Comparison of supercritical fluid extraction with classical sonication

9) and soxhlet (n = 8) extractions. The recovery data and precision of each extraction method was evaluated statistically. It was found that overall a...
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Anal. Chem. 1882, 64, 1940-1946

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Comparison of Supercritical Fluid Extraction with Classical Sonication and Soxhlet Extractions for Selected Pesticides John L. Snyder; Robert L. Grob,' Mary Ellen McNally,i and Timothy S. Oostdykt Chemistry Department, Villanova University, Villanova, Pennsylvania 19085

The supercritical fluid extractlon of organophosphate and organochlorlne pesticides from soils was Investigated and compared to the ciardcal sonlcatlon and soxhlet extractions. Four roils, sand, clay, top soil, and river sedlment,were spiked with 12 organochlorhe and Organophosphatepestlcldes. These soik were extracted with supercritical COzmodified with 3 % MeOH at a pressure of 350 atm and a temperature of 50 OC. Overall average recoverleeof the 12 pesticldes were greater than 85 % for each of the soil matrices, and the overall average relatlve standard deviation (RSD) for ail the pesticides and soils was 5.1%. Secondly, a large batch of top soli was specially prepared and fortified with the same pesticides and repetively extracted wlng SFE ( n = 9) and the classical sonIcation( n = 9) and soxhlet ( n = 8) extractlons. The recovery data and precldon of each extraction method was evaluated statistically. It was found that overall average recoveries of the 12 pestlcide compounds for the sonlcatlon, soxhlet, and eupercrltlcal fiuld e x t r a c t hwere 94.7 %,93.1% ,and 9 1.6 %, rerpectlvely. SFE demonstrated the best precision of the three extraction methods with the overall average relative standard deviation belng 2.94%. Lastly, natlve soils contaminated with organochlorine pestlcldes were repetitively extracted using SFE and the eonlcation extraction. Comparabk precklonr betweenSFE and the sonication method were demonstrated. Also SFE performed equally as well as the sonlcation extraction in recoverlng a number of organochlorine pesticides from the native soil samples.

INTRODUCTION Currently the United State Environmental Protection Agency (USEPA) espouses two primary methods for the extraction of semivolatileorganic contaminants and pesticides from soils.1tz These classical methods, the sonication extraction and the soxhlet extraction, are widely used in environmental laboratories. Unfortunately, these methods are time consuming and involve the use of large volumes of solvents that are often toxic or carcinogenic. Not only is the purchase of these solvents expensive, but their safe disposal is equallycostly. Also, during most soil extractions, the solvent extracts must be concentrated. In this process, the excess solvent is usually evaporated in a hood and vented to the atmosphere. These evaporated solvents help contribute to our present air pollution problems. The use of supercritical fluids in analytical chemistry has become increasingly apparent in the last several years.3 In the environmental area, supercritical fluid extraction (SFE) + Current address: Lancaster Laboratories, Inc., 2425 New Holland Pike, Lancaster, PA 17601. 1 Current address: E. I. du Pont de Nemours & Co., Inc., Experimental Station, Building 402, Wilmington, DE 19898 (1)Test Methods for Evaluating Solid Waste, 3rd ed.; EPA SW-846, Method 3540; US.Government Printing Office: Washington, DC, 1990. (2) Test Methods for Evaluating Solid Waste, 3rd ed.; EPA SW-846, Method 3550; U S . Government Printing Office: Washington, DC, 1990. (3) Hawthorne, S. B. Anal. Chem. 1990,62, 633A.

has been used by numerous investigators to extract organic contaminants from soils and other environmental samples. Often SFE has been compared to the soxhlet or sonication extraction techniques. Most of the time, SFE has been reported as being superior in extraction efficiency and faster. For example, Schantz and Cheder4 used SFE to extract polychlorinated biphenyls (PCB's) from sediment and polyaromatic hydrocarbons (PAHs) from urban dust. They compared the results to classical soxhlet extractions and found SFE better or equivalent to the soxhlet in extraction efficiency. Hawthorne and Miller5 reported good recoveries of PAH's using nitrous oxide modified with methanol. They also concluded that SFE gave better recoveries, was faster, and used less solvent than sonication or soxhlet extractions. McNally and Wheeler6 optimized SFE conditions necessary to extract Diuron and Linuron from soil. In their work, they reported that SFE is less labor intensive and time consuming than the classical soxhlet extraction used for these compounds. SFE has also been used to extract 2,3,7,8-tetrachlorodibenzop-dioxin from sediment7 and fly ash.8 In both cases, SFE was found to be faster than the classical method and to give better recoveries when nitrous oxide modified with methanol and pure nitrous oxide were used as the extraction fluids. In another relevant paper, Richards and Campbell9 reported on the optimization of an SFE method using COZmodified with 2 % methanol to extract 16 environmentally significant semivolatile organic compounds. In their work, they found that SFE was more efficient than soxhlet or sonication extractions for removing 13 of the 16 compounds from spiked soil and that SFE was faster and more convenient than the other two conventional methods. Because of these benefits in speed and cost and the small amounts of solvents which are necessary, SFE appears to be a very viable alternative to classical soxhlet and sonication extractions. In this study the use of SFE to extract organochlorine and organophosphate pesticides from soils was investigated. In the first part of the investigation SFE was used to extract a variety of soils which were spiked with 12 organochlorine and organophosphate pesticides. These soils included sand, top soil, clay, and river sediment. The structures of the organochlorine pesticides are shown in Figure 1 and the organophosphate structures are shown in Figure 2. Secondly, SFE is compared to the sonication and soxhlet extractions for extracting a top soil fortified with these pesticides. A special spiking method was used to ensure the freshly spiked pesticides were evenly dispersed through the soil and had maximum contact with the soil surface. Care was also taken to ensure that no residual solvent from the spiking solution remained in the soil which might influence (4) Schantz, M. M.; Cheder, S. N. J. Chromatogr. 1986, 363, 397.

(5) Hawthorne, S. B.; Miller, D. J. Anal. Chem. 1987, 59, 1705. (6) McNally, M. E.; Wheeler, J. R. J. Chromatogr. 1988, 447, 53.

(7) Onuska, F. I.; Terry, K. A. J . High Resolut. Chromatogr. 1989,12, 357. (8) Alexandrou, N.; Pawliszyn, J. Anal. Chem. 1989, 61, 2770. (9) Campbell, R. M.; Richards, M. LCGC 1991,9, 358.

0003-2700/92/0364-1940$03.00/0 0 1992 American Chemical Soclefy

ANALYTICAL CHEMISTRY, VOL. 64, NO. 17, SEPTEMBER 1, 1992

a)

c1

c+ l&cl CI e)

CI CI

CI

f)

Figure 1. Structuresof organochlorinepesticides: (a)tetrachlorometaxylene (TCMX), (b) endrin, (c)endrin aldehyde, (d) p,p'-DDT, (e) mirex, (f) decachloroblphenyl(DCB). 0 I1 Cl,C=CH-O-P(OCN3),

CI' f)

Flgure 2. Structures of organophosphate pesticides: (a) dichlorvos, (b) diazlnon, (c) ronnel, (d) parathion (ethyl),(e) methldathlon, (f) tet-

rachlorvinphos.

the extraction efficiencies of the different methods. The soil was extracted repetitively by the classical soxhlet, sonication, and supercritical fluid methods. The accuracy and precision of the three methods have been statistically evaluated and compared. A portion of this fortified soil was allowed to age for a period of 8 months and again extracted using SFE and sonication. Lastly, native soils (real soil samples) contaminated in the environment with organochlorine pesticides were extracted repetitively using SFE and the classical sonication method. The pesticide recoveries from both extraction methods have been compared and their precisions evaluated. EXPERIMENTAL SECTION Equipment. All SFE extractions were performed on a SFE50 instrument available from Suprex Corporation, Pittsburgh,

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PA. The sonication extractions were performed using a Model 385 Ultrasonic disruptor, Heat Systems-Ultrasonics, Inc., Farmingdale, NY. A Varian 3400 gas chromatograph equipped with an electron capture detector (ECD) and a Restec RTx-5, 30-m X 0.53-mm, df 3.0-pm capillary column were used for separation and quantification of the pesticides. A Hewlett-Packard 589011 gas chromatograph and a HP5971 mass selective detector were also used in this study. A Restec RTx-5, 30-m X 0.25-mm, df 1.0-pm capillary column was directly interfaced to the GC/MS. Standards and Reagents. The 12 pesticide compounds used for spiking were obtained from the Environmental Protection Agency Repository, Research Triangle Park, NC, and Chemical Services Inc., West Chester, PA. Stock solutions of each compound at 1 mg/mL were prepared in methanol. Working calibration standards were prepared in methyl tert-butyl ether (MTBE) by serial dilution of an intermediate standard prepared from the 12 individual stock standards. The standards of other organochlorine pesticides, which are EPA CLP pesticides,1° found in the native soils were prepared by dilutingconcentrated certified stock solutions purchased from Restek Corp., Bellefonte, PA. Only pesticide-grade methylene chloride (MeC12),acetone (Ac), hexane (nCe),methanol (MeOH), and MTBE were used in this study. SFC-grade carbon dioxide containing 3 % methanol (Scott Specialty Gases, Plumsteadville, PA) was used for all supercritical fluid extractions. Soils. The sand was a washed and dried Reagent sand purchased from Mallinckrodt, Inc., Paris, KY. The clay, top soil, and river sediment were collected locally (Lancaster County, PA). Each was dried at 105 "C and passed through a No. 10 wire mesh. Approximately 2.0 g of these soils were used in the initial recovery study. Twenty-five microliters of the spiking solution was added to the soil in the vessel, and the solvent was allowed to evaporate prior to extraction. Native Soils. These soils were initially analyzed by our Laboratory (Lancaster Laboratories, Inc.) and found to contain levels of organochlorine pesticides using the EPA Contract Laboratory Pesticide method.lO The presence of these pesticides was confirmed, as stated in the EPA method, by dual column confirmation or by GC/MS. These soils were extracted multiple times using the sonication and supercritical fluid extraction methods described above. Two- to three-gram portions of each of these soilswas used for the SFE extraction,and 5-10-g portions were used in the sonication extraction. These soils were spiked with surrogate standards, TCMX and DCB, to monitor the extraction. Preparation of Fortified Top Soil (FTS).Soil Pretreatment Procedure (FTS). Approximately 500 g of the sieved soil was extracted with 1:l (v/v) methylene chloride/acetone for 12 h using a soxhlet apparatus. The soil was then oven-dried again to remove the excess solvent. A small amount of deionized water (2.5%by weight)was added to the soil. Then the soilwas tumbled so that it was thoroughly mixed. At this point, 100 g of the soil was removed to serve as a blank for the soxhlet, sonication, and SFE extractions. Soil Spiking Procedure (FTS). A 400-g portion of the soil was placed in a 1-L glass container and spiked with a MeClz solution containing the 12 organochlorineand organophosphate pesticides. The container of soil was then placed in a water bath at 30 OC, and the MeClz was slowly evaporated over a period of about 24 h in a hood. During the evaporation,the soil and MeClz mixture were intermittently stirred. After the methylene chloride was completely evaporated, the soil was tumbled to ensure homogeneity. Aging. A portion of this fortified top soil was kept at 4 OC in a refrigerator for a period of 8 months. Soxhlet Extraction. Ten-gram portions of the soil were extracted according to the EPA method.' Briefly the procedure was as follows: the soils were mixed with 10 g of sodium sulfate, and then extracted with 1:l hexane/acetone for 20 h in a soxhlet apparatus. The extracts were passed through a Na2S04 drying column. The extracts were then evaporated using a Kuderna Danish (KD) apparatus and a N-evap concentrator to a volume (10) Statement of Work for Laboratory Program, 1990.

Organic Analysis, USEPA Contract

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ANALYTICAL CHEMISTRY, VOL. 64, NO. 17, SEPTEMBER 1, 1992

-

SAND SAND RSD

CLAY

-0- CLAY

-t

RSD

RIVER SED RIV SED RSD

-

TOP SOIL

TS RSD

0

ii

Flguro 3. Recovery and precision of organochlorine and organophosphate pesticldes from sand, clay, river sediment, and top soil. Spike added (ng/g): (1)dichlorvos (5201,(2)TCMX (30),(3)dlazlnon (515),(4)ronnel (25),(5)parathbn (78),(6)methidathion (loo),(7)tetrachlowinphos (36), (8)endrin (45), (9)endrin aldehyde (43), (10)p,p'-DDT (38), (11)mirex (76),(12)DCB (30).

of 1-2 mL. The solvent was exchanged to MTBE and the final volume adjusted to 5 mL. A blank soil and a total of nine spiked soil replicates were extracted. Sonic Probe Extraction. To be consistent with the soxhlet method and the SFE methods, 10-gportionsof soilwere extracted accordingto the EPA sonication method.2 Briefly the procedure was as follows: each soil was extracted ultrasonically for 3 min with three 40-mL portions of 1:l methylene chloride/acetone. The ultrasonic disruptor was equipped and operated as recommended by the method. Following each extraction, the solvent extract was filtered through Whatman 41 paper. The final extracts were dried in a Na2S04 column and concentrated using a KD apparatus and N-evap concentrator to approximately 1-2 mL. After exchanging the solvent to MTBE, the final extracts were reconstituted to a volume of 5 mL. A blank and seven replicates of the FTS were extracted. SFE Extraction Conditions. The soils were loaded into 2or 10-mL stainless steel extraction vessels (Keystone Scientific, Bellafonte, PA) depending on the sample size and extracted at 350 atm pressure and 50 "C, using a Suprex SFE-50. Carbon dioxide premixed with 3% methanol was used as the extraction fluid. Samples were extracted statistically at 350 atm for 10 min. During this time, there was no net flow of COZthrough the system. After 10 min, the system was extracted dynamically at 350 atm. The 2-mL vessels were extracted dynamically for 20 min and the 10-mLvessels were extracted for 40 min. A 50-fim capillary restrictor was used to control the flow of supercritical CO2 through the system and help maintain pressure across the extraction cell. Although there was some variability in the flow from extraction to extraction, a flow of about 1mL/min of COz in the supercritical state was obtained. The pesticide analytes were collected by bubbling the vented COz through 5 mL of MTBE. No concentration of the extracts was required, and each MTBE extract was diluted to an exact final volume of 5 mL. Separation and Quantification. The pesticides were separated and quantified by either gas chromatographywith electron capture detection (GC-ECD) or by gas chromatography/mass spectrometry (GC/MS). GC-ECD. One microliter of extract was injected directly into a gas chromatograph equipped with a DB5 megabore column. The injection port was maintained at 250 "C and the ECD was held at 300 "C. The column was temperature programmed from 140 "C, after a l-min hold, to 290 "C at 4 "C/min. The system was held at the final temperature for 25 min. Helium was the carrier gas at a flow of 5 mL/min and nitrogen was used as a make-up gas to the detector at 25 mL/min. Quantification was

performed using a five-point linear calibration curve plotting concentration and peak area. GC/MS. One microliter of the extracts were injected into a Hewlett-Packard (HP) 5890 gas chromatograph interfaced directly to a HP 5971 mass selective detector. The gas chromatograph was equipped with a DB5 narrow bore capillary column. The injection port was held at 250 "C, and a splitless injection (0.75min) was performed. The mass spectrometer was tuned so that it was most sensitive to the mid-massrange and was operated in the single ion monitoring (SIM) mode. Two or three ions prevalent in the electron impact (EI) spectrum of each pesticide were monitored around the expected retention time for each pesticide. Quantification was performed using a single-point calibration standard which was made at approximately the same concentration of the pesticides expected in the solid extracts. Safety. Because of the high pressures used in SFE, special precautions should be taken. All vessels, tubing, and fittings should be rated to withstand the pressure used for each extraction. The extraction system should be also fitted with a pressurerelief valve to prevent the accidental build up of pressure beyond the safe operating level of the system. Care must also be taken in handling the pesticide compounds to avoid human contact because of their toxicity and because many are suspected carcinogens.

RESULTS AND DISCUSSION Figure 3 shows the recovery by SFE of the organochlorine and organophosphate pesticides spiked onto the various types of sand, clay, top soil, and river sediment. The overall average recovery for all 4 soil types and the 12 pesticides was 94%. The overall precision for all pesticides and soils was 5.1 % .A blank and quadruplicateextractionswere performed for each of the soil types. However, only three extractions were performed for the top soil after it had been spiked. The overall average recovery of the organochlorine pesticides and the organophosphate pesticides were nearly equal, 93.9 % and 93.3 % , respectively. Considering only the organochlorine pesticides, the overall precision was 3.4 % The overall precision of just the organophosphate pesticides was 6.8%. Good overall recoveries of the pesticides were obtained in each of the soil matrices. If the soil matrices were ranked in terms of highest overall recovery of pesticides, the following order would be observed: top soil (10.5%), clay (98%),sand

.

ANALYTICAL CHEMISTRY, VOL. 64, NO. 17, SEPTEMBER 1, 1992

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Table I. Comparison of Extraction Methods: Organochlorine Pesticides Soxhlet vs Sonic Probe vs Supercritical Fluid Extractions amount found std % added, meth- trials, (av), dev, RSD, recvd s n ng/g % (av) compound ng/g od"

Table 11. Comparison of Extraction Methods: Organophosphate Pesticides Soxhlet vs Sonic Probe vs Supercritical Fluid Extractions amount found std ?6 added, meth- trials, (av), dev, RSD, recvd compound ng/g oda n ng/g s % (av)

TCMX

dichlorvos

30

SX

SP SFE

endrin

45

sx SP

SFE

endrin aldehyde

43

p,p'-DDT

38

sx SP

SFE

SX

SP SFE

mirex

76

SX

SP SFE

DCB

30

SX

SP SFE

8 7 9 8 7 9 7 7 9 8 7 9 8 7 9 8 7 9

23 24 22 44 49 44 35 31 36 33 31 38 71 80 74 29 31 29

2.1 0.79 1.2 2.6 0.52 1.5 1.3 1.0 1.2 4.3 0.57 0.52 8.1 1.2 2.2 1.1 0.57 0.50

8.9 3.2 5.4 6.0 1.1 3.5 3.7 3.1 3.3 13.0 1.8 1.4 11.4 1.5 3.0 3.7 1.8 1.7

78 81 74 97 108 97 81 71 84 87 81 99 94 105 97 98 104 96

a SX = soxhlet extraction, SP = sonic probe extraction, SFE = supercritical fluid extraction.

(877% ), and river sediment (85% ). This order is probably not terribly significant consideringthe closenessof the recoveries, the spiking technique, and slight variations in gas chromatographic calibrations. All of the soils had a pH between 6 and 7. The top soil, the clay, and the river sediment had organic contents of 1.7% , 1.3 76 ,and 1.2 76,respectively. Particle size analysis for the top soil showed the soil to be 53.2% sand, 37.6% silt, and 9.2% clay. Similarly particle size analysis on the clay showed it to be 27.2% sand, 37.6% silt, and 35.2% clay. The river sediment was very sandy with a composition of 93.2% sand, 3.6 76 silt, and 3.2 % clay. The larger particle size and hence smaller surface may give a clue to the lower pesticide recoveries found with the river sediment and sand. Because the spiked soils were allowed to dry after adding the spiking solution, evaporation losses of the pesticides from such a matrix were more likely. In general, recovery of the most volatile of the spiked pesticides, Dichlorvos,was the lowest and its precision was the poorest. The result of the comparison experiments on the fortified top soil (FTS) are summarized in Table I for the organochlorine pesticides which were investigated. The average recoveries of each of the organochlorine pesticides were statistically evaluated at the 955'% confidence level to determine if there was a statistical difference in means.ll When the mean recoveries from the soxhlet extraction and SFE were compared,no significant differencewas noted for TCMX, endrin, mirex, and DCB. SFE, however, showed higher recovery for p,p'-DDT and endrin aldehyde. When the sonication method mean recoveries were evaluated against the SFE mean recoveries, SFE showed improved recovery for two of the six compounds investigated, endrin aldehyde and p,p'-DDT. SFE had lower mean recoveries for TCMX, endrin, mirex, and DCB. The overall average recoveries for the six organochlorine pesticides for the soxhlet, sonication, and supercritical fluid extractions were nearly identical at 89.0%, 91.6%, and 91.2%, respectively. The precisions of the methods were also evaluated statistically for each pesticide using the F test at the 95 5%confidence level." The precision of the SFE method was not significantly different from soxhlet method for TCMX, endrin, and en(11)Anderson, R. L. Practical Statistics for Analytical Chemistry; Van Nostrand Reinhold: New York, 1987.

520

SX

SP SFE

diazinon

515

SX

SP SFE

ronnel

25

parathion (ethyl)

78

methida-

100

SX SP SFE

SX SP SFE

SX

SP

thion

SFE

tetrachlorvinphos

36

SX SP SFE

8 6 5 8 6 9 8 6 9 8 6 9 8 6 9 8 6 8

333 375 318 479 493 433 26 27 25 22 77 73 108 100 106 42 41 39

28 42 13 24 27 13 1.4 2.4 1.2 20 1.3 0.88 5.7 4.8 3.4 4.7 1.3 2.5

8.4 11 4.0 5.1 5.4 3.0 5.2 9.1 5.0 92 1.6 1.2 5.3 4.8 3.3 11 3.2 6.3

64 72 61 93 96 84 104 106 98 28 99 94 108

100 106 117 113 109

SX = soxhlet extraction, SP = sonic probe extraction, SFE = supercritical fluid extraction. (1

drin aldehyde. The SFE method showed significant improvement in precision forp,p'-DDT, mirex, and DCB. When comparing SFE to the sonication extraction, no statistical difference was noted in the standard deviations for five of the organochlorine pesticides. However, the relative standard deviation (RSD) of the sonication method was found to be slightly better then the SFE RSD for endrin (1.06 vs 3.49%). The overall average of the RSD's for the six organochlorine pesticides using the soxhlet,sonication, and supercritical fluid extractions were 7.77 % , 3.05 5% , and 2.0974, respectively. The six organophosphate pesticides were evaluated in a similar fashion. These results are shown in Table 11. When statistically evaluating the mean recoveries of the organophosphate pesticides, it was found that the average recoveries for dichlorvos,methidathion, and tetrachlorvinphos were not significantly different between the soxhlet extraction and SFE. The soxhlet extraction produced higher mean recoveries for dichlorvos and methidathion, although the average recovery of methidathion (106%) was actually greater than 100%. The soxhlet recovery (28%) for parathion was found to be much lower than either the sonication or SFE recovery. The heat required for the soxhlet extraction may have contributed to this problem and caused a breakdown reaction of the parathion to occur during the extraction, although no documentation of this could be found in the literature. Compared to SFE, the sonication extraction generated better averagerecoveries (statisticallyrelevant at the 953'% confidence level) for dichlorvos, diazinon, ronnel, and parathion. No significant statistical distinction was found between SFE and the sonication recovery for tetrachlorvinphos. SFE showed a higher mean recovery (106%) for methidathion than did the sonication method (1007% ). The overall averagerecoveries of the organophosphate pesticides by the soxhlet, sonication, and supercritical fluid extractions were very close at 97.2 5% , 97.875, and 92.876, respectively. Because of the problem associated with the soxhlet extraction of Parathion explained above, the average recovery of parathion is excluded from the soxhlet overall average. When comparing the soxhlet extraction to SFE, SFE demonstrated much improved precision for parathion, and the other five organophosphate pesticides showed equivalent precisions. When the precisions of the sonication extraction and SFE were compared, a statistical difference in the

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ANALYTICAL CHEMISTRY. VOL. 64. NO. 17. SEPTEMBER 1, 1992

120,

I

N

Table 111. Recovery of Organochlorine and Organophosphate Pesticides from Aged Top Soil' supercritical fluid sonication (n = 6 ) recovery

dichlorvos TCMX diazinon ronnel

parathion methidathion tetrachlorvinphos endrin endrin aldehyde p,p'-DDT mirex 1

2

3

L

5

6 7 8 COWlhn

9 1 O l l 1 2

Flgure 4. Comparison of mean recoveries of wganochlorineand Mganophosphate pesticides from spiked top soil uslng soxhlet, 80111cation. and supercrliical fluid exiractlons.

precisions was found for only dichlorvos and ronnel. In both cases, the SFE precision was found to he better. Dsicarding thevery poor precision for thesoxhlet extractionof parathion, the average RSDs for the soxhlet, sonication, and supercritical fluid extractions were 7.07%, 5.89%, and 3.80%, respectively. The intent of these experiments was to evaluate and compare the extraction efficiencies for the three separate methods. An attempt was made to control and minimize any variables such as the spiking techique, the time from spiking toextraction, and the gas chromatographic conditions, which might influence the recovery data other than the actual extraction procedure. The soil was spiked in a manner to ensure that each pesticide was dispersed evenly throughout the maximum contact between the soil and each pesticide was achieved. The Me& used in spiking wm evaporated to eliminate any influence it may have had on recovery. In routine proceduresfor spiking,theanalyte is seldomdispersed evenly through the soil and the spiking solvent is generally not removed. The soxhlet and sonication extractions were performed simultaneously. This was not possible for SFE, however, because of instrument limitations. The supercriti d fluid extractions were performed serially, and each required about an hour to complete. Figure 4 is a bar graph summarizing the recovery data for the extraction methods and further illustrates the effect pesticide volatility may have on recovery data in this experiment. The pesticides in Figure 4, going from left to right, are in order of their chromatographic elution and, in general, follow increasing molecular weight and decreasing volatility. The first four pesticides shown in the bar graph, dichlorvos, TCMX, diazinon, and ronnel gradually increase in overall recovery for the three extraction methods. The lower recoveries of the lighter pesticides is a limitation of the spikingprocedure. When the MeC1,solution was evaporated from the soil, the lower molecular weight and more volatile pesticides partially evaporated with the solvent, producing lower recoveries. Although all the extracts were injected under identical chromatographic conditions, the presence of artifact peaks and extracted organic material affected separation and recovery. The chromatograms for the soxhlet method contained the most extraneous peaks and the highest background noise level. Some of these extraneous peaks coeluted with analyte peaksandpreventedquantification by ECD. Because of this problem, dichlorvos, diazinon, ronnel, methidathion, and tetrachlorvinphos were quantified using GCiMS in the SIM mode.

DCB

(av), %

RSD, %

5.1 67 55 63 64 68 50

52 11 17 15 16 17 24 10 9.6 11 12 12

84 63 81 81 82

(n = 2) recovery

(av), ?% 3.6 51 55 54 57 59 33 68 28 53 67 74

Spiking levels are identical to those found in Tables I1 and 111. The spiked soil was aged 8 months prior to extraction. The presenceof humic materials in the extractsalsocaused a general increase in chromatographic response, especially for the polar organophosphate pesticides. As much as possible, this effect was corrected for by the injection of intermittentstandards. Itwasmost exaggerated in thesoxhlet and sonication extractions and is likely responsible for some of the high recoveries (>loo% ). The effects of aging the top soil were also investigated. Approximately 100g of the spiked soil, which was allowed to stand in a glass container and stored in a refrigerator for nearly 8 months, was repetitively extracted hy the supercritical fluid and sonication methods. These results are summarized in Table 111. It can be noted that the overall average recovery by SFE dropped from 91.6% to 63.5% and the overall average precision was 17.3% compared to 2.94% initially. The less stable organophosphate pesticides show the greatest fluctuation in hoth categories. Also the volatile dichlorvos was not recovered after the aging study. The overall average recovery from the sonication extraction changed from 94.7 % initially to 50.1% after aging. The sonication data, however, must be qualified in that there was only enough of the aged spiked top soil to perform two extractions, and the KD extracts were inadvertently allowed to remain on the steamhath during the concentration step for an extended period. Three native soilssuhmittedtoourLaboratoryforpesticide analysis and found to contain significant levels of organ* chlorinepesticides wererepetively extracted by SFE and sonication. These soils are designated native soil 1, soil 2,and soil 3. Soil 1 was found to contain levels of p,p'-DDT and the related breakdown products p,p'-DDD and p,p'-DDE. This soil was a dark top soil and contained significant amounts of fme gravel. The pH of the soil was 8.2, and it had a moisture content of 6.6%. A comparison of pesticide recoveries of repetitive supercritical fluid (n = 4) and sonication (n = 3) are presented in Table IV. The amount of DDT extracted by SFE (450 mg/g) and by sonication (43mg/g) agree very closely. Better precision was realized hy SFE (RSD 5.0%) than bysonication (RSD 17%),althoughthereisnostatistical significancebetweenthe two precisionsat the95% confidence level. Native soil 2 was found to contain high levels of endrin and endrin ketone. This soil was a brown sandy loam. The soil was found to have a pH of 7.41 and a moisture content of 22.1%. Quantification of the supercritical fluid sonication extractions are found in Table V. There is no significant difference between the sonication and SFE data. Most of the differences in the SFE and sonication precisions for the

ANALYTICAL CHEMISTRY, VOL. 64, NO. 17, SEPTEMBER 1, 1992

Table IV. Quantification of Organochlorine Pesticides in Native Soil Sample 1: SFE vs Sonicationn supercritical fluid sonication (n = 3)

(n = 4)

TCMX p,p’-DDE p,p’-DDD

p,p’-DDT DCB

amount (av)

RSD, %

amount (av)

RSD, %

95% 44 nglg 43 ng/g 453 ng/g 97 %

4.1 7.1 8.6

93 % 36 nglg 39 nglg 431 nglg 102%

12 15 8.4 17 2.6

5.0 2.1

a TCMX and DCB added as surrogatesprior to extractions;results expressed as percent recovered.

Table V. Quantification of Organochlorine Pesticides in Native Soil Sample 2: SFE vs Sonicationn supercritical fluid sonication (n = 6) (n = 3) amount (av) RSD, % amount (av) RSD, % 76 % 14 11 TCMX 68% 17 heptachlor 40 14 ng/g 16 ng/g 9.7 6.3 aldrin 81 ng/g 114 ng/g dieldrin 9.3 2.4 327 nglg 344 ngig a-chlordane 8.4 4.7 429 ng/g 483 nglg 9.0 6.5 y-chlordane 80 nglg 88 nglg endrin 1524 ng/g 2.5 1418 ngig 4.6 endosulfan I1 3.0 6.2 54 ngig 58 ng/g endrin aldehyde 84 3.3 29 ngig 45 ngig 2601 ng/g 14 endrin ketone 2675 ng/g 9.3 DCB 90 % 5.9 6.9 130% TCMX and DCB added as surrogatesprior to extractions; results expressed as percent recovered. Table VI. Quantification of Organochlorine Pesticides in Native Soil Sample 3 SFE vs Sonicationn supercritical fluid sonication (n = 7)

(n = 3)

amount (av) RSD, % amount (av) RSD, % TCMX aldrin a-chlordane endrin endrin aldehyde endrin ketone DCB

80% 32ng/g

23ng/g 273ng/g 17 ngig

60nglg 937%

1.3 19 14 24 36 12 15

96 76 33 nglg

20nglg

312 ng/g 31ng/g

88ngIg 110%

4.8 4.7 17 7.7 14 14 8.3

0 TCMX and DCB added as surrogatesprior to extractions;results expressed as percent recovered.

pesticides are not relevant. The very poor precision shown by SFE for dieldrin and endrin aldehyde are related to the smaller sample size used for SFE and by the fact that these compounds were barely detectable at the dilution necessary to quantify the endrin and endrin ketone. Native soil 3 was similar to soil 2 in the pesticides which were detected. However, this soil was a sand with little humic material. The pH of this soil was 7.6 and the soil was 21% moisture. The results of the SFE and sonication extractions are given in Table VI. The increase in efficiency was noted with the sonication method over SFE as was the precision. However, there is no significant statistical difference in the numbers. In practical terms, the SFE method was faster and easier to perform than either the sonication or soxhlet extraction. Less solvent was consumed by SFE. Approximately 400-500 mL of solvent was consumed per soxhlet extraction and about 150-200 mL per sonication extraction. SFE required only 5-10 mL of solvent. Because of the small amount of solvent necessary, SFE required no solvent concentration step using

1941

the Kuderna Danish apparatus. During this process the chance for analyte loss because of evaporation, breakdown, or reaction of the compound is greatly increased. Endrin, endrin aldehyde, and p,p’-DDT are known to form breakdown products when exposed to heat. It is interesting to note that recoveries of endrin aldehyde, andp,p’-DDT in the FTS study were higher for the SFE method where no solvent evaporation was necessary. The sonication and soxhlet extractions were more labor intensive than SFE to set up and perform. This labor included the soxhlet extraction set ups, the assembling and cleaning of glassware, weighing the samples, and the KD solvent concentration step. The SFE method was less labor intensive and automated by the Suprex SFE-BO. Only sample loading, adjustment of the final extract volume, and cleaning of the extraction vessel were necessary. The soxhlet extraction required 20 h to complete plus the time that was needed to prepare and set up the extractions and concentrate the solvent. The SFE method took about 1 h per extraction. The sonication method required only about 15-30 min per extraction and was relatively easy to set up but required the solvent concentration step which took about 1 h per extract to complete. Use of the ultrasonic horn required the operator’s full-time attention and was labor intensive. Because of the availability of equipment, the soxhlet and sonication extractions could be performed simultaneously in parallel (nine extractions set up at one time). The supercritical fluid extractions, because of instrument limitations, were performed serially. New instrumentation has been developed, however, to allow parallel simultaneous extractions to be performed. This instrumentation, however, may not be rugged enough to compete with soxhlet or sonication extractions on a commercial scale.

CONCLUSION SFE has been shown to be a successfulanalytical technique in extracting organochlorine and organophosphate from a variety of spiked and native soils. Overall average recoveries of the 12 pesticides spiked onto sand, clay, top soil, and river sediment were 87 5% , 9 8 % , 105 % , and 85 % The precision, as measured by an overall average RSD of 5 % ,was very good. Recoveries of organochlorine pesticides from three native soils using the sonication extraction and SFE were nearly identical, and the precision of the methods were not significantly different. Repetitive extraction of a large batch of fortified top soil by supercritical fluid, sonication, and soxhlet methods demonstrated good extraction efficiencies as measured by the overall average of the mean recoveries of the 12 pesticide compounds. Sonication was highest at 94.7% and was followed by soxhlet at 93.1% and SFE at 91.6%. The low frequency of parathion obtained by the soxhlet extractions was deleted from this average. The sonication extraction, however, gave statistically better recoveries from a majority of the individual pesticides when compared to SFE. When comparing the soxhlet to SFE, the majority of the individual pesticides had no significant difference in recovery at the 95 % confidence level. In this study SFE was found to have the best overall precision as indicated by an overall average RSD for the 12 pesticides of 2.94%. Sonication ranked second with an average RSD of 4.47 5%. However, for many of the individual pesticides, there was no significance in the difference in the precisions at the 95 % confidence level between sonication and SFE. The soxhlet extraction was found in these experiments to be the least precise, and the average RSD for the 11 of the pesticides, excluding parathion which was poorly recovered, was 7.42%.

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ANALYTICAL CHEMISTRY, VOL. 64, NO. 17, SEPTEMBER 1, 1992

SFE ranked highest of the three methods in practical terms. It was faster than either the soxhlet or sonication extractions and was less labor intensive. For a single extraction, the soxhlet extraction required about 22 h and the sonication extraction took about 2 h to complete. SFE on a single sample could be completed in less than 1 h. Less solvent was required for SFE than the other two methods, and no concentration of the solvent extracts was required. However, because of instrument limitations, simultaneous parallel extractions could not be performed using SFE. In this regard, the soxhlet and sonication extractions have a definite advantage. The equipment to perform these classical extractions is readily

available and widely used commercially. SFE has not yet reached this stage. RECEIVED for review January 6, 1992. Revised manuscript received May 6, 1992. Accepted May 8,1992. Registry No. TCMX, 877-09-8; p,p’-DDT, 50-29-3; DCB, 2051-24-3; dichlorvos, 62-73-7; diazinon, 333-41-5; ronnel, 29984-3; parathion, 56-38-2; methidathion, 950-37-8;tetrachlorvin, 1221-74-5; endrin, 72-20-8; endrin aldehyde, 7421-93-4; mirex, 2385-85-5.