Allelopathic Potential of Essential Oil from Bottle Brush (Callistemon lanceolatus DC.) on The Germination and Growth of Echinochloa crus-gall L.

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1 Interntionl Journl of Agriulturl Tehnology 2017 Vol. 13(7.3): Aville online ISSN Allelopthi Potentil of Essentil Oil from Bottle Brush (Cllistemon lneoltus DC.) on The Germintion nd Growth of Ehinohlo rus-gll L. Worhet Bunkoed *, Ptthrin Wihittrkrn nd Chmroon Losinwttn Deprtment of Plnt Prodution Tehnology, Fulty of Agriulturl Tehnology, King Mongkut s Institute of Tehnology Lndkrng, Bngkok 10520, Thilnd. Worhet Bunkoed, Ptthrin Wihittrkrn nd Chmroon Losinwttn (2017). Allelopthi Potentil of Essentil Oil from Bottle Brush (Cllistemon lneoltus DC.) on The Germintion nd Growth of Ehinohlo rus-gll L. Interntionl Journl of Agriulturl Tehnology 13(7.3): A study ws onduted to determine the llelopthi effets of essentil oil from ottle rush (Cllisremon lneoltus DC.) on seed germintion nd seedling growth of Ehinohlo rusglli (L.) Beuv. The essentil oil t onentrtions of 10, 15, 20 nd 25 µl/plte were loded on piee of filter pper tthed to the inner side of the over of the Petri-dish. The distilled wter ws used s the ontrol. The results showed tht the essentil oil from ottle rush t onentrtion of 25 µl/plte hd hightly inhiition effet on seed germintion nd seedling growth of E. rus-gll in ontinuous experiment, nturl produt heriide from C. lneoltus. Essentil oil in emusifile onentrtes formultion (50% tive ingredient; ECC). The EEC t onentrtion of µl/ml were iossyed on seed germintion of E. rus-glli. The results showed tht ECC t 0.8 µl/ml inhiited seed germintion y 36.11%. Additionlly, the effet ECC on seed imiition nd α-mylse tivities of E. rus-glli seed were studied. The result showed tht seed imiition nd α-mylse tivities of E. rus-glli seed inresed y prolonging time wheres deresed with inresing onentrtions of ECC. Keywords: Essentil oil, C. lneoltus, E. rus-glli, α-mylse, Imiition, Germintion Introdution The ontrol of weeds n e regrded s ritil issue for frmers. The interferene of weeds in griulturl res redues the qulity of griulturl produts nd uses huge eonomi loss to frms. Thus, the use of syntheti heriides hs eome very ommon prtie for the ontrol of weeds re eing despite its widespred use for the ontrol of weeds, there is extensive disussion of the dmge they use to oth mn nd the environment. (Chou et l., 2010). Therey, developing nturlly ourring grohemils to reple the syntheti grohemils eomes n interesting sujet in sustinle * Coressponding Author: Chmroon Losinwttn; E-mil ddress: klhmro@kmitl..th

2 griulture There hve een some studies out the effets of plnt synthesis for the use of heriide. llelopthy ws disovered y Molish nd ment tht plnt releses ompounds to either stimulte or inhiit the growth of other plnt growing in the sme hitt in nturl nd griulturl eosystem (Molish, 1937) The use of llelohemils involved in llelopthi intertions ould stisfy the requirements for weeds mngement nd rops protetion. Alves et l. (2014) reported tht the essentil oils of Cinnmomum zeylnium Ness, Lippi sidoides Chm. nd Cymopogom nrdus L. the voltile extrts of plnts inhiited seed germintion nd root growth of seedlings of Bidens pilos L., whih shows llelopthi potentil nd the onentrtion of 0.08% of oils used the overll deteriortion of the roots nd deth of seedlings of B. pilos. Rhimi et l. (2013) noted tht Allelopthi effet of some essentil oils on seed germintion of Lthyrus nnuus nd Vii villos it n relese poisonous effet on the L. nnuus nd V. villos it n e soon tht essentil oil of plnt hve tendeny to reple the syntheti grohemils. The ottle rush (Cllistemon lneoltus DC.) is genus of 34 speies of shrus elonging to fmily Myrtee found wildly only on the Austrlin ontinent ut now ll speies re endemi to Austrli. The genus is known in folk mediine for its ntiough, ntironhitis, nd insetiidl effets nd its voltile oils hs een used s ntimiroil nd ntifungl gents. Moreover ntiphylool, nemtiidl, lrviidl, pupiidl, ntithromoti tivities nd ntioxidnt tivities (Sumitr nd Shiv, 2014). Rsseifr et l. (2013) noted tht llelopthi effet of essentil oil extrted from leves of Eulyptus gloulus whih is the plnt in fmily Myrtee it n relese poisonous effet on the Amrnthus litoides nd Cynodon dtylon it n e soon tht essentil oil of plnt hve tendeny to reple the syntheti grohemils The ojetives of this study to e investigte the effets of essentil oil from ottle rush (Cllistemon lneoltus DC.) on seed germintion of Ehinohlo rus-glli (L.) Beuv. This pper my e of enefit for development s ioheriides in future. Mterils nd methods Plnt mteril The study ws rried out t King Mongkut s Institute of Tehnology Ldkrng, Bngkok, Thilnd. The essentil oil from the leves of C. lneoltus ws extrted y distilltion method with wter t 95 to 100 ºC for 2 hours. The experiments were onduted in lortory. Seeds of E. rus-glli were olleted from pddy fields t Phitsnulok, Thilnd, pled in the shde 2694

3 Interntionl Journl of Agriulturl Tehnology 2017 Vol. 13(7.3): t room temperture for 3 months, nd then inuted in hot-ir oven t 45 ºC for 48 h to rek their dormny (Poonpioonpipt et l., 2013). Inhiitory effet of the essentil oil from C. lneoltus on germintion nd seedling growth of E. glli seeds This study ws onduted under lortory onditions using Petridish test. The essentil oil t onentrtion of 10, 15, 20 nd 25µl were loded on piee of filter pper tthed to the inner side of the over of the Petri-dish. Distilled wter ws used s the ontrol. Petri dishes (9-m dimeter) were lined with two lyers of germintion pper moistened with 5 ml of distilled wter nd 20 seeds of E. rus-glli were pled. To test the inhiitory effet of C. lneoltus essentil oil, different mounts of oil Petri dishes ws dded on piee of filter pper tthed inside of the over Petri-dish pled in growth hmer t ºC, 12 h/12 h drk/light, nd reltive humidity of ~80%. The rte of germintion ws mesured every dy nd shoot nd root lengths of the seedlings were mesured 7 dys fter tretment. The perentge of germintion inhiition were lulted ording to the following eqution: (Chung et l., 2001) Inhiition (% of ontrol) = [1 - (smple extrts/ontrol) 100] Inhiitory effet of the nturl produt from C. lneoltus on germintion nd seedling growth of E. glli seeds The essentil oil in emulsifile onentrtes formultion (50% tive ingredient; ECC) ws prepred y mixing with tween80 nd DMF (dimethylformide). The tretments of ECC t onentrtions of 0.1, 0.2, 0.4 nd 0.8 µl/ml, distilled wter ws used s the ontrol. Petri dishes were lined with two lyers of germintion pper moistened with 5 ml of eh onentrtion tested nd 20 seeds of E. rus-glli were pled per Petri dish. Four replites were mintined per tretment in ompletely rndomized mnner in growth hmer with temperture of ºC, 12h/12h drk/light, nd reltive humidity of ~80%. After 7 dys the numer of seeds tht germinted were ounted, nd their roots nd shoots lengths were mesured. The inhiitory or stimultory perent ws lulted using the following eqution given y Chung et l. (2001). 2695

4 Inhiitory effet of the nturl produt from C. lneoltus on seed imiition nd α-mylse tivity of E. glli seeds Mesurement of seed imiitions ws performed using method of Turk nd Twh (2003) with slight modifitions. Four replites of 30 helthy seeds of E. rus-glli were weighed nd reorded s originl seed weight (W 1 ). These seeds will e seprtely germinted in ECC nd distilled wter s ontrol (ording to ove tretment). Seed weights were reorded s finl seed weight (W 2 ) for eh tretment nd exposure time. Wter uptke (seed imiition) perentge ws lulted from following eqution: Wter uptke (%) = [(W 2 - W 1 ) / W 1 ] x 100 Mesurement of tivity of α-mylse ws mde ording to the method of Bernfeld (1955) nd Sdsivm (1996). After mesuring wter uptke, seeds (30 seeds of E. rus-glli for one determintion) were homogenized with 4 ml ie-old solution of 0.1M ClCl 2 nd entrifuged t 10,000 rpm for 20 minutes t 4 ºC. Superntnt ws used s enzyme extrt. The α-mylse tivity ws ssyed y mesuring rte of genertion of reduing sugr from solule strh. The retion medium (3 ml) ontined 1 ml of 1% solule strh in ette uffter solution t ph 5.5 nd 1 ml of enzyme. The ssy medium ws inuted for 15 minutes t 37 ºC. The retion ws terminted y ddition of 1 ml DNS regent (40 mm 3.5-dinitrosliyli id, 0.4 N NOH nd 1M K-N trtrte), nd immeditely heted in oiling wter th for 5 minutes. The mixture ws ooled under running tp wter. The intensity of olor ws mesured s sorption t 560 nm in spetrophotometer. The experiment ws replited four times in ompletely rndomized design (CRD). A stndrd grph ws prepred using mltose, nd the mount of α-mylse present in smple were lulted from stndrd urve nd expressed s µmol mltose/min/g Sttistil nlysis The experimentl design ws ompletely rndomized with four repetitions. The dt were sujeted to nlysis of vrine nd when the effets of tretments showed signifint differenes (P <0.05), using Tukey s test. 2696

5 Iniition (% of ontrol) Interntionl Journl of Agriulturl Tehnology 2017 Vol. 13(7.3): Results nd Disussion Inhiitory effet of the essentil oil from C. lneoltus on germintion nd seedling growth of E. glli seeds The inhiitory effets of essentil oil form C. lneoltus on the germintion nd seedling growth of E. rus-glli shows in Figure 1. The results indited tht the germintion nd growth of E. rus-glli deresed signifintly under the effet of essentil oil. At onentrtion of 15µl, the germintion of E. rus-glli ws inhiited y 28.35%. By inresing the dose of pplition t 20 nd 25µl, the inhiition mgnitude ws inresed to nd 68.65%, respetively. In generl, the inhiitory effet ws more on root length thn on shoot length. At the highest onentrtion of 25 µl, root length ws ompletely inhiited, wheres shoot length deresed y 71.05%. Vishwkrm nd Mittl (2014) lso reported tht Eulyptus teretiornis hve high llelopthi tivity, essentil oil from leves with ws inhiited seed germintion nd seedling development of E. rus-glli seed Germintion Shoot length Root length d ontrol Conentrtions (µl/plte) Figure 1. Effets of essentil oil from C. lneoltus. on seed germintion nd seedling growth of E. rus-glli seeds. The vlues represent the mens. Different letters indite signifine differenes (p<0.05) etween tretments. 2697

6 Inhiition (% of ontrol) Inhiitory effet of the nturl produt from C. lneoltus on germintion nd seedling growth of E. glli seeds The effets of nturl produt from C. lneoltus (ECC) on germintion nd seedling growth oif E. rus-glli were showed in Figure 2. The germintion of E. rus-glli ws signifintly redued. In generl, dose-response reltionship ws oserved nd the emergene deresed with inresing in onentrtion of ECC. At the highest onentrtion of 0.8 µl/ml of ECC showed highly inhiition effet on shoot (40.06%) nd root length (44.66%), respetively. However, t the lowest onentrtion of 0.1 µl/ml, there were promotive effets on shoot nd root length. These results indite tht ECC ontins some inhiitors of the priniple of inhiiting germintion nd seedling growth. The results otined in the present study re prllel to erlier reports doumenting the growth inhiitory tivity of romti plnts, inluding Eulyptus speies whih is in the sme urreny s C. lneoltus nd their voltile oils. For instne, voltile oil ( mg/ml) from Eulyptus itriodor inhiit seedling growth nd redued dry weight umultion in Cssi oidentlis, Amrnthus viridis nd E. rus-glli y 50% (Btish et l., 2004). Also, it ws demonstrted tht the voltile oil from Eulyptus itriodor inhiits the germintion nd seedling growth of rgweed Prthenium hysterophorus. Thus, these ould e used for weeds mngement (Kohli et l., 1998) Germintion Shoot length 30 Root length d 0 d ontrol Conentrtions (µl/ml) Figure 2 Effets of nturl produt from essentil oil of C. lneoltus on seed germintion nd seedling growth of E. rus-glli seeds. The vlues represent the mens. Different letters indite signifine differenes (p<0.05) etween tretments. 2698

7 Seed imiition (%) Interntionl Journl of Agriulturl Tehnology 2017 Vol. 13(7.3): Inhiitory effet of the nturl produt from C. lneoltus on seed imiition nd α-mylse tivity of E. glli seeds The inhiition effet of nturl heriide produt from C. lneoltus on seed imiition nd α-mylse tivity of E. rus-glli seeds were shown in Figure 3. The perentge of imiition exhiited mrked inrese y prolonging the imiition periods. The time of imition in the ontrol seeds were out 12, 24 nd 36 hr, where 17.33, nd 28.07% respetively. For ll tretment onentrtions, no signifint differenes in imiition fter the 12, 24 nd 36 hr imiiton time. This is different from previous studies reported tht effet essentil oil from ssi (Cinnmomum ssi) on seed imiition of E. rus-glli exhiited seed inresed y prolonging time wheres deresed with inresing onentrtions of essentil oil (Ittiwehhi et l., 2014) Time (h) ontrol 0.1 µl/ml 0.2 µl/ml 0.4 µl/ml 0.8 µl/ml Figure 3 Effets of nturl heriide produt from C. lneoltus on seeds imiition of E. rus-glli seeds t different imiition periods. The vlues represent the mens. Different letters indite signifine differenes (p<0.05) etween tretments. 2699

8 α-mylse tivity (µmol mltose min/ g/fw) Time (h) ontrol 0.1 µl/ml 0.2 µl/ml 0.4 µl/ml 0.8 µl/ml Figure 4. Effets of nturl produt heriide from C. lneoltus on α- mylse tivity of E. rus-glli seeds t different imiition periods. The vlues represent the mens. Different letters indite signifine differenes (p<0.05) etween tretments. Figure 4 shows the tivities of α-mylse in E. rus-glli seeds treted with vrious onentrtions of ECC. The results showed tht α-mylse tivities inresed y prolonging time wheres deresed with inresing onentrtions of ECC. The tivities of α-mylse were signifintly inhiited t ll onentrtions tested for soking period of 12 nd 24 h. However, the tivity of α-mylse ws no signifintly inhiited t ll onentrtions for soking period of 36 h. This is different results from previous studies. Poonpioonpipt et l. (2013) noted tht effets of essentil oil from Cymopogon itrtus inhiit α-mylse tivities. At 36 h, the inhiitions of tivities in seeds treted with oil were % t onentrtions of 1-8 µl/petri-dish. Conlusion The results showed tht the essentil oil from C. lneoltus hd high inhiition effet on seed germintion nd seedling growth of E. rus-glli seeds. The inhiitory effet of nturl produt of essentil oil from C. lneoltus in emulsifile onentrtes formultion (50% tive ingredient; ECC) ws higher thn pure essentil oil. The inhiition on seed germintion of E. rus-glli used y inhiited seed imiition nd the tivity of α-mylse in E. rus-glli seeds. 2700

9 Interntionl Journl of Agriulturl Tehnology 2017 Vol. 13(7.3): Aknowledgement This reserh ws supported y grnts from Fulty of Agriulturl Tehnology, King Mongkut s Institute of Tehnology Lndkrng, Bngkok, Thilnd. Referenes Alves, MCS., Medeiros, FS., Mnoel NA., Brito, RC. nd Arujo, RC. (2014). Allelopthi effet of essentil oils of mediinl plnts in Bidens pilos L. Revist Brsileir de Plnts Mediinis 16(3): Btish, RD., Seti., N., Singh, HP. nd Kohli, RK. (2004). Phytotoxiity of lemon-sented eulypt oil nd its potentil use ioheriide. Crop Protetion. 23: Bernfeld, P. (1955). Amylses α nd β. In: Colowik, S.P., Kpln, N.O. (Eds.), Method in Enzymology. Ademi Press, New York: Chou, C.H. (2010). Role of llelopthy in sustinle griulture: Use of llelohemils s nturlly ourring io-grohemils. Allelopthy Journl 25:3-16. Chung, I.M., Ahn, J.K. nd Yun, S.J. (2001). Assessment of llelopthi potentil of rnyrd grss (Ehinohlo rus-glli) on rie (Oryz stiv L.) ultivrs. Crop Protetion 20: Ittiwehhi, A., Wihittrkrn, P., Chngswke, K., Teerrk, M. nd Losinwttn, C. (2014). Potentil of essentil oil from ssi (Cinnmomum ssi) on seed germintion, imiition nd α-mylse tivity of Ehinohlo rus-glli. pp In proeedings Seoul Interntionl Conferene on Biologil Engineering & Nturl Siene Courtyrd y Mrriott Seoul Times Squre, South Kore. Kohli, RK., Btish, DR. nd Singh, HP. (1998). Eulypt oil for the ontrol of prthenium (Prtheniumhysterophorus L.). Crop protetion 17: Molish, H. (1937). Der Einfluss einer Pflnze uf die ndere Allelopthie. Fisher, Jen. Poonpioonpipt, T., Pngnkorn, U., Suvunnmek, U., Teerrk, M., Chroenying, P. nd Losinwttn, C. (2013). Phytotoxi effets of essentil oil from Cymopogon itrtus nd its physiologil mehnisms on rnyrdgrss (Ehinohlo rus-glli). Industril Crops nd Produts 41: Rhimi AR., Mousvizdeh, SJ., Mohmmdi, H., Rokhzdi, A., Mjidi, M. nd Amini, S. (2013). Allelopthi effet of some essentil oils on seed germintion of Lthyrus nnuus nd Vii villos. Journl of Biodiversity nd Environmentl Sienes. 3(4): Rsseifr, M., Hosseini, N., Hsni Asl, NH., Zndi, P. nd Aghdm, MM. (2013). Allelopthi effet of Eulyptus gloulus essentil oil on seed germintion nd seedling estlishment of Amrnthus litoides nd Cyndon dtylon. Trki Journl of Sienes 1: Sdsivm, S. (1996). Biohemil methods: New Age Interntionl. Sumitr, S. nd Shiv. S. (2014). Genus Cllistemon: n updte review. World Journl of Phrmy nd Phrmetil Sienes. 3(7): Turk, MA. nd Twh, AM. (2003). Allelopthi effet of lk mustrd (Brssi nigr L.) on germintion nd growth of wild ot (Aven ftu L.). Crop Protetion 22: Vishwkrm GS.nd Mittl S. (2014). Bioheriidl potentil of essentil oil from leves of Eulyptus teretiornis ginst Ehinohlo rus - glli L. Journl of Biopestiides 7(Supp): (Reeived 5 Otoer 2017; epted 25 Novemer 2017) 2701