, total phenol, antioxidant enzymes activities and cobalt accumulation in Brassica napus L.

  • Malihe JAHANI Islamic Azad University, Department of Biology, Science and Research Branch, Tehran
  • Ramazan Ali KHAVARI-NEJAD Islamic Azad University, Department of Biology, Science and Research Branch, Tehran
  • Homa MAHMOODZADEH Islamic Azad University, Department of Biology, Mashhad Branch, Mashhad
  • Sara SAADATMAND Islamic Azad University, Department of Biology, Science and Research Branch, Tehran
Keywords: antioxidant defense system; environmental concerns; glutathione S-transferase; nanotoxicity; oxidative stress; phenolic compounds; phenylalanine and tyrosine ammonia lyase


Interaction of nanoparticles (NPs) as a significant threat to ecosystems with biological processes of plants is very important. Here, the effects of cobalt oxide (Co3O4) NPs on some physio-biochemical characteristics of Brassica napus L. were investigated. The two-weeks seedlings were sprayed with different concentrations of Co3O4 NPs (0, 50, 100, 250, 500, 1000, 2000, and 4000 mg L-1). The results showed that this treatment significantly affected the fresh and dry weights, area, relative water content (RWC) and relative chlorophyll value (SPAD) of leaves. The highest reduction of growth and biomass indexes occurred at 4000 mg L-1 NPs. The content of H2O2 and electrolyte leakage (EL) increased respectively, after 100 and 250 mg L-1 of Co3O4 NPs and showed a maximum level at 4000 mg L-1. The activities of phenylalanine ammonia lyase (PAL), ascorbate peroxidase (APX) and superoxide dismutase (SOD) increased after 100 mg L-1 of Co3O4 NPs. However, tyrosine ammonia lyase (TAL) activity enhanced after 500 mg L-1. The catalase (CAT) activity and protein content decreased after 1000 mg L-1 of Co3O4 NPs. Application of concentrations higher than 500 mg L-1 of Co3O4 NPs induced polyphenol oxidase (PPO) activity but reduced glutathione reductase (GR). The activities of guaiacol peroxidase (GPX) and glutathione S-transferase (GST) increased at 250-1000 mg L-1 of Co3O4 NPs and then decreased. These results suggested that low concentrations of Co3O4 NPs induced a positive effect on growth parameters but high levels caused extensive oxidative damage and mediated defense responses by organization of phenolic compounds and antioxidative system.


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Aebi H (1984). Catalase in vitro. Methods in Enzymology 105:121-126.

Agati G, Azzarello E, Pollastri S, Tattini M (2012). Flavonoids as antioxidants in plants: location and functional significance. Plant Science 196:67-76.

Alexieva V, Sergiev I, Mapelli S, Karanov E (2001). The effect of drought and ultraviolet radiation on growth and stress markers in pea and wheat. Plant, Cell and Environment 24(12):1337-1344.

Ayeni OO, Ndakidemi PA, Snyman RG, Odendaal JP (2010). Chemical, biological and physiological indicators of metal pollution in wetlands. Scientific Research and Essays 5(15):1938-1949.

Bakkaus E, Gouget B, Gallien JP, Khodja H, Carrot F, Morel JL, Collins R (2005). Concentration and distribution of cobalt in higher plants: the use of micro-PIXE spectroscopy. Nuclear Instruments and Methods in Physics Research Section B Beam Interactions with Materials and Atoms 231(1-4):350-356.

Beaudoin-Eagan LD, Thorpe TA (1985). Tyrosine and phenylalanine ammonia lyase activities during shoot initiation in tobacco callus cultures. Plant Physiology 78(3):438-441.

Borowiak K, Budka A, Hanć A, Kayzer D, Lisiak M, Zbierska J, ... Łopatka N (2018). Relations between photosynthetic pigments, macro-element contents and selected trace elements accumulated in Lolium multiflorum L. exposed to ambient air conditions. Acta Biologica Cracoviensia Series Botanica 60(1):35-44.

Bradford MM (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 72(1-2):248-254.

Chichiriccò G, Poma A (2015). Penetration and toxicity of nanomaterials in higher plants. Nanomaterials 5(2):851-873.

Dalton DA, Boniface C, Turner Z, Lindahl A, Kim HJ, Jelinek L, ... Taylor CG (2009). Physiological roles of glutathione S-transferases in soybean root nodules. Plant Physiology 150(1):521-530.

Dionisio-Sese M, Tobita S (1998). Antioxidant responses of rice seedlings to salinity stress. Plant Science 135(1):1-9.

Du W, Gardea-Torresdey JL, Ji R, Yin Y, Zhu J, Peralta-Videa JR, Guo H (2015). Physiological and biochemical changes imposed by CeO2 nanoparticles on wheat: a life cycle field study. Environmental Science & Technology 49(19):11884-11893.

Eckhardt U, Grimm B, Hörtensteiner S (2004). Recent advances in chlorophyll biosynthesis and breakdown in higher plants. Plant Molecular Biology 56(1):1-14.

Foltête AS, Masfaraud JF, Bigorgne E, Nahmani J, Chaurand P, Botta C, ... Cotelle S (2011). Environmental impact of sunscreen nanomaterials: ecotoxicity and gentoxicity of altered TiO2 nanocomposites on Vicia faba. Environmental Pollution 159(10):2515-2522.

Foyer CH, Halliwell B (1976). The presence of glutathione and glutathione reductase in chloroplast: a proposed role in ascorbic acid metabolism. Planta 133(1):21-25.

Gál J, Hursthouse A, Tatner P, Stewart F, Welton R (2008). Cobalt and secondary poisoning in the terrestrial food chain: data review and research gaps to support risk assessment. Environment International 34(6):821-838.

Giannopolitis CN, Ries SK (1977). Superoxide dismutase: I. occurrence in higher plants. Plant Physiology 59(2):309-314.

Gorczyca A, Pociecha E, Kasprowicz M, Niemiec M (2015). Effect of nanosilver in wheat seedlings and Fusarium culmorum culture systems. European Journal of Plant Pathology 142(2):251-261.

Habig WH, Pabst MJ, Jakoby WB (1974). Glutathione S-transferase. The first enzymatic step in mercapturic acid formation. The Journal of Biological Chemistry 249(22):7130-7139.

Hajra A, Mondal NK (2017). Effects of ZnO and TiO2 nanoparticles on germination, biochemical and morphoanatomical attributes of Cicer arietinum L. Energy, Ecology and Environment 2(4):277-288.

Hasanabadi T, Lack S, Modhej A, Ghafurian H, Alavifazel M, Ardakani MR (2019). Feasibility study on reducing lead and cadmium absorption by alfalfa (Medicago scutellata L.) in a contaminated soil using nano-activated carbon and natural based nano-zeolite. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 47(4):1185-1193.

Hong J, Peralta-Videa JR, Rico C, Sahi S, Viveros MN, Bartonjo J, ... Gardea-Torresdey JL (2014). Evidence of translocation and physiological impacts of foliar applied CeO2 nanoparticles on cucumber (Cucumis sativus) plants. Environmental Science & Technology 48(8):4376-4385.

Hossain MA, Piyatida P, da Silva JAT, Fujita M (2012). Molecular mechanism of heavy metal toxicity and tolerance in plants: central role of glutathione in detoxification of reactive oxygen species and methylglyoxal and in heavy metal chelation. Journal of Botany 2012:1-37.

Iannone MF, Groppa MD, de Sousa ME, van Raap MBF, Benavides MP (2016). Impact of magnetite iron oxide nanoparticles on wheat (Triticum aestivum L.) development: evaluation of oxidative damage. Environmental and Experimental Botany 131:77-88.

Jahani S, Saadatmand S, Mahmoodzadeh H, Khavari-Nejad RA (2018). Effects of cerium oxide nanoparticles on biochemical and oxidative parameters in marigold leaves. Toxicological and Environmental Chemistry 100(8-10):677-692.

Jahani S, Saadatmand S, Mahmoodzadeh H, Khavari-Nejad RA (2019). Effect of foliar application of cerium oxide nanoparticles on growth, photosynthetic pigments, electrolyte leakage, compatible osmolytes and antioxidant enzymes activities of Calendula officinalis L. Biologia 74(9):1063-1075.

Kalra YP (1998). Handbook of reference methods for plant analysis. CRC Press, Boca Raton, FL.

Khan MN (2016). Nano-titanium dioxide (nano-TiO2) mitigates NaCl stress by enhancing antioxidative enzymes and accumulation of compatible solutes in tomato (Lycopersicon esculentum Mill.). Journal of Plant Sciences 11(1-3):1-11.

Kheirizadeh Arough Y, Seyed Sharifi R, Sedghi M (2016). Effect of zinc and bio fertilizers on antioxidant enzymes activity, chlorophyll content, soluble sugars and proline in triticale under salinity condition. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 44(1):116-124.

Kitamura Y, Ohta M, Ikenaga T, Watanabe M (2002). Responses of anthocyanin-producing and non-producing cells of Glehnia littoralis to radical generators. Phytochemistry 59(1):63-68.

Kooti M, Gharineh S, Mehrkhah M, Shaker A, Motamedi H (2015). Preparation and antibacterial activity of CoFe2O4/SiO2/Ag composite impregnated with streptomycin. The Chemical Engineering Journal 259:34-42.

Kováčik J, Klejdus B, Hedbávný J, Štork F, Bačkor M (2009). Comparison of cadmium and copper effect on phenolic metabolism, mineral nutrients and stress-related parameters in Matricaria chamomilla plants. Plant and Soil 320:231-242.

Lee CW, Mahendra S, Zodrow K, Li D, Tsai YC, Braam J, Alvarez PJ (2010). Developmental phytotoxicity of metal oxide nanoparticles to Arabidopsis thaliana. Environmental Toxicology and Chemistry 29(3):669-675.

Lei Z, Mingyu S, Xiao W, Chao L, Chunxiang Q, Liang C, ... Fashui H (2008). Antioxidant stress is promoted by nano-anatase in spinach chloroplasts under UV-B radiation. Biological Trace Element Research 121(1):69-79.

López-Luna J, Camacho-Martínez MM, Solís-Domínguez FA, González-Chávez MC, Carrillo-González R, Martinez-Vargas S, ... Cuevas-Díaz MC (2018). Toxicity assessment of cobalt ferrite nanoparticles on wheat plants. Journal of Toxicology and Environmental Health Part A 81(14):604-619.

López-Moreno ML, Avilés LL, Pérez NG, Irizarry BÁ, Perales O, Cedeno-Mattei Y, Román F (2016). Effect of cobalt ferrite (CoFe2O4) nanoparticles on the growth and development of Lycopersicon lycopersicum (tomato plants). Science of the Total Environment 550:45-52.

Ma C, Liu H, Guo H, Musante C, Coskun SH, Nelson BC, ... Dhankher OP (2016). Defense mechanisms and nutrient displacement in Arabidopsis thaliana upon exposure to CeO2 and In2O3 nanoparticles. Environmental Science: Nano 3(6):1369-1379.

Ma C, White JC, Dhankher OP, Xing B (2015). Metal-based nanotoxicity and detoxification pathways in higher plants. Environmental Science & Technology 49(12):7109-7122.

MacAdam JW, Nelson CJ, Sharp RE (1992). Peroxidase activity in leaf elongation zone of tall fescue. Plant Physiology 99(3):872-878.

Majumdar S, Almeida IC, Arigi EA, Choi H, VerBerkmoes NC, Trujillo-Reyes J, ... Gardea-Torresdey JL (2015). Environmental effects of nanoceria on seed production of common bean (Phaseolus vulgaris): a proteomic analysis. Environmental Science & Technology 49(22):13283-13293.

Marchiol L, Mattiello A, Pošćić F, Fellet G, Zavalloni C, Carlino E, Musetti R (2016). Changes in physiological and agronomical parameters of barley (Hordeum vulgare) exposed to cerium and titanium dioxide nanoparticles. International Journal of Environmental Research and Public Health 13(3):332.

Matysik G, Wójciak-Kosior M, Paduch R (2005). The influence of Calendulae officinalis flos extracts on cell cultures, and the chromatographic analysis of extracts. Journal of Pharmaceutical and Biomedical Analysis 38(2):285-292.

Michalak A (2006). Phenolic compounds and their antioxidant activity in plants growing under heavy metal stress. Polish Journal of Environmental Studies 15(4):523-530.

Minz A, Sinha AK, Kumar R, Kumar B, Deep KP, Kumar SB (2018). A review on importance of cobalt in crop growth and production. International Journal of Current Microbiology and Applied Sciences 7:2978-2984.

Nair PMG, Chung IM (2015). Physiological and molecular level studies on the toxicity of silver nanoparticles in germinating seedlings of mung bean (Vigna radiata L.). Acta Physiologiae Plantarum 37(1):1719.

Nair PMG, Kim SH, Chung IM (2014). Copper oxide nanoparticle toxicity in mung bean (Vigna radiata L.) seedlings: physiological and molecular level responses of in vitro grown plants. Acta Physiologiae Plantarum 36:2947-2958.

Nakano Y, Asada K (1981). Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts. Plant and Cell Physiology 22(5):867-880.

Nath UK, Kim HT, Khatun K, Park JI, Kang KK, Nou IS (2016). Modification of fatty acid profiles of rapeseed (Brassica napus L.) oil for using as food, industrial feed-stock and biodiesel. Plant Breeding and Biotechnology 4(2):123-134.

Palit S, Sharma A, Talukder G (1994). Effects of cobalt on plants. The Botanical Review 60(2):149-181.

Pandey N, Pathak GC, Pandey DK, Pandey R (2009). Heavy metals, Co, Ni, Cu, Zn, and Cd, produce oxidative damage and evoke differential antioxidant responses in spinach. Brazilian Journal of Plant Physiology 21(2):103-111.

Pandey SK, Singh H (2011). A simple, cost-effective method for leaf area estimation. Journal of Botany 2011:6 pp.

Pauly N, Pucciariello C, Mandon K, Innocenti G, Jamet A, Baudouin E, ... Puppo A (2006). Reactive oxygen and nitrogen species and glutathione: key players in the legume-rhizobium symbiosis. Journal of Experimental Botany 57(8):1769-1776.

Pošćić F, Mattiello A, Fellet G, Miceli F, Marchiol L (2016). Effects of cerium and titanium oxide nanoparticles in soil on the nutrient composition of barley (Hordeum vulgare L.) kernels. International Journal of Environmental Research and Public Health 13(6):577.

Prakash M, Gopalakrishnan N, Chung IM (2017). Evaluation of stress effects of copper oxide nanoparticles in Brassica napus L. seedlings. 3 Biotech 7(5):293.

Prasad TNVKV, Sudhakar P, Sreenivasulu Y, Latha P, Munaswamy V, Raja Reddy K, ... Pradeep T (2012). Effect of nanoscale zinc oxide particles on the germination, growth and yield of peanut. Journal of Plant Nutrition 35:905-927.

Puła J, Barabasz-Krasny B, Lepiarczyk A, Zandi P, Możdżeń K (2019). Activity of the photosynthetic apparatus in Phaseolus vulgaris L. leaves under the cadmium stress. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 47(2):405-411.

Raymond J, Rakariyatham N, Azanza JL (1993). Purification and some properties of polyphenol oxidase from sunflowers seeds. Phytochemistry 34(4):927-931.

Rico CM, Hong J, Morales MI, Zhao L, Barrios AC, Zhang JY, ... Gardea-Torresdey JL (2013). Effect of cerium oxide nanoparticles on rice: a study involving the antioxidant defense system and in vivo fluorescence imaging. Environmental Science & Technology 47(11):5635-5642.

Rizwan M, Ali S, Qayyum MF, Ok YS, Adrees M, Ibrahim M, ... Abbas F (2017). Effect of metal and metal oxide nanoparticles on growth and physiology of globally important food crops: a critical review. Journal of Hazardous Materials 322(A):2-16.

Salehi H, Chehregani A, Lucini L, Majd A, Gholami M (2018). Morphological, proteomic and metabolomic insight into the effect of cerium dioxide nanoparticles to Phaseolus vulgaris L. under soil or foliar application. The Science of The Total Environment 616-617:1540-1551.

Schwab F, Bucheli TD, Lukhele LP, Magrez A, Nowack B, Sigg L, Knauer K (2011). Are carbon nanotube effects on green algae caused by shading and agglomeration? Environmental Science & Technology 45(14):6136-6144.

Sharma P, Jha AB, Dubey RS, Pessarakli M (2012). Reactive oxygen species, oxidative damage, and antioxidative defense mechanism in plants under stressful conditions. Journal of Botany 217037:1-26.

Singleton VL, Rossi JA (1965). Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. American Journal of Enology and Viticulture 16:144-158.

Skórska E, Grzeszczuk M, Barańska M, Wójcik-Stopczyńska B (2019). Long wave UV-B radiation and asahi SL modify flavonoid content and radical scavenging activity of Zea mays var. saccharata leaves. Acta Biologica Cracoviensia Series Botanica 61(1):87-92.

Skórska E, Murkowski A (2018). Photosynthetic responses of Chlorella vulgaris L. to short-term UV-B radiation exposure. Acta Biologica Cracoviensia Series Botanica 60(1):65-71.

Vicas SI, Cavalu S, Laslo V, Tocai M, Costea TO, Moldovan L (2019). Growth, photosynthetic pigments, phenolic, glucosinolates content and antioxidant capacity of broccoli sprouts in response to nanoselenium particles supply. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 47(3):821-828.

Wagner GJ (1979). Content and vacuole/extravacuole distribution of neutral sugars, free amino acids, and anthocyanin in protoplasts. Plant Physiology 64(1):88-93.

Wang H, Arakawa O, Motomura Y (2000). Influence of maturity and bagging on the relationship between anthocyanin accumulation and phenylalanine ammonia-lyase (PAL) activity in ‘Jonathan’ apples. Postharvest Biology and Technology 19(2):123-128.

Wheatherley PE (1950). Studies in the water relations of the cotton plant I. the field measurements of water deficit in leaves. New Phytologist 49:81-97.

Zhang P, Ma Y, Liu S, Wang G, Zhang J, He X, ... Zhang Z (2017). Phytotoxicity, uptake and transformation of nano-CeO2 in sand cultured romaine lettuce. Environmental Pollution 220(Pt B):1400-1408.

Zhang W, Ebbs SD, Musante C, White JC, Gao C, Ma X (2015). Uptake and accumulation of bulk and nanosized cerium oxide particles and ionic cerium by radish (Raphanus sativus L.). Journal of Agricultural and Food Chemistry 63(2):382-390.

Zhao L, Peng B, Hernandez-Viezcas JA, Rico C, Sun Y, Peralta-Videa JR, ... Gardea-Torresdey JL (2012). Stress response and tolerance of Zea mays to CeO2 nanoparticles: cross talk among H2O2, heat shock protein, and lipid peroxidation. ACS Nano 6(11):9615-9622.

Zhu Y, Xu F, Liu Q, Chen M, Liu X, Wang Y, … Zhang L (2019). Nanomaterials and plants: positive effects, toxicity and the remediation of metal and metalloid pollution in soil. Science of The Total Environment 662:414-421.

How to Cite
JAHANI, M., KHAVARI-NEJAD, R. A., MAHMOODZADEH, H., & SAADATMAND, S. (2020). Effects of cobalt oxide nanoparticles (Co3O4 NPs) on ion leakage, total phenol, antioxidant enzymes activities and cobalt accumulation in Brassica napus L. Notulae Botanicae Horti Agrobotanici Cluj-Napoca, 48(3), 1260-1275. https://doi.org/10.15835/nbha48311766
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