Anal. Chem. 2000, 72, 1294-1300
High Extraction Efficiency for POPs in Real Contaminated Soil Samples Using Accelerated Solvent Extraction Andreas Hubert,† Klaus-Dieter Wenzel,*,† Michael Manz,† Ludwig Weissflog,† Werner Engewald,‡ and Gerrit Schu 1u 1 rmann†
Department of Chemical Ecotoxicology, UFZ Center for Environmental Research, Permoserstrasse 15, D-04318 Leipzig, Germany, and Faculty of Chemistry and Mineralogy, Institute for Analytical Chemistry, University of Leipzig, Linne´ strasse 3, D-04103 Leipzig, Germany
Systematic investigations were performed to study the dependence of the extraction efficiency of persistent organic pollutants (POPs), including chlorobenzenes, HCH isomers, DDX, PCB congeners, and PAHs, on the accelerated solvent extraction (ASE) operating variables solvent and temperature. Mixed soil samples from two locations with considerable differences in soil properties and contamination in the Leipzig-Halle region (Germany) were used. The objective was to optimize ASE for the extraction of POPs from real soil samples and to improve on the results achieved with Soxhlet extraction (SOX). Solvents with differing polarities were tested. Quadruple and triple determinations were performed on the two soils, respectively, between 20 and 180 °C in 20 °C steps. All the results were compared with those obtained by SOX, as well as, in some cases during preliminary studies, by ultrasonic extraction (USE). In ASE, the optimum conditions proved to be two extraction steps at 80 and 140 °C (average RSD 10.7%) with three static cycles (extraction time 35 min) using toluene as solvent and at a pressure of 15 MPa. Owing to the superior analyte/ matrix separation by ASE, in many cases for real soil samples analytical values better by up to 1 order of magnitude or even more were obtained compared to SOX results. The advantage of an extraction procedure over other methods can particularly be recognized from its extraction efficiency for diverse, complex, real environmental samples reflecting long-term trends and changes in pollutant behavior in soil matrixes. Owing to continuous multiple pollutant entries with a broad spectrum of other unknown pollutants, decomposition products and a larger number of undefined pollutant-matrix conjugates may be present in addition to the substances to be analyzed. These can exert varying influences on the quality of extraction, cleanup, and analysis. The extraction efficiency of individual extraction methods for persistent organic pollutants (POPs) from soil material in † ‡
UFZ Center for Environmental Research. University of Leipzig.
1294 Analytical Chemistry, Vol. 72, No. 6, March 15, 2000
particular also depends on physicochemical properties1 of the pollutants and on matrix factors such as the organic carbon content and other soil parameters (e.g., clay and secondary oxides and hydroxides of inorganic elements), which can affect sorption and desorption, degradation, and the formation of conjugates with the matrix. POPs persistence and lipophilic affinity lead to the accumulation and enrichment of these substances in soils with high organic contents.2,3 Various correlations can be used to estimate the adsorption coefficient Koc determining these qualities.4,5 After optimization, the extraction method accelerated solvent extraction (ASE), which has been used since about 1995, represents an exceptionally effective extraction technique6 compared to alternatives such as Soxhlet extraction (SOX),7 steam distillation,8,9 microwave extraction (MWE),10,11 ultrasonic extraction (USE),12,13 and partly also supercritical fluid extraction (SFE).14-16 (1) Schu ¨u ¨ rmann, G. Ecotoxic modes of action of chemical substances. In: Ecotoxicology; Schu ¨u ¨ rmann, G., Markert, B., Eds.; John Wiley and Spektrum Akademischer Verlag: New York, 1998; Chapter 22. (2) Cousins, I. T.; Hartlieb, N.; Teichmann, C.; Jones, K. C. Environ. Pollut. 1997, 97, 229-238. (3) Marschner, B. Z. Pflanzenernaehr. Bodenkd. 1999, 162, 1-14. (4) Sabljic, A.; Gu ¨ sten, H.; Verhaar, H.; Hermens, J. Chemosphere 1995, 31, 4489-4514. (5) Lyman, W. J. Adsorption Coefficient for Soils and Sediments. In: Handbook of Chemical Property Estimation Methods; Lyman, W. J., Reehl, W. F., Rosenblatt, O. H., Eds.; American Chemical Society: Washington, DC, 1990; Chapter 4. (6) Wenzel, K.-D.; Hubert, A.; Manz, M.; Weissflog, L.; Engewald, W.; Schu ¨u ¨ rmann, G. Anal. Chem. 1998, 70, 4827-4835. (7) Tremolada, P.; Burnett, V.; Calamari, D.; Jones, K. C. Environ. Sci. Technol. 1996, 30, 3570-3577. (8) Veith, G. D.; Kiwus, L. M. Bull. Environ. Contam. Toxicol. 1977, 17, 631636. (9) Ramos, L.; Tabera, J.; Hernandez, L. M.; Gonzalez, M. J. Anal. Chim. Acta 1998, 376, 313-323. (10) Lopez-Avila, V.; Young, R.; Benedicto, J.; Ho, P.; Kim, R.; Beckert, W. F. Anal. Chem. 1995, 67, 2096-2102. (11) Kodba, Z. C.; Marsel, J. Chromatographia 1999, 49, 21-27. (12) Burford, M. D.; Hawthorne, St. B.; Miller, D. Anal. Chem. 1993, 65, 14971505. (13) Wenzel, K.-D.; Mothes, B.; Weissflog, L.; Schu ¨u ¨ rmann, G. Fresenius Environ. Bull. 1994, 3, 734-739. (14) Wenzlawiak, B.; Hoffmann, A.; Bodusz, M. Fresenius’ J. Anal. Chem. 1992, 343, 116-117. (15) Langenfeld, J. J.; Hawthorne, St. B.; Miller, D. J.; Pawliszyn, J. Anal. Chem. 1993, 65, 338-344. (16) Li, K.; Landriault, M.; Fingas, M.; Llompart, M. Analusis 1998, 26, 365369. 10.1021/ac991005l CCC: $19.00
© 2000 American Chemical Society Published on Web 02/11/2000
Table 1. Extraction Efficiencies for Pollutant Groups Depending on the Extraction Method and Solventa ASEb
USE pollutant group
SOX toluene
hexane
hexane/ CH2Cl2, 1:1
hexane
hexane/ CH2Cl2, 1:1
hexane/ methanol, 1:1
toluene
∑HCHs ∑DDX ∑Cl-Bz ∑PCBs ∑PAHs
8.07 7.43 0.35 0.54 66.8
0.21 1.99 nd 0.12 13.7
2.69 7.84 0.36 0.54 82.5
3.78 7.44 0.41 0.25 112
1.58 3.72 0.13 0.11 101
6.33 10.6 0.39 0.20 202
17.2 8.03 0.55 0.23 261
∑POPs
83.2
16.0
93.9
124
107
220
287
a Site Ro ¨sa. Concentrations are given as ng/g of dry weight (nd ) not detectable). Concentrations are average values of double (SOX, USE) or triple (ASE) determinations. b Extraction temperatures are 40 and 120 °C.
Table 2. Comparison of the Extraction Efficiencies for ASE and SOX in Soils from Southern Russiaa Caucasus pollutant group
ASEc
∑HCHs ∑DDX ∑Cl-Bz ∑PCBs ∑PAHs
4.24 5.90 0.56 0.69 219
Elbb SOX 0.83 0.65 0.11 nd 26.1
f
ASEc
Ki-Pb SOX
5.1 9.1 5.1
0.83 0.70 0.40 0.87 6.87
1.23 0.23 0.16 nd 10.3
8.4
f
ASEc
Ki-Mb SOX
0.7 3.0 2.5
1.43 0.41 0.05 0.34 22.6
1.01 0.16 0.11 nd 5.52
0.7
f
ASEc
Klub SOX
1.4 2.6 0.5
9.76 3.30 0.18 77.5 149
0.97 0.17 0.06 nd 6.32
4.1
Lower Volga
b
Krab
pollutant group
ASFc
SOX
∑HCHs ∑DDX ∑Cl-Bz ∑PCBs ∑PAHs
11.6 13.2 0.09 0.69 42.6
3.37 0.78 0.13 0.03 8.10
f
SOX
3.4 17.0 0.7 23.0 5.3
2.65 9.36 0.14 0.32 22.8
0.82 3.32 0.13 nd 9.38
range of f
10.1 19.4 3.0
0.7-10.1 2.6-19.4 0.5-5.1
23.6
0.7-23.6
Kalmykian Steppe
Cerb ASEc
f
Astb
Elib
f
ASEc
SOX
3.2 2.8 1.1
22.5 157 0.25 0.92 183
4.08 19.7 0.17 0.19 66.9
2.4
f
ASEc
SOX
5.5 9.9 1.5 4.8 2.7
1.23 1.47 nd 0.08 5.68
0.52 0.08 0.40 nd 2.86
f
ASEc
Godb SOX
2.4 18.4
5.46 1.35 nd 0.71 9.73
0.72 0.60 0.11 0.20 4.05
2.1
f
range of f
7.6 2.3
2.4-7.6 2.3-18.4 0.7-1.5 3.6-23.0 2.1-5.3
3.6 2.4
a Concentrations are given as ng/g of dry weight (nd ) not detectable). Solvent is toluene. f ) concentration for ASE/concentration for SOX. For locations, see the section “Sampling Locations” in the text. c Extraction temperatures are 40 and 120 °C.
The advantages are its considerably shorter extraction times and the lower consumption of solvent than with SOX, the universal use of solvents and solvent mixtures differing in polarity, and individually variable pressures of 35-200 atm at possible extraction temperatures ranging from room temperature to 200 °C. The recovery data, including precision and systematic deviations of certified and spiked soils or sediments using ASE, are comparable to those of other extraction methods.17-21 Use of the solvent n-hexane/acetone, 1:1, in ASE allowed the simultaneous determination of alkanes, PAHs, and chlorinated compounds. In comparison to general criteria such as the solvent consumption, extraction time, and practical conditions (variation possibilities) of all the various methods, SFE and ASE are the superior techniques, but they also require the most expensive equipment.19 These remarks are based on the soil extraction conditions of 100 °C and n-hexane/acetone, 1:1, as solvent, referred to in Dionex (17) Ho ¨fler, F.; Ezzell, J.; Richter, B. LaborPraxis 1995, 3, 62-67. (18) Ho ¨fler, F.; Ezzell, J.; Richter, B. LaborPraxis 1995, 4, 58-62. (19) Heemken, O. P.; Theobald, N.; Wenzlawiak, B. W. Anal. Chem. 1997, 69, 2171-2180. (20) Saim, N.; Dean, J. R.; Abdullah, M. P.; Zakaria, Z. Anal. Chem. 1998, 70, 420-424. (21) Popp, P.; Keil, P.; Mo ¨der, M.; Paschke, A.; Thuss, U. J. Chromatogr., A 1997, 774, 203-211.
application notes.22 According to our latest findings concerning the extraction of POPs from plant materials such as pine needles and mosses, ASE can, after optimization, enable successful analyses from much more contaminated locations with remarkable increases in extraction efficiency,6 which is of enormous interest for ecotoxicological and toxicological assessment. The main aim of this work was to use ASE’s better extraction efficiency for natural plant samples compared to that of traditional procedures6 by optimizing the extraction parameters solvent and temperature for the simultaneous extraction of POPs from different real soil sampling materials, also. EXPERIMENTAL SECTION Sampling. General Procedures. Topsoil samples (0-5 cm) were taken in October 1997 in Germany and in July 1997 at nine locations between the Caspian Sea and the Black Sea in southern Russia. The soil was kept cool during transport to the laboratory, where it was air-dried for 24 h at room temperature, and it was then frozen at -80 °C until further preparation. Sampling Locations. The soils used for the investigations came from Hettstedt and Ro¨sa in the Leipzig-Halle region of Germany. (22) Application Notes ASE 313, 316, 318, and 320; Dionex: Sunnyvale, CA, 1994.
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1295
Table 3. Extraction Efficiencies for the Hettstedt Samples Depending on Temperaturea temp (°C) compd R-HCH β-HCH γ-HCH δ-HCH -HCH
20 1.39 1.31 7.09 0.27 0.13
40
60
80
100
120
140
160
2.89 2.05 12.8 0.53 0.33
4.60 8.37 20.4 0.65 0.45
4.52 11.8 21.1 0.55 0.33
5.03 13.1 14.9 0.93 1.39
4.96 16.4 14.1 0.64 0.54
6.13 13.7 18.6 1.16 0.73
3.48 7.10 11.6 0.58 0.75
∑HCHs
10.2
18.6
34.5
38.3
35.4
36.6
40.3
23.5
DDT DDE DDD
40.6 5.44 0.81
59.4 7.58 1.11
87.3 10.8 1.65
120 14.2 2.38
71.8 10.3 1.41
68.0 11.6 2.19
64.2 11.1 2.96
15.4 8.48 3.16
∑DDX
46.9
68.1
137
83.5
81.8
78.3
27.0
100
PCB 28 PCB 52 PCB 101 PCB 138 PCB 153 PCB 180 PCB 194
0.16 0.66 1.27 2.22 1.51 1.36 0.40
0.19 0.78 1.56 2.45 1.71 1.41 0.42
0.21 0.88 1.93 2.76 1.91 1.51 0.49
∑PCBs
7.58
8.52
9.69
PHE ANT FLUOR PYR BaP
36.6 3.26 76.8 61.2 36.5
84.8 7.28 123 87.7 37.1
212 18.9 227 146 45.3
337 32.0 330 205 56.9
417 42.0 304 181 45.7
340
649
961
990
∑PAHs
214
1.12 1.05 3.82 4.11 2.96 2.29 0.64 16.0
1245tetraCB 1234tetraCB pentaCB hexaCB
nd nd 0.11 1.04
0.08 0.08 0.41 1.89
0.26 0.26 1.04 3.76
0.10 0.12 1.17 5.20
∑Cl-Bz
1.15
3.19
5.32
6.59
180 1.72 4.30 4.00 0.65 0.29 11.0 8.36 6.49 3.06 17.9
av RSD (%)b 17.7 12.1 10.6 39.9 36.7 13.5c 17.4 10.9 16.8 15.0
0.87 0.49 1.83 1.92 1.25 0.91 0.44
0.86 0.43 2.09 1.59 1.15 0.76 0.43
0.95 0.43 0.99 1.96 1.50 0.83 0.40
0.87 0.37 0.88 1.95 1.21 0.91 0.35
0.64 0.24 0.78 1.64 1.39 nd nd
16.7 17.9 22.3 15.9 16.0 18.2 16.6
7.71
7.31
7.06
6.54
4.69
17.7
1.86 1.45 2.77 5.65 11.7
460 49.5 353 222 34.2 1120 2.23 1.75 3.05 5.64 12.7
477 55.0 388 226 36.8 1180 2.31 1.86 3.34 6.06 13.6
413 35.4 291 117 25.2
370 32.8 337 129 49.5
10.3 12.9 11.6 12.8 16.8
882
918
12.9
1.51 1.34 2.92 4.49 10.3
1.33 1.12 2.07 3.23
23.4 25.5 15.5 14.2
7.75
19.7
a Average values are based upon four determinations. Concentrations are given as ng/g of dry weight (nd ) not detectable). Solvent is toluene. SOX values by way of comparison (ng/g of dry weight): ∑HCHs, 21.8; ∑DDX, 6.80; ∑PCBs, 3.56; ∑PAHs, 1053; ∑Cl-Bz, 5.65 b Average RSD for each substance resulting from nine individual RSD values of the various temperature steps. c RSD without δ-HCH and -HCH.
Hettstedt is 70 km west of Leipzig, near local heavy metal plants used to smelt shredded copper cables until the mid-1990s. The smelting/combustion processes liberated organochlorines such as hexachlorobenzene, DDT, and HCH (lindane),23 and soil in the area contains highly enriched levels of organic compounds. Ro¨sa is located 30 km north of Leipzig in a pine forest area with predominantly sandy soil. The nine measuring sites in southern Russia between the Caspian Sea and the Black Sea were geographically divided into three sampling areas, as follows. 1. The Caucasus: Klu (Kluchorpass) and Elb (Elbrus), both ∼2000 m above sea level (high mountains); Ki-P (Kislovodsk Plateau), ∼800 m above sea level, near the city Kislovosk; Ki-M (Kislovodsk Mountain), ∼1200-1400 m above sea level (both foothills). 2. Lower Volga: Kra (Krasnyj Jar), ∼50 km east of Astrakhan; Cer (Cernyj Jar), ∼300 km north of Astrakhan; Ast (Astrakhan, Volga delta), ∼50 km from the Caspian Sea. 3. Kalmykian Steppe: God (Godschur), ∼300 km north of Elista; Eli (Elista), ∼300-400 km north of the Caspian Sea. Standards, Materials, and Solvents. The extracts were quantified using p,p′-DDT, p,p′-DDE, γ-HCH, the internal standard (23) Weissflog, L.; Wenzel, K.-D.; Manz, M.; Kleint, F.; Schu¨u ¨ rmann, G. Environ. Pollut. 1999, 105, 341-347.
1296 Analytical Chemistry, Vol. 72, No. 6, March 15, 2000
mixture containing a 13C-labeled chlorobenzene cocktail (EM-1725-A), PCB 28 (EC-1413), PCB 153 (EC-1406), and the deuterated PAH surrogate cocktail (ES-2044). These standards were supplied by Promochem (Wesel, Germany). All solvents (nhexane, dichloromethane, toluene, acetone, methanol, diethyl ether) used were of analytical grade (Merck GmbH, Darmstadt, Germany). Forty gram samples of Florisil, 60-100 mesh, (Promochem) were conditioned with twice the volumes of the solvents methanol and dichloromethane. After drying at 100-120 °C in a drying cupboard, the samples were activated under nitrogen for 4 h at 180 °C. Soxhlet. General Conditions. Ten gram samples of air-dried soils were extracted with 150 mL portions of toluene in a DET 5 Soxhlet unit (Behr Labor-Technik GmbH, Du¨sseldorf, Germany) for 24 h. USE. General Conditions. Extractions of soil samples (bath sonication; cooling water was circulated through the bath; bath temperature 15-20 °C) were carried out using a Sonorex Super RK 255H (HF 35 kHz, power 320 W; Brandelin Electronics, Berlin). Ten gram samples of air-dried soils (∼24 h until a constant final weight had been reached) were extracted into beakers 3 × 10 min). Each extraction step was performed using 100 mL of solvent (cf. Table 1). The extracts were then filtered, and the filtrates were combined.
Figure 1. ASE efficiencies of POPs from Ro¨sa soil depending on extraction temperature (average values based upon three determinations). Solvent is toluene. SOX values for comparison (ng/g of dry weight): ∑HCHs, 14.2; ∑DDX, 10.5; ∑PCBs, 0.75; ∑PAHs, 122; ∑Cl-Bz, 0.66.
ASE. General Conditions. Extractions of 10 g samples of airdried soils were carried out using 33 mL stainless steel vessels of a Dionex ASE 200 (Dionex GmbH, Idstein, Germany). The
stagnant volume of each vessel was filled up with a mixture of ∼10 g of Florisil/Al2O3, 2:1 (as precleanup). Each extraction step consisted of a heating phase of 5 min (8 min at 180 °C) and three Analytical Chemistry, Vol. 72, No. 6, March 15, 2000
1297
Table 4. Ratio Factors f for the Extraction Efficiencies of ASE vs SOXa pollutant group tempb
HCHs
20 40 60 80 100 120 140 160 180
0.47 0.85 1.58 1.76 1.62 1.68 1.85 1.08 0.50
20 40 60 80 100 120 140 160 180
0.27 0.63 0.47 0.65 1.10 0.72 2.41 1.61 1.63
DDX
PCBs
(a) Hettstedt 6.90 10.0 14.7 20.2 12.3 12.0 11.5 3.97 2.63
Samplesc 2.13 2.39 2.72 4.49 2.17 2.05 1.98 1.84 1.32
(b) Ro ¨sa Samplesd 0.64 0.73 1.04 0.84 0.92 0.91 0.98 0.81 1.62 1.21 1.46 1.47 3.54 3.53 3.16 2.87 2.98 2.28
PAHs
Cl-Bz
0.80 0.68 0.62 0.91 0.94 1.06 1.12 0.84 0.87
0.20 0.57 0.94 1.17 2.07 2.25 2.41 1.82 1.37
0.62 0.93 1.28 1.85 3.03 3.25 9.10 7.39 6.47
0.94 2.42 1.53 2.24 4.96 5.71 13.1 9.77 8.62
a f ) concentration of pollutant group for ASE/concentration of pollutant group for SOX. b Temperature in °C. c For individual values, see Table 3. d For individual values, see Figure 1.
10 min static cycles. Extractions were performed with ∼60 mL of solvent, and the vessels were rinsed with ∼2 mL of the same solvent, the time for rinsing (nitrogen) being 120 s. Conditions for Preliminary Investigations. Extractions to test solvents using soil samples from Ro¨sa (cf. Table 1) and the Russian samples (all nine locations) were performed in two steps at 40 and 120 °C.11 The solvent for the Russian samples was toluene. Basic Temperature Investigations. Quadruple determinations (Hettstedt) and triple determinations (Ro¨sa) of mixed soil samples were performed using the solvent toluene. The extractions were carried out in the range from room temperature (20 °C) up to a maximum of 180 °C in 20 °C steps. Cleanup. After the extraction procedures (ASE, Soxhlet, USE), internal standard solutions were added. Each extract was concentrated to 2 mL and transferred to a column (diameter 0.5 cm, length 20 cm) containing ∼3.5 g of activated Florisil to separate out lipophilic matrix compounds. The column was eluted with 100 mL of n-hexane/dichloromethane, 1:1. The eluate was concentrated to dryness, and the residue was extracted three times with 2 mL of diethyl ether. The coimbined extracts were transferred to a vial and evaporated to dryness under nitrogen, and the residue was dissolved in 200 µL of toluene. Analysis. GC/MS analysis was carried out using a HewlettPackard (HP) 5971 mass spectrometer in the SIM mode coupled with an HP 5890 capillary column gas chromatograph equipped with an HP 7673 autosampler. An HP ultra 2 capillary column (25 m × 0.32 mm i.d. × 0.52 µm film thickness) was used. The carrier gas was helium (purity 5.0; Linde GmbH, Ho¨llkriegelskreuth, Germany). One microliter of sample was injected in the splitless mode at 0.75 min. The temperature program was as follows: initial temperature 60 °C held for 1 min, followed by a 10 °C min-1 ascent to 260 °C, maintained for 1 min. Analytical parameters such as detection limit and identification and quantification of the POPs 1298 Analytical Chemistry, Vol. 72, No. 6, March 15, 2000
were determined using internal standard solutions. Recovery in the soil matrix for organochlorines ranged from 90% for γ-HCH to 97% for PCB 153. PAH recovery was 80-90% for pyrene, phenanthrene, anthracene, and fluoranthene and 50-75% for benzo[a]pyrene.24 RESULTS AND DISCUSSION Preliminary Investigations. Extraction Efficiency Depending on the Method and Solvent Used. Some initial investigations were performed to optimize the ASE procedure. First of all, the dependence of extraction efficiency on different extraction methods and solvents was studied. The traditional SOX and USE procedures were compared with ASE by using double or triple determinations of mixed samples from Ro¨sa (sandy soil being a fairly uncomplicated matrix) and testing different solvents (cf. Table 1). On the basis of previous studies involving plant material, the extraction temperatures for the ASE soil extractions chosen were 40 and 120 °C.6 All samples were extracted at a constant pressure of 15 MPa. Varying the pressure in this range does not affect the results.16,20 Comparison between the method used and different solvents showed that, under these conditions, ASE with toluene as the extraction solvent produced the best yields for the simultaneous extraction of POPs (cf. the ∑POPs column in Table 1). The largest increases compared to the SOX results were found for the HCHs (about 2-fold) and the PAHs (about 4-fold). By contrast, comparison with results for the solvent combination n-hexane/CH2Cl2, 1:1, revealed the extraction yields for organochlorines to be even higher with USE than with ASE, emphasizing how important it is to choose the right extraction solvent. According to previous experience, the extraction of POPs depends on both the level of total pollution by contaminants at the location (multiple contamination) and the degree of complexity of the matrix (composition of the soil components, e.g. high proportions of humic substances). Toluene was therefore proposed instead of acetone/n-hexane, 1:1, for the better extraction of DDX, HCHs, and PAHs from highly contaminated soils (toxic wastes). The effect of using toluene was especially striking in the case of PAHs, their extraction efficiency being increased at least 2-fold.21 Comparison of ASE and Soxhlet Using Real Soils. ASE and SOX were performed with the extraction solvent toluene using two extraction steps at 40 and 120 °C for samples from the contaminated sites in southern Russia. The locations were divided into three different test areas by geographical factors (cf. Table 2). The sample material was only sufficient for a single determination. Apart from a few exceptions (e.g., at site Ki-P for HCHs and PAHs), the ASE extraction yields were considerably higher than those with SOX. The difference in the efficiencies of the two extraction methods at each location is expressed in Tables 2 by the factor f (f ) concentration of pollutant group for ASE/ concentration of pollutant group for SOX). The extraction yields with ASE were up to 10 times higher for HCHs, up to 19 times higher for the DDX group, up to 5 times higher for the chlorobenzenes, and as much as 24 times higher for the PAHs. In many cases, PCBs could only be detected by use of ASE. It turned out that the degree of improvement of the extraction efficiency greatly depended upon the matrix composition at each (24) Wenzel, K.-D.; Weissflog, L.; Paladini, E.; Gantuz, M.; Guerreiro, P.; Puliafito, C.; Schu ¨u ¨ rmann, G. Chemosphere 1997, 34, 2505-2518.
Figure 2. RSD’s for Ro¨sa soil samples depending on temperature (triple determinations).
location. These promising findings were the reason to investigate ASE’s dependence on temperature. ASE Optimization Attempts To Determine Extraction Efficiency Depending on Temperature. The temperature is one of the most important parameters for ASE. Applications generally use temperatures between 75 and 125 °C. Although the standard temperature for all environmental applications recommended by Dionex is 100 °C,25 two extraction steps at 40 and 120 °C were found to be more suitable for biomonitoring samples of pine needles and mosses.6 Therefore, the dependence of extraction efficiency on temperature was determined and compared to that of SOX. Real soil samples were studied over a very wide temperature range to ascertain the highest extraction efficiency. Table 3 (Hettstedt) and Figure 1 (Ro¨sa) show the extraction efficiencies of ASE resulting from quadruple and triple determinations, respectively. Extraction was carried out in the temperature range between 20 and 180 °C in 20 °C steps using the solvent toluene. Surprisingly, the highest extraction yields were achieved for samples from Hettstedt at 80 °C for two groups of pollutants (DDX and PCBs), whereas for all other substances, the highest yields were only attained at 140 °C. For Ro¨sa samples, the best extraction temperature was always 140 °C. At Hettstedt, ASE produced extraction yields between 1.1 and 20 times higher than those of SOX (cf. Table 4a). At Ro¨sa, the increases for the individual groups of pollutants were between 2.4- and 13-fold (cf. Table 4b). Key Temperatures for Extraction Efficiency in Soil. The difference in extraction behavior of the groups of pollutants studied may be attributable to the sharp differences in soil composition, with the sandy soil from Ro¨sa contrasting with soil from Hettstedt, which contains a high level of carbon and consists of the fertile Magdeburg Bo¨rde soil. It could also be attributable to the multiple contamination of the Hettstedt samples (due to local large-scale industrial activities) compared to the lower pollution in the woodland area at Ro¨sa. The extraction temperature (as had already been found for plant samples6) also played a crucial role in the (25) Technical Note ASE 208; Dionex: Sunnyvale, CA, 1999.
case of real soil samples. Contacts between the pollutants and the matrix in such samples can (in contrast to studies with spiked soil material) continue for long times, lasting months or even years. Depending on the level of organic substances (including humic substances), this can lead for individual substances in the long term to strong bonds in the soil and thus to soil-bound residues.26,27 Certain intermolecular bonds can only be broken at higher temperatures and pressures, and this is probably the decisive advantage of ASE compared to traditional techniques. The extraction efficiency continuously increased with temperature for the soil from Ro¨sa until about 140 °C but then declined as of 160 °C. It is conceivable that in a certain temperature range (e.g., >80 and >140 °C), adhesions or fusions of organic soil material containing pollutant fractions limit the extraction efficiency, which depends on the content of organic matrix substances. What is also remarkable is the increase for all pollutant groups by a factor of 2-3 by increase of the extraction temperature from 100 to 140 °C (cf. Figure 1), especially in view of Dionex’s recommendation of 100 °C.22 Even in the soil from Hettstedt, an extraction temperature of 100 °C proved not to be optimal. Eighty and 140 °C could be two key temperatures at which the highest extraction efficiency is reached, depending on soil parameters, the degree of contamination, and the pollutant group. Possible biological processes between 80 and 140 °C cannot at present be explained. During the extraction of POPs from moss samples for biomonitoring, these key temperatures were (in contrast to the case of soils) 40 and 120 °C. This certifies that the extraction efficiency greatly depends on the composition of the matrix, the solvent, and the temperature selected. Dependence of the RSD on the Extraction Temperature. The mean relative standard deviations in the temperature range 40-140 °C were 10.7% (cf. Figure 2). The only exceptions were δ- and -HCH, which had average RSDs of 33.3% and 24.0%, respectively, and which represent problematical substances when (26) Klein, W.; Scheunert, I. Bound pesticide residues in soil, plants and food with particular emphasis on the application of nuclear techniques. In: Agrochemicals; IAEA: Wien, 1982; pp 177-205. (27) Lichtenstein, E. P. Residue Rev. 1980, 76, 147-153.
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analytical parameters are determined for both contaminated soils and plant compartments.6 The reasons for this could be conversion or degradation reactions in the extraction step or the thermal conditions in GC/MS, and they are therefore not taken into account for the HCHs in Figure 2. Depending on the substance of interest, the higher temperatures of 160 and 180 °C lead to a considerable increase in the RSD up to 47%. This can be attributed to the high fractions of matrix components in the extract, which may have a negative impact on cleanup and analysis (interference peaks). At 20 °C, the quality of the reproducibility of the findings is inadequate owing to the excessively mild extraction conditions in the extraction cell (RSD ∼24.9%). The partly lower RSD values obtained with the use of certified and spiked materials19 are understandable because, in contrast to real soil material, the matrixes are simpler and are not multiply contaminated. CONCLUSIONS The real quality of a procedure for the extraction of POPs from environmental samples can only be evaluated using natural samples, assuming the various extraction methods can be compared in terms of extraction efficiency for recovery values in spiked and/or certified materials. When the operating variables solvent and temperature are optimized, ASE is an extraordinarily effective extraction method, especially for highly contaminated sample material and a complicated soil matrix (higher content of organic matter, complicated soil composition, dependence on physicochemical parameters such as molecule size, aromaticity and
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lipophilicity of substances analyzed, and modification by the chemical environment, e.g. pH and salts). The best extraction solvent used in this study for the ASE of real soil samples is toluene. The extractions should be performed in two temperature steps at 80 and 140 °C for an extraction time of 3 × 10 min. When using ASE, the extraction efficiency for POPs was increased by a factor of 10 or even more compared to SOX. The extraction temperatures of 80 and 140 °C represent the key temperatures for the extraction of POPs from the soil matrix at which (just as with plant samples, albeit at 40 and 120 °C) the highest extraction efficiency can be achieved depending on the soil parameters, the degree of multiple contamination of the sample, and the physicochemical properties of the substances to be analyzed. ACKNOWLEDGMENT We thank Mrs. B.Mothes, Mrs. M. Petre, Mrs. A. Sperreuter, Mrs. J. Kru¨ger, and Mrs. M. Heinrich (Dipl.-Ing.) for their excellent technical assistance in performing sample preparation, cleanup, and GC/MSD analysis of the comparative samples. The soil samples from southern Russia used in this work were provided by INTAS-COPERNICUS (Project ECCA, No. PL 963203), which is supported by the European Union. Received for review August 31, 1999. Accepted December 15, 1999. AC991005L
