SORPTION KINETICS OF ISOPROTURON AND ASSESSMENT OF ITS ECOTOXICITY ON Lemna minor

The study was carried out to investigate the ecotoxicity of Isoproturon on Lemna minor and the interaction of the EC 50 value of the chemical with sorption behaviour of the chemical in soil. The sorption isotherms (K f and K OC ) for Isoproturon were determined for three different soils having various organic carbon and clay content. EC 50 for Lemna was determined both with and without soil. Both the Freundlich adsorption coefficient (K f ) and normalized sorption coefficient (K OC ) values varied with different types of soil. There was moderate correlation between the log K f and log K OC existed. The regression study revealed a strong relationship between log K OC and organic carbon and between log K OC and soil clay content. There EC 50 value for Lemna grown with soil was higher than that grown without soil. However, the difference was statistically insignificant. Greater degree of inconsistency in various data suggests the reiteration of the study. DOI: http://dx.doi.org/10.3329/ralf.v1i1.22347 Res. Agric., Livest. Fish. 1 (1): 13-18, Dec 2014


INTRODUCION
Isoproturon (IPU) is a widely used phenylurea derived systemic herbicide [3-(4isopropylphenyl)-1, 1-dimethylurea] for the control of annual grasses and broad-leaved weeds in agricultural fields.This herbicide is specific for monocots and mainly inhibits the electron transport in photosystem II and thereby disrupts the photosynthetic pathway of the target plant (Arnaud et al., 1994).It is one of the most common herbicide species, applied and to be found in water samples.Though IPU has a short environmental half-life (soil DT50 6-28 d) (Tomlin, 2001); reports on frequent finding of IPU in both surface and ground water at concentrations exceeding the European Union limit for drinking water (0.1 µg liter -1 ) (Johnson et al., 2001;Spliid and Køppen, 1998;Stangroom et al., 1998) has arisen the issue of environmental concern.
Sorption is one of the soil phenomena which directly affects the retention, volatilization, biodegradation, transport of organic chemicals in soil and uptake by plants.Therefore, the sorption constant is an important parameter used in a variety of models for predicting the ecotoxicity of organic chemicals entering in the environment (Wu et al., 2000;Blume and Ahlsdorf, 1993).Chiou et al. (1979) and Green and Karickhoff (1990) observed that the value of KOC for a compound measured in different soils approach towards a constant, and thus the Kf value for any given compound is closely related to the organic carbon constant of the soil.Sorption behaviour of a chemical in soil may assist in predicting the toxicity in the natural environmet.Because, it help simulates the actual amount of exposure of an organism in the environment.Since IPU is a major herbicide found in the aquatic ecosystem, duckweed (Lemna minor) was selected as the test organism for assessing the IPU's ecotoxicity in the aquatic environment.The present research was undertaken to determine the toxicity (EC50) of isoproturon on aquatic test plant duckweed (Lemna minor) as well as the influence of soil sorption of the herbicide on the same.

Setting IPU sorption experiment
The sorption of experiment was carried out using three different soils in a batch-slurry system with five sorbate concentrations (as mentioned above).The organic carbon and clay contents of the test soils are shown in Table 1. 10 g of air dried soil were weighed into a polypropylene centrifuge bottle and mixed with 20 ml standard IPU solutions.At least three replicates were run for the sorption experiment.A control was also carried out using the top concentration (i.e.0.2 mgl -1 ) without soil.The tubes were placed in end-over-end shaker for 16 hrs.

Sampling of the sorption study
After shaking the centrifuge tubes were removed from the shaker and allowed to stand for about one hrs and then the liquid supernatant were decanted into glass sample tubes.For the ease of chemical analysis by HPLC, the control and the three lower concentrations were extracted and concentrated in methanol by Solid Phase Extraction (SPE) method using SPE cartridges and transferred to glass sample vial.The concentrations of IPU in the extracted sample and the two unextracted higher samples were then determined by HPLC.Using the peak area data obtained from HPLC concentrations in the test samples were calculated.

Lemna Growth Inhibition Test
The Lemna growth inhibition experiment was conducted in two batches with five replications.The first set was prepared taking only the standard IPU solutions (20ml) in a 250 ml conical flask (i.e.without soil).The second set of ecotoxicity test was undertaken by taking 15 g of soil D in 60ml of each standard IPU solutions and then stirred to mix the soil and solution well.In both batches of conical flasks 2-3 Lemna colonies were placed and photographed before incubation in a plant growth cabinet for 2 week (20°C, 18 hrs light, 8 hrs dark).After 2 weeks, photographs for the batches without soil were taken.For the rest, frond numbers were counted to determine the growth of Lemna.

Growth rate estimation
Growth of Lemna was determined using both area of fronds and change in frond numbers.The areas of colonies were determined using the image analysis software Image J. It was done after converting the images at 8-Bits in a binary format.The area (in cm 2 ) covered by the fronds was measured by analyzing particles using a fixed length (rular).Then inhibition rate (I) was calculated using the following formula,

A 100
Where, Ac = ln (final area of the control) -ln (starting area of the control) At = ln (final area of the treated Lemna) -ln (starting area of the treated Lemna) I = % inhibition in growth The computation of the growth rate using the frond number was done by the following equation, ln ln Where, GR = Average specific growth rate Ni = No. of fronds determined at time i Nj = No. of fronds determined at time j ti = Moment time for start of the period tj = Moment time for end of the period The inhibition data was then used in the Probit programme to develop concentrationresponse relationship.The EC50 of IPU was determined both excluding and including soil data set.In the Lemna growth study, for most cases highest inhibition in growth was observed highest at the highest concentration and the lowest at control and 0.01 mg l -1 level (see Appendix).The EC50 value for Lemna using soil or without soil was found 0.171 and 0.121 mg l -1 (Table 3).It is noticeable that the value is greater in with soil treatment than the treatment lacks soil.However, statistical analysis revealed no significant difference between the two data sets.The reason behind may be the values dispersed very widely.The comparatively lower mean EC50 value in treatment lacking soil is presumable.Because it is assumed that some portion of the IPU will be adsorbed to the soil adsorption sites (organic matter and clays).Hence, the toxicity of the herbicide towards Lemna will be buffered to some extent.Because organic matter is a major sorbent of organic chemicals (Chiou, 1989 andLuthy et al., 1997).
A fair degree of irregularity was observed in the various data sets.And this strongly indicated the errors might have been occurred during the conduction of the study.A second attempt may be undertaken to assess the results of the present study.

Figure 1 .
Figure 1.Regression for (a) soil sorption data (log KOC values) on the Freundlich sorption coefficient (log Kf values) for Isoproturon in different soils; (b) soil organic carbon (OC) and Log Kf, (c) OC and Log KOC; and (d) soil clay content and log KOC.Ecotoxicity assessment of IPU on Lemna

Table 1 .
Organic carbon and clay content of test soils

Table 2 .
Mean Kf , nf and KOC values for different soils

Table 3 .
Mean and standard deviation for EC50 values in presence and absence of soil D