Published in J. Environ. Qual. 33:406-408 (2004).
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A Simple High Performance Liquid Chromatography Method for Analyzing Paraquat in Soil Solution Samples
Ying Ouyang*,a,
Robert S. Mansellb and
Peter Nkedi-Kizzab
a Department of Water Resources, St. Johns River Water Management District, P.O. Box 1429, Palatka, FL 32178-1429
b Soil and Water Science Department, University of Florida, Gainesville, FL 32611-0290
* Corresponding author (youyang{at}sjrwmd.com).
Received for publication May 1, 2003.
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ABSTRACT
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A high performance liquid chromatography (HPLC) method with UV detection was developed to analyze paraquat (1,1'-dimethyl-4,4'-dipyridinium dichloride) herbicide content in soil solution samples. The analytical method was compared with the liquid scintillation counting (LSC) method using 14C-paraquat. Agreement obtained between the two methods was reasonable. However, the detection limit for paraquat analysis was 0.5 mg L–1 by the HPLC method and 0.05 mg L–1 by the LSC method. The LSC method was, therefore, 10 times more precise than the HPLC method for solution concentrations less than 1 mg L–1. In spite of the high detection limit, the 12C (nonradioactive) HPLC method provides an inexpensive and environmentally safe means for determining paraquat concentration in soil solution compared with the 14C-LSC method.
Abbreviations: HPLC, high performance liquid chromatography • LSC, liquid scintillation counting
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INTRODUCTION
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CONTAMINATION of soil and ground water resources by herbicides such as paraquat is an increasing environmental concern. Paraquat is an effective herbicide for both terrestrial and aquatic plants. It is very soluble in water, methanol, and polar solvents but is insoluble in nonpolar organic solvents. Paraquat has been used extensively as a pre- and post-emergence herbicide for weed control, sod destruction before reseeding in lieu of mechanical cultivation, and desiccation before harvest. The dichloride salt of paraquat is a quaternary dipyridylium herbicide. The toxicant portion of the molecule is an organic cation with an electrical charge of +2 and a molecular weight of 186.2 g mol–1 (Weed Science Society of America, 1967). Two characteristics of paraquat are its deactivation by adsorption on soil and its stability in soil (Tucker et al., 1967). Paraquat has a very long persistence in soils, which is only limited by microbial breakdown of the strongly sorbed chemical (Weed Science Society of America, 1967). It has been reported that paraquat is toxic to human organs such as lungs, heart, liver, kidneys, cornea, adrenal glands, skin, and digestive system (USEPA, 1987).
Public health concern about paraquat contamination requires methods to determine paraquat concentrations that are accurate, safe, and not demanding of time and sophisticated equipment. Several methods have been reported to determine paraquat in various matrices, including a colorimetric method (Gilreath and Duranceau, 1986), a spectrophotometric method (Willard et al., 1965; Weber et al., 1969), and a radioassay or 14C method (Bray, 1960). Although these methods have provided useful means for analysis of paraquat in soil solution and water samples, they commonly have one or two limitations such as low accuracy, poor precision, high time consumption, or limited safety. Recently, USEPA Method 549.2 (Munch and Bashe, 1997) was developed from Method 549.1 (Hodgeson et al., 1992) to determine diaquat and paraquat in drinking water by liquid–solid extraction and HPLC with ultraviolet detection. Although this method can be employed to measure paraquat in drinking water, its applicability to analyze paraquat in soils is untested. The objective of this study was to develop a method for measuring paraquat in soil solution using HPLC (hereafter referred to as the 12C-HPLC method). The 12C-HPLC method was evaluated by comparing the results with those obtained using the 14C-LSC method.
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Materials and Methods
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Soil types used in this study included Eustis fine sand (siliceous, thermic Psammentic Paleudults) and Webster silt loam (fine-loamy, mixed, superactive, mesic Typic Endoaquolls). Both soils were obtained from a repository in the Soil and Water Science Department, University of Florida. These soils were air-dried and passed through a 2-mm sieve. Selected soil properties are shown in Table 1. Analytical-grade paraquat was purchased from ChemService (West Chester, PA). Radioactive 14C-paraquat (labeled in methyl group) with a specific activity of 374 KBq mmol–1 and radiopurity of >98% was purchased from Sigma Radio Chemical (St. Louis, MO). Acetonitrile, NaCl, KCl, and HPLC water were of analytical grade.
Paraquat standards were prepared according to procedures described by Chichila and Walters (1991). Nonradioactive 12C-paraquat stock solution at 100 mg L–1 and working standards with 0.1, 1, 5, and 10 mg L–1 were prepared in volumetric flasks by diluting with HPLC water. The 14C-paraquat stock solution was prepared by dissolving 2.8 mg 14C-paraquat into 49.1 mL deionized water (57 mg L–1). Calculated radioactivity (4.97 x 106 dpm mL–1, where dpm is disintegrations per minute) for the stock solution agreed closely with experimentally measured (4.99 x 106 cpm mL–1, where cpm is counts per minute). Working standards of 5, 50, and 100 mg L–1 paraquat containing 14C material used for the Eustis soil experiments were prepared by pipetting 1 mL of 14C-paraquat stock solution into 500-mL volumetric flasks. Working standards of 300 and 500 mg L–1 paraquat with 14C material used for the Webster soil experiments were prepared by pipetting 2 mL of 14C-paraquat stock solution into a 250-mL volumetric flask. The working standards of 50 and 100 mg L–1 for the Webster soil experiments were prepared by pipetting 10 and 20 mL of 500 mg L–1 paraquat into a 100-mL volumetric flask. All standards were then immediately transferred into plastic bottles to avoid sorption of paraquat onto glass surfaces.
The HPLC system used for analyzing paraquat consisted of a Spectra-Physics (Mountain View, CA) SP-8800 ternary HPLC pump, an Alltech Associates (Deerfield, IL) silica analytical column (25 cm in length with a 4.6-mm-i.d.), a Waters (Milford, MA) 490 programmable multiwavelength ultraviolet detector, an SP 8780 autosampler, and an SP 4290 integrator. The HPLC mobile phase was prepared by dissolving 5.0 g NaCl in 600 mL HPLC water that was previously adjusted to pH 3.0 with HCl, and then mixed with 400 mL of acetonitrile. A flow rate of 1.0 mL min–1 with an operating pump pressure of approximately 4.14 MPa and a chart speed of 40 cm h–1 were used. The UV detector wavelength was set at 257 nm with 0.15 AUFS (absorbance units full scale). A 20-µL volume of each working standard was injected into the column by the autosampler. One or two injections of mobile phase or a blank sample were made to elute accumulated co-extractives in the injector and the column before injecting the standards. At the end of the analyses, the column was flushed with HPLC water.
Duplicate 20-mL volumes of paraquat (containing 12C or 14C materials with concentrations ranging from 50 to 100 mg L–1 for the Eustis soil experiments or 100 to 500 mg L–1 for the Webster soil experiments) and 10 g air-dried soil were added to 50-mL plastic centrifuge tubes and shaken on a platform shaker for 0.5, 1, 6, 12, and 24 h. The soil samples were then centrifuged at 15000 rpm for 15 min. For 12C-paraquat analysis, an aliquot of 20 µL supernatant was injected into the HPLC column. In addition, spiked samples were used to estimate the paraquat recovery rate in the soil samples. For 14C-paraquat analysis, 1 mL of the supernatant and 10 mL of ScintiVerse scintillation solution (Fisher Scientific, Hampton, NH) were pipetted into plastic vials and analyzed for paraquat contents using LSC. A radiation background check on LSC was conducted by pipetting 1 mL of deionized water and 10 mL of ScintiVerse into a plastic vial and counting by LSC. Possible sorption of paraquat onto the surfaces of plastic tubes was estimated by pipetting 20 mL of 100 mg L–1 (with duplication) into plastic centrifuge tubes and incubating for 24 h without soil.
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Results and Discussion
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Soil paraquat concentrations were determined successfully using the 12C-HPLC method. The retention time for paraquat for the experimental conditions used in this study was 4.35 min (Fig. 1) . Table 2 revealed that no paraquat was detected in the soils by the 12C-HPLC method when initial paraquat concentrations were 50 mg L–1 for the Eustis soil and 100 mg L–1 for the Webster soil due to strong sorption of paraquat on soils. However, as the initial paraquat concentrations increased, soil solution paraquat contents were detected by the 12C-HPLC method (Table 2). Note that the 14C-LSC method was able to detect paraquat in soil solution even for the lowest initial paraquat concentration (50 and 100 mg L–1, Table 2). For each initial paraquat concentration added to the soil, the soil solution concentration decreased as the contact time increased due to sorption kinetics.
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Fig. 1. Chromatogram of paraquat concentration of 10 mg L–1 measured by the 12C-high performance liquid chromatography (HPLC) method. The peak height is at 4.35 min.
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Table 2. Comparison of paraquat concentrations in soil solution for Eustis and Webster soils measured by the 12C-high performance liquid chromatography (HPLC) and 14C-liquid scintillation counting (LSC) methods.
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Statistical comparison of measured paraquat concentrations in soil solution between the 12C-HPLC and 14C-LSC methods is shown in Fig. 2
. With a linear regression equation YHPLC = 0.8958XLSC and R2 = 0.9333, we concluded that a reasonable agreement was obtained between the two methods. Further comparison of paraquat concentrations measured by the two methods revealed that the 12C-HPLC method had a much higher detection limit than that of the 14C-LSC method. However, both methods showed similar experimental results for the higher soil solution paraquat concentrations (Table 2). The detection limit for paraquat analysis was 0.5 mg L–1 by the 12C-HPLC method and 0.05 mg L–1 by the 14C-LSC method. For solution concentrations less than 1 mg L–1 the 14C-LSC method was, therefore, 10 times more sensitive than the 12C-HPLC method. In spite of the high detection limit, the 12C-HPLC method was determined to be an inexpensive and environmentally safe means for determining paraquat concentrations of >1 mg L–1 in soil solution as compared with the 14C-LSC method.
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Fig. 2. Comparison of paraquat concentration in soil solution measured by the 14C-liquid scintillation counting (LSC) method and the 12C-high performance liquid chromatography (HPLC) method.
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In this investigation, elaborate experimental procedures and conditions were used to demonstrate that paraquat content in soil solution can be measured using HPLC. Although a detection limit of 0.5 mg L–1 is not impressive, we hope this report will stimulate further refinement of the 12C-HPLC method for soil solution analysis of paraquat content in the future. This detection limit could be possibly lowered by choosing a more appropriate HPLC column and adjusting the mobile phase pH value by trial and error.
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NOTES
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Florida Agric. Exp. Stn. Journal Ser. no. R-09604.
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REFERENCES
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- Bray, G.A. 1960. A simple efficient liquid scintillator for counting aqueous solutions in a liquid scintillation counter. Anal. Biochem. 1:279–285.[ISI]
- Chichila, T.M., and S.M. Walters. 1991. Liquid chromatographic determination of paraquat and diquat in crops using a silica column with aqueous ionic mobile phase. J. Assoc. Off. Anal. Chem. 74:961–976.[Medline]
- Gilreath, J.P., and S.J. Duranceau. 1986. Photodegradation of paraquat applied to polyethylene mulch film. HortScience 21:1145–1146.
- Hodgeson, J.W., W.J. Bashe, and J.W. Eichelberger. 1992. Method 549.1: Determination of diquat and paraquat in drinking water by liquid-solid extraction and high performance liquid chromatography with ultraviolet detection. EPA/600/R-92/129. USEPA Environ. Monitoring Systems Lab., Cincinnati, OH.
- Munch, J.W., and W.J. Bashe. 1997. Method 549.2. Determination of diquat and paraquat in drinking water by liquid-solid extraction and high performance liquid chromatography with ultraviolet detection. Revision 1.0. USEPA Natl. Exposure Res. Lab., Office of Res. and Development, Cincinnati, OH.
- Tucker, B.V., D.E. Pack, and J.N. Ospenson. 1967. Adsorption of bipyridylium herbicides in soil. J. Agric. Food Chem. 15:1005–1008.
- USEPA. 1987. Health advisory draft report: Paraquat. USEPA Office of Drinking Water, Washington, DC.
- Weber, J.B., R.C. Meek, and S.B. Weed. 1969. The effect of cation-exchange capacity on the retention of diquat+2 and paraquat+2 by three-layer type clay minerals: II. Plant availability of paraquat. Soil Sci. Soc. Am. J. 33:382–385.[Abstract/Free Full Text]
- Weed Science Society of America. 1967. Herbicide handbook of the Weed Science Society of America. Academic Press, New York.
- Willard, H.H., L.L. Merritt, Jr., and J.A. Dean. 1965. Instrumental methods of analysis. D. Van Nostrand Co., New York.
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ABSTRACT
INTRODUCTION
