Turkish Journal of Agriculture - Food Science and Technology, 3(6): 472-477, 2015 Turkish Journal of Agriculture - Food Science and Technology www.agrifoodscience.com, Turkish Science and Technology The Symptoms of Herbicidal Action: The Case of Aclonifen Özgür Kılınç* Department of Plant Production and Technologies, Faculty of Agricultural Sciences and Technologies, Niğde University, 51240 Niğde, Turkey A R T I C L E I N F O A B S T R A C T The symptoms which were directly bound to the biochemical mode of action of an Article history: herbicide, other symptoms result from an indirect consequence of this action. The Received 20 March 2015 symptoms of herbicidal action deeply differed and that the climatic factors during the two Accepted 01 April 2015 first weeks after treatment could change definitely the result of the selective herbicide Available online, ISSN: 2148-127X action. The repetitive observation of symptoms allows to inform the farmer about the tolerance or resistance of certain plant species, including the culture, for instance through Keywords: the appearance of symptoms on the first leaves of the seedlings and their absence in the Aclonifen following leaves. As a whole, the accurate observation of herbicidal symptoms on plants Selectivity is the essential, rapid and non-expensive analysis of treatment effectiveness at the field Herbicide scale. The purpose of the current report is to describe the symptoms of a very complex Treatment herbicidal action, that of aclonifen involving two modes of action for the same molecule, Carotenoids approximately at the same concentration, and to compare these symptoms under field conditions and under controlled conditions, for a better understanding. * Corresponding Author: E-mail: [email protected] Introduction Selectively killed weed close to cultivated plants was founded on the structural analogy of the concerned which do have to remain unaffected, is the necessary herbicidal family with indol-3-acetic acid (I.A.A.) and on condition for using herbicides in agronomy (Tissut et al., the fact that these herbicides were chemically and 2006) biologically stable in marked contrast with I.A.A. This challenge is generally not a simple one and (Grossmann, 2000; PerrotRechenmann et al., 2005.) farmers do have to pay much attention to the effectiveness The observation of such symptoms by the farmers and selectivity of such a chemical operation which teached them that the herbicidal action may be rapid or depends on several factors, especially climatic slow, depending on the type of biochemical mode of (temperature, rain, hygrometry) and agronomic factors action, that the effectiveness of a herbicide is tightly (soil nature, development stage of the culture, weed bound to the development stage of the target-plant, that, species and nature of the planted cultivars). That is why in the two main strategies of treatment (pre-emergence the careful observation of the symptoms of the herbicidal and post-emergence), the symptoms of herbicidal action action on the culture and on the weeds is a precious deeply differed and that the climatic factors during the source of information (Scalla, 1991; Gauvrit, 1996) two first weeks after treatment could change definitely the Before the sixties, two main types of symptoms of result of the selective herbicide action. herbicidal actions were commonly observed which were Beside the symptoms which were directly bound to due to either membrane disruption (H2SO4, the biochemical mode of action of an herbicide, other dinitrophenols) or synthetic phytohormones (2, 4-D, 2, 4, symptoms result from an indirect consequence of this 5-T, etc.). Membrane disruption obtained through the action. For instance, the inhibition of cell division often structural change in the polar fatty acids induced rapid induces an increase in leaf chlorophyll concentration, water loss, oxidations and cell death on the places where probably due to a higher nitrogen disponibility (Devine et droplets of the herbicide solution are in close contact with al., 1993; Tissut et al., 2006). In the same way, the cells of the aerial parts. The major symptom is that of inhibition of protein synthesis may induce a strong dark spots suggesting a burning effect. (Fedtke, 1982; anthocyanin synthesis or the accumulation of the Bourdin, 1983; Cabanette, 1986; Hascoet and Bourdin, secondary pigments, as it is the case for glyposate (Franz 1988). et al., 1997) The main symptom of phytohormones action was the All these symptoms contribute to evaluate in the field disorganization of the stem elongation process which the effectiveness of a herbicide strategy and, often, to affected selectively dicotyledonous plants. This symptom deduce the type of herbicide which was used. Kılınç / Turkish Journal of Agriculture - Food Science and Technology, 3(6): 472-477, 2015 Furthermore, the repetitive observation of symptoms Materials and Methods allows to inform the farmer about the tolerance or Plant culture: Six plant species including Zea mays L. resistance of certain plant species, including the culture, var Furio, Triticum aestivum L., Alopecurus myosuroides for instance through the appearance of symptoms on the Hudson, Cucumis sativus L., Sinapis arvensis L. and first leaves of the seedlings and their absence in the Vicia faba L. were concurrently cultivated. The weed following leaves. seeds were supplied by Arbiotech, 35590 Saint Gilles, As a whole, the accurate observation of herbicidal France. The pre-emergent treatments were comparatively symptoms on plants is the essential, rapid and non- carried out either with pure aclonifen dispersed in water + expensive analysis of treatment effectiveness at the field 2% dimethylsulfoxide or with the formulated preparation scale. Challenge 600 dispersed in water (30 ml.m-2). The purpose of the current report is to describe the Plants (Alopecurus myosuroides and Brassica napus) symptoms of a very complex herbicidal action, that of were grown in a greenhouse with an electric heating aclonifen involving two modes of action for the same supply when temperature was lower than 10°C. A 150 molecule, approximately at the same concentration, and to µE.m-2.s-1.PAR light supply was added for 16 hours/day. compare these symptoms under field conditions and under In other experiments, plants (Zea mays, Cucumis sativus controlled conditions, for a better understanding. L. and Triticum aestivum) were grown in culture chamber Aclonifen (2-chloro-6-nitro-3-phenoxyaniline) is an (day: 16 h, 25°C; night: 8 h, 18°C R.H. 80%) and Vicia herbicide having the typical diphenylether structure faba and Sinapis arvensis were cultivated in the fields. (Figure 1). Among this chemical family numerous The formulated preparation Challenge 600 (with derivatives are known herbicides inhibiting one enzyme surfactant and other formulating agents) was used as a protoporphyrinogen oxidase (EC 1.3.3.4) in the preplant treatment at different doses, among which 2.7 kg biochemical pathway leading to chlorophyll or a.i/ha in 300 l water which is the conventional cytochromes biosynthesis (Matringe et al., 1989, Witkowski concentration for agronomic uses in France. and Halling, 1989; Arnould and Camadro, 1998). 14C labelling:14C aclonifen (1167 Bq.nmol-1) was a Several of these diphenylether derivatives were generous gift from Bayer CropScience AG Wuppertal authorized for agronomic uses (acifluorfen, lactofen, Germany. Unlabelled aclonifen was extracted from the oxyfluorfen, etc.) in different countries. One of them, commercial product “Challenge 600” (Bayer nitrofen has been forbidden since the end of the XXth CropScience) containing 600g/l of a.i. The acetonic century (Tomlin, 2006; Tissut et al, 2006) extract was fractionated with petrol ether 40-60°C. The As shown in Figure 1, aclonifen has a specific latter solution was evaporated to dryness, dissolved in the substitution allowing a good inhibitory activity on the minimal amount of acetone and stored at 4°C until chlorophyll synthesis pathway but, at the same crystallization. This process was repeated three times and concentration, it has also a noticeable inhibitory activity gave pure aclonifen as controlled by TLC on carotenoid biosynthesis. These two modes of action chromatography and spectrophotometry. determine a target space inside the plants (Figure 2) Critical concentration measurement: Plants were which is limited to the aerial parts (Kilinc et al, 2009). treated by 14C aclonifen at different concentration. Plant Furthermore, these two modes of action seem fresh samples (aerial and underground parts separately) complementary or even synergistic as they contribute were submitted to three successive extractions with together to induce membrane disruption through; acetone. The acetonic solutions were evaporated to 1 The emission of toxic O2 resulting from plastidial dryness and redissolved in 70% EtOH. An aliquot of the protoporphyrinogen oxidase inhibition (Matrige et dry residual pellet was dissolved with 10 mL of al., 1989; Dayan et al., 2001). mineralization mixture, H2O2/perchloric acid/H2O (1/1/1, The suppression of the carotenoid plastidial v/v/v), in closed polyethylene flasks. After 2-3 days of accumulation playing the role of ROS scavengers mineralization at 25°C, the radioactivity of the for the biological membranes (Böger and mineralization mixture as well as that of ethanol 70% Sandmann, 1992; Edge et al., 1997; Kilinc et al., solutions were measured through scintillation counting 2011). (1414 Winspectral EG&G Wallac), using PerkinElmer ULTIMA GOLD AB liquid. During the mineralization Such a double complementary mode of action 14 step, the absence of CO2 leaching from the flasks was expressed with its highest effectiveness in young plants controlled using a KOH trap. The critical concentration leads to choose the pre-emergence treatment as the best was the lower concentration inducing lethal symptom. agronomic strategy. This requires to treatment of the Autoradiography: TLC chromatography was carried superficial soil layer just after planting the seeds of the out on silica gel thin layer plates (Macherey-Nagel TLC culture. Afterwards, aclonifen has to be absorbed by the plates SIL G-25 0.25 mm silica gel).