Synthetic Pyrethroids

Chapter 8

Solid-Phase Microextraction (SPME) Methods to Measure Bioavailable Concentrations in Sediment 1

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S. Bondarenko , J. Gan , and F. Spurlock Downloaded by UNIV LAVAL on June 15, 2014 | http://pubs.acs.org Publication Date: August 19, 2008 | doi: 10.1021/bk-2008-0991.ch008

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Department of Environmental Sciences, University of California, 2258 Geology Building, Riverside, C A 92521 California Department of Pesticide Regulation, 1001 I Street, Sacramento, C A 95812 2

The freely dissolved concentration (C ) in porewater can be used to improve prediction of sediment toxicity by pyrethroids. We used solid-phase microextraction (SPME) to analyze C of eight pyrethroids in sediment porewater. External calibration was applied to obtain C of chemicals, whereas internal calibration with C-c/s-permethrin was used to determine total concentration ( C ) . Total porewater concentration measured by using the isotopic-SPME method was well correlated with data obtained by exhaustive liquidliquid extraction (LLE). Method detection limits (MDLs) of the S P M E methods were lower than the 10 percentile of the reported LC50s for aquatic invertebrates, with relative standard deviation < 20%. The S P M E method was further used on field contaminated samples. Measuring C by SPME may represent a good alternative to the estimation of total or OC normalized sediment concentrations for predicting sediment toxicity from pyrethroid contamination. free

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© 2008 American Chemical Society In Synthetic Pyrethroids; Gan, J., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2008.

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Introduction Hydrophobic organic compounds (HOCs) such as organochlorine insecticides, polychlorinated biphenyls, polycyclic aromatic hydrocarbons, and pyrethroids, tend to accumulate in the bed sediment due to their high organic carbon partitioning coefficient (K ). Consequently potential toxicity to benthic organisms is an important concern. Fate and transport of HOCs in sediment depend on many factors, including sediment characteristics, sediment-chemical contact time and properties of HOCs. Exposure of sediment dwelling invertebrates to HOCs occurs primarily through transport of freely dissolved molecules of pollutants across cell membranes or direct ingestion of contaminated food particles. For soil invertebrates uptake of chemicals depends on physical characteristics of the organisms (soft and hard bodied) and physiology of the gut. In contrast, sediment invertebrate uptake mechanisms are not yet well understood (/, 2). Several studies have demonstrated that uptake through porewater is likely the dominant route for sediment exposure (3, 4). Equilibrium partitioning theory (EqP) is widely used to describe bioavailability of HOCs in sediment-water systems. EqP is based on the assumption of HOC partition equilibrium between porewater phase and sediment phase, leading to the conclusion that sediment toxicity can be estimated either from porewater or from organic carbon (OC)-normalized sediment concentration, regardless of the exposure route (3). OC-normalization of sediment concentrations has been often used for estimating sediment toxicity caused by HOCs, including that by pyrethroids (5-8). The preference is mainly due to the availability of relatively robust methods for measuring sediment concentration that are exclusively based on exhaustive extraction techniques. The bioavailability of HOCs under field conditions often differs from that predicted by using a constant K for different types of sediments along with EqP (2, 9). Numerous studies showed variable K values for the same compound due to sediment O C characteristics such as aromaticity, lipid content, black carbon content, and environmental factors, such as the contact time between the sediment and the contaminant, i.e., aging (10-13). Thus, the use of OC-normalization along with constant K is likely to yield inaccurate estimates of HOC sediment toxicity in many cases. The use of porewater concentration also has complications because the total porewater concentration C is the sum of both the freely dissolved concentration C d the concentration complexed with dissolved organic carbon (DOC). The presence of D O C in porewater complicates the measurement of C for HOCs with log K > 5 (14). Lately, a wide variety of extraction techniques have been tested for quantifying Cf . These methods include equilibrium dialysis, ultracentrifugation, reversed phase separation, size exclusion chromatography, fluorescence quenching, headspace, semipermeable membrane devices, and

Downloaded by UNIV LAVAL on June 15, 2014 | http://pubs.acs.org Publication Date: August 19, 2008 | doi: 10.1021/bk-2008-0991.ch008

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In Synthetic Pyrethroids; Gan, J., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2008.

151 solid-phase microextraction (SPME). Overview of these techniques and their limitations was given in detail elsewhere (14, 15). Solid-phase microextraction (SPME) was introduced by Pawliszyn and colleagues (16) and has been successfully used to measure C of HOCs in porewater (17-20). This technique is based on either "matrix-SPME" that uses the whole sediment for sampling or "negligible-depletion S P M E " that uses a small amount of porewater under negligibly depletive conditions. For instance, S P M E was used to quantify C of polyaromatic hydrocarbons (PAHs) in sediment porewater, and those SPME-determined concentrations were closely correlated with bioaccumulation of PAHs by benthic organisms (20). In recent studies, we applied S P M E in analyzing pyrethroids in runoff effluents and surface water samples (21-23), as reviewed in a separate chapter in this book by Hunter et al. Those studies showed that both uptake of pyrethroids by Daphnia magna and acute toxicity to Ceriodaphnia dubia demonstrated a higher correlation to the SPME-detected concentrations than to C obtained with L L E . In this chapter, we review the development of SPME methods for analysis of pyrethroids in sediment porewater. More details can be found in our published studies (24, 25). A significant amount of new information is also included, especially on the use of C-c/s-permethrin in G C - M S - M S analysis. The later application allows for the simultaneous determination of Cf and C in the same run. The proposed methods may be used for screening sediments for potential toxicity from pyrethroid contamination. free


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Materials and Methods Sediment and Porewater Preparation Several sediments were used in this study. The sediments were collected at the surface (0-5 cm) with a hand shovel into plastic containers and transported to the laboratory. Prior to use and characterization, sediments were wet sieved through a 2-mm screen to remove large particles. The OC content was determined by high temperature combustion of acidified sediments and subsequent analysis of the evolved C 0 . Fresh sediment porewater was prepared by centrifuging 200 g (wet weight) of the sieved sediment in a 250-mL polyethylene centrifuge bottle at 10,000 rpm for 30 min. The supernatant was carefully pipetted from multiple replicates into a 250-mL glass bottle for use in the following experiments. The D O C level in porewater samples was measured on an Apollo 9000 Carbon Analyzer (Teledyne Instruments, Mason, OH), using a high temperature combustion method. 2


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SPME Analysis The manual and automatic injector-type P D M S fibers were purchased from Supelco (Bellefonte, PA). Before use, new fibers were conditioned by heating at 320 °C in the G C inlet for 2 h, while reused fibers were cleaned and activated by heating at 260 °C for 3 min. For manual SPME, 10 mL of sample in 20-mL glass scintillation vials was used. The fiber immersion depth in the sample solution was fixed at 2 cm from the surface, and the solution was stirred at 600 rpm with a disposable magnetic bar made of 12 * 0.12 mm (diameter) rust-resistant steel wire. After exposure, the fiber was manually injected into the G C for analysis. For automatic S P M E analysis, 9 mL of sample in 10-mL amber glass vial with PTFE-coated septa was placed on a C T C Combi-PAL autosampler (Varian, Palo Alto, C A ) . Agitation of sample was performed by an automatic device at 250 rpm. To avoid carry-over between samples, the fiber was desorbed for an additional 5 min at 320 °C between consecutive sample runs. External calibration standards for S P M E analysis were prepared in deionized water and analyzed under the same conditions on the same day of analysis at six different concentrations (1000, 500, 100, 40, 10, 5, and 1 ng L" ) for G C - E C D and G C M S - M S analyses. For internal standard calibration using C-cw-permethrin on G C - M S - M S a concentration of 1 ng L" was used. 1

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Measuring SPME Water Partition Coefficients (#SPME)> Uptake (ki) and Elimination (Ar) Rates 2

Pesticide uptake was determined for both the 7- and 30-um polydimethylsiloxane (PDMS) fibers. Eight common pyrethroids were selected in this study: bifethrin, fenpropathrin, cw-and /raws-permethrin, lambdacyhalothrin, cyfluthrin, cypermethrin, esfenvalerate, and deltamethrin. Sediment porewater was spiked with pyrethroids at 40 ng L" , and then shaken for 10 min. The amount of acetone in each sample was < 0.01% (v/v). Exposure time was 10, 20, 30, 40, 60, 90, 120, 180, and/or 240 min. For each analysis a new vial was used and analysis was performed in three or four replicates. After exposure, the fiber was injected into the G C for analysis. The amount of pesticide desorbed from the fiber was calibrated by injecting pyrethroid standards in hexane under the same chromatographic conditions as for S P M E analysis. The volume of P D M S on the fiber was 0.028 uL and 0.132 uL for 7- and 30-um PDMS fiber, respectively. The data obtained from manual and automated S P M E analysis was fit to one-compartment model (26) using GraphPad Prism (v.4.03, GraphPad Software, San Diego, CA). From the curve fitting, uptake rate (£j), elimination (k ) rate, S P M E water partition coefficient (A^SPME), and minimum sampling time (/ ) were determined. 1

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153 Measuring Pyrethroid Concentrations in Porewater Spiked porewater samples were analyzed in parallel by S P M E with the 30um P D M S fiber (to measure C ) and L L E (to measure C ) to evaluate the capability of S P M E for detecting C . Sediment porewater samples were spiked with a mixture of all eight pyrethroids at 500, 100 or 40 ng L" . The content of acetone in the spiked samples was < 0.01% (v/v). A l l spiked samples were vigorously shaken for 5 min and equilibrated for 30 min or more before analysis. Preliminary experiments showed that pyrethroids added to a porewater sample reached an apparent equilibrium within 10 min after pesticide addition. Four replicates of 10-mL porewater samples were used for each concentration level. The S P M E sampling interval was fixed at 20 min, and the stirring speed was 600 rpm. Field contaminated samples were analyzed using S P M E to evaluate method performance. The samples were manually collected at two agricultural runoff sites in Orange County, C A . Wet sediment was weighed into a 250-ml polyethylene centrifuge bottle and the sample was centrifuged at 10,000 rpm to generate porewater. The porewater was analyzed using 30-um PDMS fiber under the same conditions as described above. To obtain C in porewater after S P M E sampling, the same porewater sample was extracted with ethyl acetate. The water sample was transferred to a glass separatory funnel and vigorously mixed with 10 mL of ethyl acetate for 1 min, followed by the collection of the ethyl acetate phase into a 50-mL pearshaped flask. The same extraction step was repeated for two additional times and solvent extracts were combined. The extract was dried by passing through 2 g of anhydrous sodium sulfate and then condensed to 0.5-1.0 mL on a vacuum rotary evaporator. A 1.0-uL aliquot was injected into the 6890 G C for analysis. Preliminary experiments showed that the recovery of the L L E procedure was 7598% for the selected pyrethroids. Calibration standards were prepared in hexaneacetone (1:1) and analyzed on the same day as the samples. free

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Measuring C

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Internal calibration with stable isotope labeled C-c/s-permethrin was used in S P M E and G C - M S - M S analysis to estimate C . In the same analysis, external calibration was performed using deionized water with known concentrations of pyrethroids (1-1000 ng L" ) to estimate C . The external calibration was found to be linear over the entire concentration range. The amount of acetone in each sample was < 0.01% (v/v). Fiber exposure time was 20 min, and other SPME sampling conditions were the same as given above. w

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154 G C - E C D and G C - M S - M S Analysis Manual S P M E and L L E analyses were carried out on an Agilent 6890 G C coupled with two electron capture detectors (ECD). After injection, the sample was split through a Y-connector into a DB-5MS column (30 m x 0.25 mm x 0.25 urn) and a DB-1701 column (30 m x 0.25 mm x 0.32 urn) (J&W Scientific, Folsom, C A ) . The dual columns were used to provide confirmation of the resolved peaks. The column temperature was held at 160 °C for 1 min, ramped to 300 °C at 10 °C min" , and then held at 300 °C for 6 min. The column flow rate was 1.5 mL min" (helium). The inlet temperature was 260 °C, and the detector temperature was 320 °C. The make-up gas flow rate was 60 mL min" (nitrogen). The injector port was used in pulsed splitless mode (50 psi at 3 min). External calibration was used for quantification. For compounds with multiple peaks, the sum of all peak areas was used for calibration and quantitation. Automated S P M E analyses were performed on a 3800 Varian G C system (Varian, Palo Alto, C A ) equipped with a DB-5MS column (30 m x 0.25 mm * 0.25 urn) (J&W Scientific, Folsom, C A ), a 1200 triple quadrupole mass spectrometer detector (Varian, Palo Alto, C A ) , and a Combi-PAL automated S P M E sampler (Varian, Palo Alto, C A ) . The column program was the same as for manual S P M E - G C - E C D analysis. The injector was used in the pulsed split mode (40 psi at 3 min). The injector, transfer line, and source temperature were 300 °C, 280 °C, and 170 °C, respectively. Electron impact mode at 70 eV and argon as collision gas were used. The following parent (precursor) ions were used for pyrethroids identification and quantitation: 181 (166) for bifenthrin, 181 (152) for fenpropathrin, lambda-cyhalothrin, and deltamethrin, 183 (153) for cisand /ra^-permethrin, 189 (174) for C-cw-permethrin, 163 (127) for cyfluthrin and cypermethrin, and 167(125) for esfenvalerate. 1


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Method Validation Method detection limits (MDLs) and method precision were determined using spiked sediment porewater samples. The M D L of each pyrethroid in a given matrix was determined by multiplying the one-sided 99% / statistic by the standard deviation obtained from four analyses of a matrix spike at 40 ng L" . The sampling and analytical conditions were the same as given above. The method precision was obtained by calculating the relative standard deviation (RSD) of pesticide concentrations from replicated analyses that were derived during the method development phase. 1


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Theory Concentration in the S P M E fiber assuming one compartment system with a first-order kinetics can be described as (26)

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where C (ng mL" ) is initial analyte concentration at 0 time, C M E , t ( g mL" ) is analyte concentration in the fiber coating at time / (min), k is uptake rate constant (min" ), and k is elimination rate constant (min' ). This model can be used when depletion is negligible or when the following condition is met 1

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where K P E (mL) is the volume of the fiber coating, and V (mL) is the volume of sample. At equilibrium, partition coefficient A " M E can be measured as ratio of k\ to k In order to measure C in the kinetic state, the freely dissolved amount that is extracted from sample into the fiber coating should be negligibly small and matrix presented in the aqueous phase should not interfere with S P M E measurement (26). The minimum S P M E sampling time (/ ) can be calculated as (26) S

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Results and Discussion Fiber Uptake Kinetics Uptake curves for the 7-|im and 30-um P D M S fibers are shown in Figure 1 for bifenthrin and Figure 2 for cypermethrin. Similar curves were also observed for the other pyrethroids. Since the depletion was less than 10 % (27) at all sampling time intervals, the first-order compartment model was used for



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Figure I. Uptake curves of bifenthrin into 7- and

PDMS-coated

SPME fibers

Figure 2. Uptake curves of cypermethrin into 7- and 30-jum PDMS-coated SPME fibers


157 describing the kinetics. The parameters derived from curve fitting are presented in Table 1 and Table 2. Uptake rate constants (k ) in matrix-free solutions ranged from 194 to 417 min" for the 7-um fiber, and from 74 to 274 min' for the 30um fiber, indicating slightly different diffusion kinetics of pyrethroids into fibers with different thicknesses of P D M S coating. Elimination rate constants (k ) were relatively consistent, ranging from 0.010 to 0.037 min" for all pyrethroids and different coating thicknesses. It has been observed that for highly hydrophobic compounds, one of the limiting steps in kinetic phase is diffusion of free chemicals through the stagnant water layer surrounding the fiber coating (28-31). Extensive mixing of the sample, which may be achieved using automated agitation or sonication, can significantly reduce the unstirring water layer and improve kinetic uptake (28, 32, 33). Similar diffusion and elimination rates were obtained by using automated S P M E coupled with an agitation device. These results suggest that under the conditions used here for both manual and automated S P M E analyses, the effect of the stagnant layer may be negligible. The ratio ^ S P M E ^ ^ was < 0.089 for the 7-um P D M S fiber and

Synthetic Pyrethroids

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