The Effect of Milk Thistle (Silybum marianum L.) Extract on Growth and Enzyme Activities of Common Cocklebur (Xanthium strumarium L.) Seedling

Document Type : Research Article


Islamic Azad University-Shoushtar Campus


Introduction Allelopathy means any process involving the secondary metabolites (allelochemicals) produced by plants, algae, bacteria, and fungi (excluding animals) that influences the growth and development of agricultural and biological systems, with positive or negative effects. Plants produce secondary metabolites and in certain circumstances, these can act as phytotoxins, inhibiting or promoting some biochemical or physiological processes in the other plants or organisms. Toxicity of allelochemical compounds include effects on growth, chlorophyll content, photosynthesis, transpiration, stomatal conductance, relative growth rate, and oxygen uptake in many crops. Furthermore, the influence of allelochemicals on alpha amylase, oxidase, sucrose synthase activity and malon dialdehyde concentration were analysed. The role of numerous plant secondary metabolites is still unclear, and this raises curiosity for both plant physiologist and ecologists. Inhibitory effects on germination and establishments of crops caused by residues of either crops or weeds have lead to investigation of the release of toxic compounds from such residues. This present study was designed to evaluate the allelopathic potential of milk thistle for controlling common cocklebur in vegetables.
Materials and Methods In order to investigate the effect of milk thistle aqueous extract on growth and enzyme activities of common cocklebur seedlings, two separate experiments were carried out in randomized complete design and randomized complete block designs with five and four replications, respectively, in spring 2012 in seed technology laboratory and greenhouse of Azad University, Shoushtar branch, Shoushtar, Iran. Milk thistle aqueous extract concentrations were involved (0, 5, 10 and 15% (v/v)) in petridish and (0, 10, 20 and 30% (v/v)) as spraying on common cocklebur seedlings under greenhouse conditions. Whole plants of milk thistle were gathered from medicine plant farm of Azad University. Then, their flowers were cut and their shoots were dried in 60 °C and grinded. For preparing aqueous extract 100 g powder of milk thistle dissolved in 1000 ml distilled water and maintained at 20 °C. The solution was filtered and cleared. The aqueous extract was considered as stock and other aqueous extracts were prepared from it. After application extract concentration in petridish and pot, seedling fresh weight, activity of sucrose synthase, alpha amylase, catalase enzymes and malon dealdehyde concentration of common cocklebur were measured. Statistical calculations were accomplished through the SPSS software and graphs were drawn by the Excel software.
Results and Discussion Results showed that fresh weight of common cocklebur seedlings were affected by milk thistle extract. Milk thistle extract reduced alpha amylase enzyme activity and seedling growth of common cocklebur. In addition, increase in aqueous extract of milk thistle resulted in reduction and increase in catalase and malon dialdehyde concentration in cocklebur seedling; respectively. The minimum alpha amylase enzyme activity (2.1 nM/g seed/min) and seedling fresh weight (0.32 g) were observed in 15% aqueous extract. Malon dialdehyde concentration of cocklebur seedlings was 0.4 (nM/g FW) in 15% aqueous extract of milk thistle. Catalase enzyme activity was the lowest (1.4 nM H2O2/mg Protein/min) and the greatest (27 protein absorption/60s) in 15% aqueous extract of milk thistle. In greenhouse experiment, also, with increasing aqueous extract of milk thistle was reduced seedling fresh weight, catalase and sucrose synthase enzyme activities. Catalase and sucrose enzyme activities were 1.8 (nM H2O2/mg Protein/min) and 2 (nM/ mg protein/min) at 30% aqueous extract of milk thistle. Futhermore, increase in aqueous extract of milk thistle resulting in increase cell memberane destruction and malon dialdehyde concentration in cocklebur seedling tissue. The lowest (0.0029 nM/g FW) and greatest (0.93 nM/g FW) malon dialdehyde concentration was at 30% aqueous extract.
Conclusions In this study, with increasing the amount of milk thistle extract, values of measured traits including fresh weight, sucrose synthase and catalase enzymes of cocklebur seedlings compared to control treatment (distilled water) have a decreasing trend, but malon dialdehyde concentration of cocklebur seedlings had increased compared to the control. The destruction of cell membranes under the influence of cocklebur allelopathic compounds can be one reason for the reduced growth of weed seedlings. Reduced sucrose synthase activity, led to reduced production of sucrose that was associated with reduced seedling growth. The present study confirmed that milk thistle can be used as an allelopathic plant for weed suppression in agroecosystems through its release of allelochemicals. Knowledge about the challenges related to the demonstration of allelopathy, as an ecological significant mechanism, is important in the assessment of ecological effects of allelopathic plants.


1- Avigad G. 1964. Sucrose-uridine diphosphate glucosyltransferase from Jerusalem artichoke tubers. Journal of Biological Chemistry, 239: 3613-3618.
2- Azizi G., Alimoradee L. and Rashedmahassel, M.H. 2006. Allelopatic effects of Bunium persicum and Cuminum cyminum essential oils on seed germination of some weeds species. Iranian Journal of Medicinal and Aromatic Plants, 22: 198-208. (in Persian with English abstract).
3- Bohm P.A.F., Zanardo F.M.L. and Ferrarese O. 2006. Peroxidase activity and lignification in soybean root growth-inhibition by juglone. Biology Plantarum, 50: 315-317.
4- Chance B. and Maehly A. C. 1995. Assay of Catalase and Peroxidases. Method in Enzymology, 2:764-775.
5- Counce P.A. and Gravois K.A. 2006. Sucrose Synthase Activity as a potential indicator of high rice grain yield. Crop Science, 46:1501-1508.
6- Daremi Zadeh N. 2012. Allelopathic effect of aqueous extract of barley (Hordeum vulgare) on germination, vegetative growth and activity of antioxidant and photosynthetical enzymes of wild barley (Hordeum spontaneum) and barnyardgrass (Echinochloa crus-galli). MSc thesis. Islamic Azad University, Shoushtar branch, Shoushtar, Iran.
7- Farhoudi R. 2012. Effect of safflower aqueous extracts foliar application on peroxidase enzyme activity and cell membrane leakage of hairy vetech. Pajouhesh and Sazandegi, 102: 45-53. (In Persian with English abstract).
8- Farhoudi R. and Lee D.J. 2012. Evaluation of sufflower (Carthamus tinctorius cv. Koseh) extract on germination and induction of a-amylase activity of wild mustard (Sinapis arvensis) seeds. Seed Science and Technology, 40: 134-138.
9- Farhoudi R. and Parham F. 2010. Investigation the allelopatic effects of aqueous extracts of safflower (Carthamus tinctorius) on peroxidase enzyme activity and leaf electrical leakage of hairy vetech (Vicia villosa). The Proceedings of 3rd Iranian Weed Science Congress, Babolsar, Iran. P. 383-386. (In Persian with English abstract).
10- Glenn A. 2008. Allelopathic Interference of Invasive Acacia dealbata on the rice seedling growth, 5th World Congress on Allelopathy, New York, USA.
11- Kaffashan E., Farhoudi R., Modhej A. 2010. The allelopatical effect of safflower on seedling characteristics and peroxidase enzyme activity in Hordeum spontaneum, Lolium spp., Phalaris spp. and wheat (Triticum aestivum). The Proceedings of 3rd Iranian Weed Science Congress, Babolsar, Iran. P. 508-511. (In Persian with English abstract).
12- Hossein Zadeh M., Kiarostami K., Ilkhani Zadeh M. and Saboora A. 2009. A study on allelopathic compounds derived from Hordeum spontaneum on carbohydrates, proteins and some enzymes of wheat (Triticum aestivum L.). Iranain Journal of Biology, 22: 392-406. (in Persian with English abstract).
13- Kato-Noguchi H. 2003. Assessment of allelopathic potential of shoot powder of lemon balm. Scientia Horticulturae. 97: 419-423.
14- Kourosh Nejad, N. 2012. The allelopatical effect of wild oat (Avena ludoviciana) and wheat (Triticum aestivum) on each other with emphasis on enzyme activities. MSc thesis. Islamic Azad Univercity, Shoushtar branch, Shoushtar, Iran.
15- Lorenzo P., Palomera-Pe´Rez A., Reigosa M.J. and Gonzal L. 2011. Allelopathic interference of invasive Acacia dealbata link on the physiological parameters of native understory species. Plant Ecology, 212: 403-411.
16- Maffei M., Bertea C.M., Garneri F. and Scanneri S. 1999. Effect of benzoic acid hydroxy- and methoxy-ring substituents during cucumber (Cucumis sativus L.) germination. I.: Isocitrate lyase and catalase activity. Plant Science, 141:139-147.
17- Mojab M. and Mahmoodi S. 2009. Allelopatic effects of shoot and root water extracts of hoary cress (Cardaria draba) on germination characteristic and seedling growth of Sorghum (Sorghum bicolor L.). Electronic Journal of Crop Protection, 1: 65-78. (in Persian with English abstract).
18- Morian M., Doa A. and Zhang L. 2009. Investigation the allelopathic effects of aqueous extracts of safflower on wheat germination. Weed Science Conference, Melbourne, Australia, 36-41.
19- Muscdo A., Panucci M. R. and Sidari M. 2001. The effect of phenols on respiratory enzymes in seed germination respiratory enzyme activities during germination of Pinus larico seeds treated with phenols extracted from different forest soils. Plant Growth Regulation, 35: 31-35.
20- Niakan M., Aroudi M. and Kiaei E. 2008. Investigating antioxidant and nitrate reductase enzyme activities of soybean and wheat in response to aqueous extract of canola under hydroponic conditions. Plant and Ecosystem. 13: 42-53. (in Persian with English abstract).
21- Oracz K., Bailly C., Gniazdowska A., Côme D., Corbineau D. and Bogatek R. 2007. Induction of oxidative stress by sunflower phytotoxins in germinating mustard Seeds. Journal of Chemichal Ecology, 33: 251-264.
22- Rezaie F. and Yarnia M. 2009. Allelopathic effects of Chenopodium album, Amaranthus retroflexus and Cynodon dactylon on germination and growth of safflower. Journal of Food, Agriculture and Environment. 7: 516-521. (in Persian with English abstract).
23- Rezaei Nodehi A., Khangholi S. and Nouri M. 2003. Allelopathic potential of Cardaria draba, Brassica reflexa and Brassica napus on germination and seedling growth of Mathiola incana and Amaranthus caudatus. Pajouhesh and Sazandegi, 60:65-71. (in Persian with English abstract).
24- Scott S. J., Jones R. A. and Williams W. A. 1984. Review of data analysis methods for seed germination. Crop Science. 24: 1192-1199.
25- Valentovic P., Luxova M., Kolarovi L. and Gasparikora O. 2006. Effect of osmotic stress on compatible solutes content, memberane stability and water relation in two maize. Plant Soil Enviroment. 52: 186-191.
26- Xiao Z., Storms R. and Tsang A. 2006. A quantitative starchiodine method for measuring alpha-amylase and gluco amylase activities, Analytical Biochemistry. 351: 146-148.
27- Yu J. Q., Fye S., Zhang M. F. and Hu W. H. 2003. Effects of root exudates and aqueous root extract of cucumber and allelochemicals on photosynthesis and antioxidant enzymes in cucumber. Biological Systems and Ecology, 31: 129- 139.