Journal of Microbiology Research 2014, 4(4): 161-169 DOI: 10.5923/j.microbiology .20140404.02 Toxicity of Binary Mixtures of Formulated Glyphosate and Phenols to Rhizobium Species Dehydrogenase Activity C. O. Nweke 1,* , N. C. Ahumibe 2 , J. C. Orji 1 1 Department of Microbiology, Federal University of Technology, P.M.B. 1526, Owerri, Nigeria 2 Department of Biotechnology, Federal University of Technology, P.M.B. 1526, Owerri, Nigeria Abstract Acute toxicities of formulated glyphosate (Roundup®) in binary mixtures with 2,4-dichlorophenol, 4-chlorophenol and phenol were determined based on inhibition of 2-(p-Iodophenyl)-3-(p-nitrophenyl)-5-phenyl tetrazolium chloride (INT)-dehydrogenase activity in Rhizobium species. The phenolic compound: glyphosate mixture ratios (%) were 100:0, 20:80, 50:50, 60:40, 20:80 and 0:100 for the respective mixtures in the concentration range of 0 – 5000 mg/l. The effective doses (IC p s) were estimated using hormetic dose-response model. The median inhibitory concentrations (IC 50 ) of the formulated glyphosate, phenol, 4-chlorophenol and 2,4-dichlorophenol were 661.614 ± 33.234 mg/l, 1064.190 ± 87.286 mg/l, 158.884 ± 8.185 mg/l and 46.378 ± 2.504 mg/l respectively. The joint action toxicity of the mixtures on test organism was evaluated with isobolographic representations and toxicity index (TI) model. The isobole analysis indicated additive interaction between glyphosate and phenol. With the exception of 20:80 of 2,4-dichlorophenol:glyphosate mixture ratio that was synergistic, other ratios of the mixture were additive. A synergistic (especially 20:80 4-CP:glyphosate mixture) and additive interaction was observed for some ratios of glyphosate and 4-chlorophenol mixtures. However, the TI of most isoboles are within the range of 0.5 – 2.0 and are considered additive. Synergistic and additive interaction of formulated glyphosate with intermediates of 2,4-D was possible against the dehydrogenase activity of Rhizobium species, an important soil bacterium. The dynamics of the toxic effects thus would depend on the relative amounts of these compounds. Keywords Roundup®, Herbicides, Dehydrogenase Activity, Phenols, Toxicity 1. Introduction Increased application of herbicides for improved and sustained agriculture has resulted in the contamination of agricultural soils with organic and inorganic pollutants. One of most commonly used herbicide is glyphosate (N-phosphonomethylglycine), a post-emergence herbicide. Glyphosate is desirable due to its effective control of weeds, rapid inactivation in soil and low mammalian toxicity [1, 2]. However, environmental concern over glyphosate has grown due to its undesirable side effects on non-target organisms and persistence in soil and groundwater [3]. Glyphosate inhibits amino acid synthesis in bacteria and fungi via the enzyme, 5-enolpyruvyl-shikimate-3-phosphatase synthase in shikimic acid pathway [4, 5]. However, applications of glyphosate at high rate have been reported to stimulate microbial respiration [6, 7]. Glyphosate is often used in mixture with other herbicides such as 2,4-dichlorophenoxyacetic acid (2,4-D) amines and esters to widen spectrum, improve herbicide action and save * Corresponding author: xrisokey@yahoo.com (C. O. Nweke) Published online at http://journal.sapub.org/microbiology Copyright © 2014 Scientific & Academic Publishing. All Rights Reserved cost [8, 9]. In one of its commercial formulations, Roundup®, glyphosate is formulated as isopropylamine (IPA) salt of glyphosate (36% glyphosate a.i.) and a surfactant, polyoxyethyleneamine (POEA). The increased toxicity of this glyphosate formulation in comparison to glyphosate is attributed to the surfactant component [10]. Field application of glyphosate at recommended rates have been generally found to be non- toxic to soil microorganisms [11]. Both antagonistic and synergistic effect of 2,4-D on glyphosate activity have been reported. Glyphosate and 2,4-D mixture has resulted in reduced herbicide activity against grass compared to glyphosate applied alone [12, 13] but 2,4-D enhanced the activity of glyphosate against Rape [12]. Glyphosate applied with 2,4-D increased leafy Spurge control by 10-fold after 3 months of exposure compared to glyphosate alone [8]. Combinations of glyphosate with 2,4-D was generally more effective against cutleaf eveningprimrose than glyphosate applied alone [14]. Synergistiic effect of 2,4-D and glyphosate mixture against Brazil Pusley has also been reported by Sharma and Singh [15]. The environmental fate of these herbicides includes degradation by plants and microorganisms, hydrolysis and photolysis. A wide range of microorganisms have been