EFFECT OF DIFFERENT WHEAT STRAW EXTRACTS ON GERMINATION AND GROWTH OF THREE DOMINANT WEED SPECIES

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1 International Journal of Botany and Research (IJBR) ISSN Vol. 3, Issue 2, Jun 2013, TJPRC Pvt. Ltd. EFFECT OF DIFFERENT WHEAT STRAW EXTRACTS ON GERMINATION AND GROWTH OF THREE DOMINANT WEED SPECIES P RAJPUT & P B RAO Department of Biological Sciences, College of Basic Sciences & Humanities, G.B. Pant University of Agriculture & Technology, Pantnagar, Uttarakhand, India ABSTRACT The phytotoxic potential of straw of 15 different varieties of wheat (Triticum aestivum L.) against three dominant weed species viz., Avena ludoviciana Durieu., Celosia argentea L., and Echinochloa crus-galli (L.) P. Beauv. was examined under laboratory conditions. The aqueous extracts (10%) exhibited inhibitory as well as stimulatory effect on rate of germination, per cent seed germination and growth of different weed species. Among the varieties, RAJ 4077 was more effective in reducing > 50% germination, while both MACS 6272 and PBW 550 exhibited maximum inhibitory effect on length and dry weight of different weed species. HPLC analysis of different extracts indicated that the maximum phenolics (six) were found in MACS 6272 followed by PBW 550, RAJ 4014 and RAJ 4077 (five, each) and minimum in both RAJ 4048 and WH 1003 (two). Thus, these wheat varieties can be utilized for controlling these weed species in the crop fields and also useful for the preparation of natural phyto-chemicals for sustainable agriculture productivity. KEYWORDS: Phytotoxic, Eco-friendly, Wheat Varieties, Weed Species, Sustainable Agriculture INTRODUCTION Wheat is one of the vital food crops in India which contributes nearly 35 % to national food basket. Increase in productivity is the only option, to pull off the food refuge with the escalating population. The mismatch between population growth and cereal production has led India in a paradox where huge food surpluses at the aggregate national level coexist with large undernourished and poor population. Weeds are major impediment for crop production systems in all over the world, particularly in India. Weed control is attained by manual weeding or chemical herbicides leads to the increased labour input, cost of chemical control, resistance in weeds, population shifts of weed species, abridged accessibility of new herbicide molecules and issues related to eco-friendly environment (Chauhan, 2012). Singh et al. (2001) opined that allelopathy is influence of one plants on neighbouring plants through the release of allelochemics that plays an important role in many agro-ecosystems. Wheat (Triticum aestivum L.) is a potent allelopathic plant and its phytotoxicity has been attributed to hydroxamic acids (Wu et al., 2000 a; Villagrasa et al., 2006) as well as phenolic acids (Wu et al., 2001). Its aqueous extracts were also allelopathic to Ipomoea lacunosa, Ambrosia artemisiifolia, and also to herbicide susceptible and resistant biotypes of Lolium rigidum (Liebl & Worsham, 1983; Wu et al., 2000 b). Wheat straw can equip selective weed control upon decomposition or due to its physical presence on the soil surface thus shading and altering the red to far red ratio of transmitted light (Bhowmik & Inderjit, 2003). Thus, the present study was designed to evaluate the phytotoxic effect of wheat straw extracts on germination and growth (both length and dry weight) against three dominant weed species of Terai region of Uttarakhand Himalaya, India. MATERIALS AND METHODS Straw of 15 different varieties (DBW 14, JOB 668, K 9006, KAUZ/STAR, MACS 6272, PBW 550, PBW 588,

2 72 P Rajput & P B Rao RAJ 4014, RAJ 4026, RAJ 4048, RAJ 4077, UP 2382, UP 2526, UP 2565 and WH 1003) of wheat (Triticum aestivum L.) and seeds of three dominant weed species viz., Avena ludoviciana Durieu., Celosia argentea L., Echinochloa crus-galli (L.) P. Beauv. were collected from the fields of Crop Research Centre (C.R.C.), G.B. Pant University of Agriculture & Technology, Pantnagar, India and brought to the laboratory for experimentation. Preparation of Aqueous Extracts Aqueous straw extracts ((10 %, w/v) of 15 wheat varieties were prepared by taking 10 g of powdered straw in 100 ml sterilized distilled water and kept for 24 h. The solutions were filtered through four layers of cheese cloth and centrifuged at 3000 rpm for 4 h and the supernatant was filtered by using Whatman No. 42 filter paper and finally made 100 ml by using distilled water for germination experiments. Healthy seeds were surface sterilized with 0.01% mercuric chloride (HgCl 2 ) solution for 5 min and washed thoroughly with distilled water and placed on double filter papers layered Petri dishes (9 cm dia.) and moistened either with distilled water (control) or extracts (treatments) in triplicates and kept at laboratory conditions (temperature 25 ± 1ºC and diffused light). Germination was recorded at an interval of 24 h up to 9 days. After 9 days, three seedlings from each Petri dish were selected and their length (cm) and dry weight (mg) were measured. The per cent reduction was calculated by using the following formula: Per cent reduction (%) = [(Control mean - Treatment mean)/control mean] x 100. Germination Relative Index (GRI) was calculated according to Srivastava and Sareen (1972): GRI = Xn (h-n), Where, Xn = number of seeds germination at n th count, h = total number of counts, and n = count number IDENTIFICATION OF PHENOLICS TROUGH REVERSE PHASE HPLC Phenolics were identified by acid hydrolysis procedure given by Harbourne (1977) and Charpentier and Cowles (1981). Powdered straw of each verity was extracted with diethyl ether, anhydrous sodium sulphate (Na 2 SO 4 ), active charcoal and filtratered and re-dissolved in 5 ml MeOH (HPLC grade) and filtered through Millipore filters (0.22 µ) and analyzed by HPLC. The standards of different phenolic acids (HPLC grade) such as cinnamic, chlorogenic, benzoic, gallic, ferulic, p-hydroxybenzoic, salicylic, syringic and vanillic were used for identification of compounds in sample. HPLC system (Dionex 300), comprising LC -10 AT VP pumps, SCL - 10 A VP auto injector and Phenomenex Luna C 18, 5 µm, (250 x 4.6 mm) column was used at ambient temperature. Isocratic elution was carried out with mobile phase, water : MeOH (77:23) consisting 2% glacial acetic acid at flow rate of 1 ml/min. STATISTICAL ANALYSES All data were analysed by one way analysis of variance (ANOVA). The mean values were separated on the basis of least significant difference (LSD) at the 0.05 probability level. RESULTS AND DISCUSSIONS Seed Germination The application of different wheat straw extracts significantly (P< 0.01) inhibited rate of germination in all three weed species (Figs. 1 a - c). The rate of seed germination was delayed in all weed species with all extracts except in A.

3 Effect of Different Wheat Straw Extracts on Germination and Growth of Three Dominant Weed Species 73 ludoviciana with UP 2526 and C. argentea with RAJ 4048, in which it was stimulated/promoted. Interestingly, the inhibition in seed germination was with all the varieties (100 %) in E. crus-galli; 14 (93.3%) in C. argentea; and 13 (86.7%) in A. ludoviciana (Table 1). Interestingly, more than 50 % reduction in seed germination was observed in all the three weed species and it was with maximum and minimum number of extracts in E. crus-galli (five) and A. ludoviciana (two), respectively. The extracts of RAJ 4077 caused maximum inhibitory effect (> 50%) on seed germination in all weed species. However, the extracts of RAJ 4048 exhibited stimulatory effect in two weed species and none of the extract in E. crus-galli. The Germination Relative Index (GRI) values were maximum and minimum in E. crus-galli with RAJ 4026 (18.34) and PBW 550 (3.00), respectively. The highest GRI values in different weed species with different extracts followed the order: RAJ 4026 in E. crus-galli (18.34) > UP 2382 in A. ludoviciana (18.00) > both RAJ 4026 and UP 2565 in C. argentea (14.00) (Fig. 2). Similar observations were also reported by Satsangi et al. (2010) and reported that the inhibitory effect of wheat straw on seed germination in Chenopodium album L., Boerhaavia diffusa L. and Parthenium hysterophorus L. Nikneshan et al. (2011) reported inhibitory effect of wheat extracts on germination of different weed species. Thus, the present results indicated that the extracts of MACS 6272, RAJ 4014 and RAJ 4077 caused maximum reduction on per cent seed germination in all weed species whereas the extracts of RAJ 4048 was least affective. Hence, it can be explained that former varieties possess ininate allelopathic potential or phytotoxicity compared to later. (a) A. ludoviciana (b) C. argentea

4 74 P Rajput & P B Rao (c) E. crus-galli Figure 1 a-c: Effect of Different Rice Extracts on Rate of Seed Germination in Three Different Weed Species 1= DBW 14; 2= JOB 668; 3= K 9006; 4= KAUZ/STAR; 5= MACS 6272; 6= PBW 550; 7= PBW 588; 8= RAJ 4014; 9= RAJ 4026; 10= RAJ 4048; 11= RAJ 4077; 12= UP 2382; 13= UP 2526; 14= UP 2565; 15= WH 1003 Figure 2: Effect of Different Wheat Straw Extracts on Germination Relative Index (GRI) in all Weed Species Length (cm) Radicle and plumule length was significantly (P< 0.01) inhibited with different extracts (Table 1). The varieties, MACS 6272 and PBW 550 exhibited maximum per cent inhibition (76) in A. ludoviciana. However, K 9006 and RAJ 4048 showed positive effect in all weed species except in E. crus-galli. The maximum average (average of all three weed species) per cent inhibition (60 %) was with MACS 6272 and PBW 550 and least with RAJ 4048 (1%) and K 9006 (3%), respectively on radicle and plumule length. These results are in agreement with Zwain (1999) and Al Saadawi (2001), who

5 Effect of Different Wheat Straw Extracts on Germination and Growth of Three Dominant Weed Species 75 reported the inhibitory effect of aqueous extracts of wheat residues on different weed species. Root length may be a key parameter to verify allelopathic strength of different wheat varieties because it had high sensitivity to allelochemicals. Randhawa et al. (2002) examined sorghum water extracts on Trianthema portulacastrum and reported inhibition on germination, root and shoot length. Hashem and Adkins (2010) studied the allelopathic effect of 19 accessions of Triticum speltoides on Avena fatua L. and eight on Sisymbrium orientate L. Thus, the present study results indicated inhibitory or stimulatory effect of different wheat extracts exhibited selective response of allelochemicals on the radical and plumule length of different weed species. Dry Weight (mg) The inhibition (%) in radicle and plumule dry weight was maximum with MACS 6272 (69) and RAJ 4077 (67) in A. ludoviciana and minimum with JOB 668 (2) in E. crus-galli and UP 2526 (1.76) in C. argentea, respectively (Table 2). However, among the extracts, JOB 668 in A. ludoviciana and RAJ 4048 in both A. ludoviciana and C. argentea showed positive effect. The inhibition (%) on total seedling dry weight was maximum with RAJ 4077 (67) in A. ludoviciana and minimum with UP 2382 (3) in C. argentea. However, positive effect was observed with the extracts of JOB 668 in A. ludoviciana and RAJ 4048 in C. argentea. The extracts of MACS 6272 exhibited the maximum average (average of all three weed species) per cent inhibition (53.92%) while minimum with WH 1003 (15.31%). Thus, the straw extracts of MACS 6272, PBW 550 and RAJ 4077 caused maximum inhibitory effect on dry weight (radicle, plumule and total) in different weed species. Ma (2005) studied the reduction in seedling dry weight of Amaranthus retroflexus, Stellaria media and Digitaria ciliaris, when treated with different wheat extracts. The suppressive phytotoxic effects of wheat against both grassy and broad leaved weeds are also reported by Mathiassen et al. (2006) and Labbafi et al. (2010). Khaliq et al. (2012) reported the highest suppression in shoot and root dry weight (58 and 67%, respectively) by wheat straw extracts. The straw of different crops contains phenolic acids, hydroxamic acids, terpenoids, alkaloids and other secondary metabolites. In the present study, different phenolic acids were identified in all 15 wheat varieties through reverse phase HPLC. 1= DBW 14; 2= JOB 668; 3= K 9006; 4= KAUZ/STAR; 5= MACS 6272; 6= PBW 550; 7= PBW 588; 8= RAJ 4014; 9= RAJ 4026; 10= RAJ 4048; 11= RAJ 4077; 12= UP 2382; 13= UP 2526; 14= UP 2565; 15= WH Figure 3: Effect of Different Wheat Straw Extracts on Total Seedling Dry Weight of Different Weed Species Identification of Phenolics HPLC analysis showed that the highest phenolics (six) was identified in MACS 6272 followed by PBW 550, RAJ 4014 and RAJ 4077 (five, each) and lowest in both RAJ 4048 and WH 1003 (two each) (Table 3). Among the phenolics,

6 76 P Rajput & P B Rao the presence of gallic acid was more frequent (14 varieties) while both syringic and cinnamic acids seldom (two varieties). Interestingly, chlorogenic acid could not be detected in any wheat variety. On the basis of presence of number of different phenolics in different varieties of wheat followed the order: MACS 6272 > PBW 550 = RAJ 4014 = RAJ 4077 > DBW 14 = JOB 668 = K 9006 = PBW 588 = RAJ 4026 = UP 2526 = UP 2565 > KAUZ/STAR = UP 2382 > RAJ 4048 = WH Likewise, Wu et al. (2000 b) identified p-hb, trans-p-coumaric, cis-p-coumaric, syringic, vanillic, trans-ferulic and cisferulic acids; and 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) from wheat seedlings. Mpofu et al. (2006) detected vanillic, caffeic, syringic, p-coumaric, ferulic and o-coumaric acids from spring wheat. CONCLUSIONS The present study results revealed that the varieties- MACS 6272, PBW 550 and RAJ 4077 with maximum number of phenolics exhibited significant inhibitory effect on different parameters in different weed species while the varieties- both RAJ 4048 and WH 1003 which exhibited minimum number of phenolics exerted stimulatory effect. The disparity in reduction was clearly ascribed to the genetic differences and presence of phenolics among the wheat varieties, since the extracts were of same concentration. Thus, these varieties can be exploited for the control of these weed species in the crop fields of Terai region and also for the development of natural bio-herbicides for the sustainable agricultural productivity and eco-friendly environment. REFERENCES 1. Alsaadawi, I. S. (2001). Allelopathic influence of decomposing wheat residues in agroecosystems. In: Allelopathy in agro-ecosystems. New York: Haworth Press, pp Bhowmik, P. C., & Inderjit. (2003). Challenges and opportunities in implementing allelopathy for natural weed management. Crop Protection, 22, Chauhan, B. S. (2012). Weed ecology and weed management strategies for dry-seeded rice in Asia. Weed Technology, 26, Chon, S. U., & Kim, Y. M. (2004). Herbicidal Potential and Quantification of Suspected Allelochemicals from Four Grass Crop Extracts. Journal of Agronomy and Crop Science, 190, Hashem, A., & Adkin, S. W. (2010). Introduction of allelopathy to wheat: effect of triticum speltoides on two weeds of wheat. Eleventh Australian Weeds Conference Proceedings, Pp Khaliq, A., Matloob, A., Aslam, F., Mushtaq, M. N., & Khan, M. B Toxic action of aqueous wheat straw extract on horse purslane. Planta Daninha, Viçosa-MG, 30(2), Labbafi, M. R., Assadollah, H. A., Maighany, F., Khalaj, H., & Mehrafarin, A. (2010). Evaluation of allelopathic potential of Iranian wheat (Triticum aestivum L.) cultivars against weeds. Agriculture and Biology Journal of North America, 1(3), Liebl, R. A., & Worsham, A. D. (1983). Inhibition of pitted morning glory (Ipomoea lacunosa L.) and certain other weed species by phytotoxic components of wheat (Triticum aestivum L.) straw. Journal of Chemical Ecology, 9, Ma, Y. Q. (2005). Allelopathic studies of common wheat (Triticum aestivum L.). Weed Biology and Managment, 5,

7 Effect of Different Wheat Straw Extracts on Germination and Growth of Three Dominant Weed Species Mathiassen, S. K., Kudsk, P., & Mogensen, B. B. (2006). Herbicidal effects of soil-incorporated wheat. Journal of Agriculture and Food Chemistry, 54(4), Mpofu, A., Sapirstein, H. D., & Beta, T. (2006). Genotype and environmental variation in phenolic content, phenolic acid composition and Antioxidant activity of hard spring wheat. Journal of agriculture and Food Chemistry, 54, Nikneshan, P., Karimmojeni, H., Moghnibashi, M., & Hosseini, N. A. S. (2011). Allelopathic potential of sunflower on weed management in safflower and wheat. Australian Journal of Crop Science, 5, Satsangi, G. P., Bartariya, G., & Saxena, A. (2010). Allelopathic response of wheat (Triticum aestivum L.) straw on germination and growth of selected weed species. Seventeenth Australasian Weeds Conference, pp Singh, H. P., Kohli, R. K., & Batish, D. R. (2001). Allelopathy in agro-ecosystems: an overview. Journal of Crop Production, 4, Villagrasa, M., Guillamon, M., Labandeira, A., Taberner, A., El jarrat, E., & Barcelo, D. (2006). Benzoxazinoid allelochemicals in wheat: distribution among foliage, roots, and seeds. Journal of Agriculture and Food Chemistry, 54, Wu, H., Haig, T., Pratley, J., Lemerle, D., & An, M. (2000a). Allelochemicals in wheat (Triticum aestivum L.): variation of phenolic acids in root tissues. Journal of Agriculture and Food Chemistry, 48, Wu, H., Haig, T., Pratley, J., Lemerle, D., & An, M. (2000b). Laboratory screening for allelopathic potential of wheat (Triticum aestivum) accessions against annual ryegrass (Lolium rigidum). Australian Journal of Agriculture and Research, 51, Wu, H., Haig, T., Pratley, J., Lemerle, D., & An, M. (2001). Allelochemicals in wheat (Triticum aestivum L.): production and exudation of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one. Journal of Chemical Ecology, 8, Zwain, K. H. Y., Alsaadawi, I. S., & Shahata, H. A Effect of decomposing wheat residues on growth and APPENDICES biological nitrogen fixation of blue green algae. Allelopathy Journal, 6, Table 1: Allelopathic Effect of Different Wheat Straw Extracts on Per Cent Reduction in Seed Germination, Radicle and Plumule Length of Different Weed Species Wheat Varieties % Reduction Seed Germination Radicle Length Plumule Length C. E. E. A. A. C. C. Argent crusgallgalli crus- ludovician ludoviciana Argentea Argentea ea a A. ludovici ana E. crus-galli JOB K KAUZ/STAR MACS PBW PBW RAJ RAJ RAJ

8 78 P Rajput & P B Rao Table 1:Contd., RAJ UP UP UP WH means stimulation. Table 2: Allelopathic Effect of Different Wheat Straw Extracts on Per Cent Reduction in Radicle and Plumule Dry Weight of Diferent Weed Species % Reduction in Dry Weight Wheat Radicle Plumule Varieties A. C. E. C. E. A. ludoviciana ludoviciana argentea crus-galli argentea crus-galli DBW JOB K KAUZ/STAR MACS PBW PBW RAJ RAJ RAJ RAJ UP UP UP WH Table 3: Identification of Various Phenolic Acids in the Straw of Different Varieties of Wheat by HPLC Wheat Phenolic Acids Varieties Number Name DBW 14 4 Gallic, Benzoic, p-hb and Vanillic JOB Gallic, p-hb, Caffeic and Vanillic K Gallic, p-hb, Caffeic and Vanillic KAUZ/STAR 3 Gallic, Ferulic and Salicylic MACS Cinnamic, Gallic, Benzoic, p-hb, Salicylic and Syringic PBW Gallic, Benzoic, p-hb, Caffeic and Vanillic PBW Gallic, p-hb, Caffeic and Vanillic RAJ Gallic, Ferulic, Salicylic, Vanillic and Syringic RAJ Gallic, p-hb, Caffeic and Salicylic RAJ Benzoic and Salicylic RAJ Cinnamic, Gallic, Ferulic, Salicylic and Vanillic UP Gallic, Ferulic and Salicylic UP Gallic, p-hb, Caffeic and Vanillic UP Gallic, Ferulic, Salicylic and yringic WH Cinnamic and Gallic