Anal. Chem. 1986, 58,721-723
Registry No. XAD-2 resin, 9060-05-3; AS-THC, 1972-08-3; ll-OH-AS-THC,36557-05-8; 11-nor-9-carboxy-Ag-THC,5635406-4. LITERATURE CITED (1) Foltz., R . L. Adv. Anal. Toxicol. 1984, 7 , 125-157. (2) Borman, S. A. Anal. Chem. 1985, 5 7 , 651A. (3) St. Onge, L. M.; Dolar, E.; Anglim, M. A,; LeIst, C. J., Jr. Clin. Chem. (Winston-Salem. N . C . ) 1979, 2 5 , 1373-1376. (4) Hux, R. A,; Mohammed, H. Y.; Cantwell, F. F. Anal. Chem. 1982, 5 4 , 113-1 17. ( 5 ) Balkon. J. B.; Donnelly, B.; Prendes, D. J . Forensic Sci. 1982, 2 7 , 23. (6) Kennedy, E. R.; Hill, R. Anal. Ch8m. 1982, 5 4 , 7739-1742. (7) Lipori, E. Anal. Chem. 1984, 5 6 , 1820-1826.
721
(8) Zhu, A. N.; Xu, Gui-Yun J . Chromatogr. 1984, 314, 421-428. (9) Rosenfeld, J. M.; Murelka-Russell, M.: Phatak, A. J . Chromatogr. 1984, 283, 127-135. (IO) Poole, C. F.; Schuette, S.A. HRC C C , J . High Resolut. Chromatogr. Chromatogr. Commun. 1983, 6 , 526-547. (11) Rosenfeld, J. M.; Taguchl, V . Y. Anal. Chem. 1976, 47, 726-729. (12) Rosenfeld, J . M. Anal. Lett. 1978, 10, 917-925.
RECEIVED for review August 1, 1985. Accepted October 25, 1985. This work was supported by the National Institute on Drug Abuse of the USA and was presented in part at the Satellite Symposium on Cannabinoids held a t Oxford, England, August 1984.
Extraction of Anthracene and Benzo[ a Ipyrene from Soil Peter J. A. Fowlie* and Terri L. Bulman Wastewater Technology Centre, Canada Centre for Inland Waters, Burlington, Ontario L7R 4A6, Canada
Extraction of 14C labeled benzo[a Ipyrene and anthracene from contamlnated sol1 samples by Soxhlet and Poiytron techniques was studled in a replicated 24 factorlal experlment. Soxhlet extractlon gave higher recoveries than Polytron extraction. Percent recoverles from both techniques were greater at a 50 pg/g contamination level than at 5 pg/g. Pyrolysls of the extracted residue followed by trapping of I4CO2 corroborated the extraction results. I n addition, analysis of the residue showed a slgnificant increase in I4C due to soil steriliratlon with HgCi, and an interaction effect between PAH and concentratlon. The effects of PAH, concentratlon, and HgCi, treatment on extractlon and sorption were the same with both extractlon methods.
The extraction of 14Clabeled polynuclear aromatic hydrocarbons (PAH’s) from contaminated soil samples has been studied as part of the Wastewater Technology Centre’s (WTC) program to determine the fate of PAH’s applied to land. This program, including the Donnybrook sandy loam, has been described by Bulman et al. (1). The soil samples were spiked with labeled and unlabeled benzo[a]pyrene, or anthracene at 5 and 50 Fg/g soil. The samples were incubated in biometer flasks a t 20 “C for 3 and 5 months for anthracene and benzo[a]pyrene, respectively, allowing degradation to be monitored and the PAH to interact with the soil matrix. These samples provided a unique opportunity to assess the effects of extraction method on PAW extractability and sorption onto soil under conditions that approach “natural incorporation” of the spikes as suggested by MacDougall et al. (2). Haddock et al. (3)compared a 1-and 8-day “spike time” for anthracene and found significantly reduced recovery a t the longer time. A common problem encountered in the interpretation of PAH data is the variability resulting from different extraction procedures. In this study, subsamples were taken from each flask and extracted by one of two methods: by overnight Soxhlet extraction with 1:1 hexane:acetone (Maybury et al. (4))or by extraction in a Polytron homogenizer (Brinkman Instruments, Ltd.) with three successive 25-mL portions of acetone (Afghan and Wilkinson (5)). Both of these extractions are routinely used in the WTC laboratory.
EXPERIMENTAL SECTION Radiochemicals. 14C-labeledchemicals used as tracers were obtained from California Bionuclear Corp., Sun Valley, CA. Benzo[a]pyrene (7,1O-l4C,CBN 114) and anthracene (9-14C,CBN 086) were purchased at 98% radiochemical purity and were used as received. The labeled compounds were dissolved in toluene, 1mL of which was pipetted evenly over each 50 g biometer flask soil sample. The moist soil was mixed for several minutes with a glass rod and then left open to air overnight to evaporate the toluene. Benzo[a]pyrene was added to the soil samples at 2.76 x IO5 dpm/g while anthracene was added at 2.40 X lo5 dpm/g. Instruments. All samples were counted for 14C on a LKB Model 1217 liquid scintillation counter. The external standard ratio (ESR) method of automatic window setting and the ESR method of quench correction for disintegrations per minute calculation WBS used. Chemiluminescence was routinely monitored by the instrument. Combustion analysis of the samples was carried out in a Biological Material Oxidizer (BMO/R. J. Harvey Instrument Corp.). Standards and Reagents. All samples were counted in 22-mL plastic vials with 10 mL of Amersham PCS liquid scintillator and varying volumes of sample. Standards were prepared by dissolving one LKB-Wallac 14C-0 Internal Standard pellet (14C= 105400 dpm) in 10 mL of PCS and adding various volumes of blank sample to the PCS. Several standards formed the quench calibration curve which was stored in the instrument memory for automatic disintegrations per minute calculation. The C02 trapping solution was Oxisorb CO, (New England Nuclear) mixed 1:2 with PCS. Methods. Soxhlet. A 10-g soil sample of 29% moisture content was weighed into a Soxhlet extraction tube and extracted 16-18 h with 300 mL of 1:l hexane:acetone. The recovered solvent was evaporated to 15 mL on a rotovap. One milliliter was taken for 14Ccounting, 50 fiL for thin-layer chromatography, and the soil residue reserved for combustion analysis. Polytron. A 10-g soil sample of 29% moisture content was weighed into a 40-mL centrifuge tube and 25 mL of acetone added. The sample was extracted 2 min with a Polytron homogenizer (a high-velocity mixer/shearer). The tube was then centrifuged for 5 min in a clinical centrifuge at 2000 rpm and the supernatant decanted. This procedure was repeated two more times with acetone and the three extracts combined. The volume of recovered acetone was recorded and a 1-mL sample taken for 14Ccounting. The soil residue was reserved for combustion analysis. Thin-Layer Chromatography (TLC). With a microliter syringe, 50 fiL of the Soxhlet extract was applied to the preadsorbent area
0003-2700/86/0358-0721$01.50/0 Published 1986 by the American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 58, NO. 4, APRIL 1986
Table I. Percent Recovery of 14Cin Extract and Residue" extraction Soxhlet
PAHb concn, pg/g HgClz, % BaP
50 5
Anth
50
5
Polytron
BaP
50 5
Anth
50 5
extract
residue
0.5 0 0.5 0
79.3 77.1 74.9 77.8
3.1 2.5 9.3 6.6
0.5 0 0.5 0
76.4 75.5 75.8 61.1
3.3 3.0 5.3 4.5
0.5 0 0.5 0
68.2 71.9 63.0 64.9
4.6 4.6 12.7 9.6
0.5 0 0.5 0
67.5 68.3 52.2 50.5
7.4 3.8 10.5 8.9
" Each value is the mean of 3 replicates. bBaP,benzo[a]pyrene; Anth, anthracene. of each of two TLC plates (Whatman LK5DF silica gel, 250 pm thickness, 20 X 20 cm plate). Several other samples and a standard were applied to alternating lanes of the TLC plate. The plates were developed in ascending fashion, one plate in 1:1:1 hexane:acetone:toluene (Rf = 0.9) and the other plate in hexane (Rf = 0.3) to a height of 12-14 cm. After drying, the lanes were divided into 1 cm sections and the silica gel recovered for counting. Residue Analysis. After removal of solvent by air-drying, a sample of 0.05-0.08 g of extracted soil was burned at 900 "C in a stream of O2in a biological material oxidizer. After catalytic oxidation of combustion gases, the C02 produced was trapped and counted for 14C. Statistical Analysis. The data from the incubation experiment were analyzed as a 24 factorial experiment, which was replicated three times for a total of 48 extractions. The factors were as follows: (1) Soxhlet vs. Polytron extraction, (2) anthracene vs. benzo[a]pyrene, (3) 5 pg/g vs. 50 pg/g concentration, and (4) HgCl, (0.5% w/w) for sterility vs. no HgC12for biologically active soil. Dependent variables measured were (1)14Cin the solvent after extraction and (2) 14Cin the extracted soil residue by combustion in a stream of O2 and trapping of 14C02. The I4C in the solvent was confirmed to be parent PAH by thin-layer chromatography (TLC). Identification of residual 14C compound was not possible as the 14C was burned to 14C02.
RESULTS AND DISCUSSION Analysis of variance was used to assess the effects on PAH extraction a t the 99% confidence level for the four factors varied. The percentage of 14C in the extract and soil residue does not total 100% because of degradation and volatilization during incubation as discussed by Bulman et al. (I) and due to losses during analysis. The data are presented in Table I and represent the average of the three replicates for the extract or soil residue. Extraction into Solvent. PAH recovery of the individual extracts ranged from 36.1% to 87.4%. Significant differences at the 99% confidence level were observed for the extraction technique and for the PAH concentration in the soil (Table 11). The average recovery by the Soxhlet technique was 74.5% whereas 62.8% was the average Polytron recovery. A much higher proportion was extracted with PAH at the 50 pg/g level (72.6%) than a t the 5 pg/g level (64.6%) suggesting that the extraction efficiency is not constant with concentration, an effect that was observed for both PAHs. A similar effect was found by Harrison et al. (6) in their extraction of [I4C]benzo[a]pyrene from fly ash. No significant interaction effects were observed among the variables indicating that the main effects operate in an additive manner. Significant effects are summarized in Table 111. No zone of activity other than that
Table 11. Analysis of Variance Table for Extraction and Soil Residue
source of variation extraction (E) PAH (P) concentration (c) sterility (S) EP EC ES PC PS
cs
EPC EPS ECS PCS EPCS residual
extraction mean square F
square
F
6.034 1.626 2.813 0.046 0.057 0.509 0.316 0.929 0.439 0.166 0.123 0.036 0.005 0.209 0.246 0.346
4.078 0.239 7.829 0.863 0.192 0.242 0.076 0.733 0.009 0.058 0.029 0.159 0.077 0.406 0.034 0.055
74.15" 4.34 142.35" 15.69" 3.49 4.40 1.38 13.33" 0.16 1.05 0.53 2.89 1.40 7.38 0.62
soil residue mean
17.44" 4.70 8.13" 0.13 0.16 1.47 0.91 2.68 1.27 0.48 0.36 0.10 0.01 0.60 0.71
degrees of freedom:
total, 47 residual, 32 others, 1
F (1,32, 0.01) = 7.50
Significant effect at 99% level. Table 111. Table of Means for Significant Effects at the 99% Confidence Level Extract extraction
Soxhlet Polytron
74.5% 62.8%
concentration
50 pg/g level 5 pg/g level
72.6% 64.6%
Soil Residue extraction
Soxhlet Polytron
4.5% 7.4%
concentration
50 pg J g level 5 pg/g level
4.0% 8.1%
sterility
sterile (Hg) unsterile
6.6% 5.2%
PAHconcentration interaction
Anth" at 5 Anth at 50 BaP at 5 BaP at 50
7.0% 4.2% 9.3% 3.8%
=Anth,anthracene; BaP, benzo[a]pyrene. for parent PAH was observed on any of the thin-layer chromatograms. Soil Residue by Combustion. After overnight Soxhlet treatment with 1:lhexane:acetone or three successive Polytron extractions with acetone, 2.2%-16.1% of the added radioactivity was found associated with the solids. Results of the analysis of variance are presented in Table 11. Significant main effects a t the 99% confidence level were found for the extraction technique and for the PAH concentration and correspond to the significant effects found for the extract data. Polytron and Soxhlet treatment left 7.4% and 4.5% of activity, respectively, associated with the soil, which is in accordance with the greater extraction efficiency noted for the Soxhlet technique. A higher percentage (8.1%) of activity was found on the soil spiked initially with 5 pg/g PAH than with 50 pg/g (4.0%),also consistent with the extract data. In addition, a significant difference was found with HgCl, addition. A higher percentage (6.6%) of the activity was
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Anal. Chem. 1906, 58,723-725
associated with HgClz treated soils than with untreated soils (5.2%). The difference of 1.4% due to HgClz was significant for the soil residues but was too small to have produced a significant effect in the extract. There was also a significant interaction term between PAH and PAH concentration. The highest percentage of bound 14C (9.3%) occurred with benzo[a]pyrene with the 5 pg/g treatment and the smallest percentage (3.8%) occurred with benzo[a]pyrene at the 50 pg/g treatment. The difference in bound 14Cwas much less pronounced for anthracene where the soil residue contained 7.0% of 14Cat the low concentration and 4.2% at the high concentration and was consistent for both extraction procedures. The mechanism for this effect is not clear because the effect of PAH is confounded with differences in experimental conditions such as the length of incubation time. Although the PAH main effect was not significant, the significant interaction term between type of PAH and concentration could have been caused by differences in experimental conditions. Recovery of PAH's from environmental matrices is influenced by many factors. Results of this study indicate that PAH concentrations are likely to be underestimated and that the degree of underestimation is likely to be greater for low than for high concentrations. Underestimation should be minimized by using the most efficient extraction technique, in this case the Soxhlet method. The most efficient technique
should be determined for each sample matrix.
ACKNOWLEDGMENT The authors thank Kevin Hosler for preparing the contaminated soil samples and David Ide for his work on the extractions and TLC. Registry No. Benzo[a]pyrene, 50-32-8;anthracene, 120-12-7. LITERATURE CITED (1) Bulman, T. L.; Lesage, S.; Fowlie, P. J. A.; Webber, M. D. "The Persistence of Polynuclear Aromatic Hydrocarbons in Soil"; Petroleum Assoclatlon for Conservation of the Canadian Environment (PACE), Report No. 85-2, Ottawa, Ontario, November 1985. (2) MacDougall, D.; Crummett, W. Anal. Chem. 1980, 52, 2242-2249. (3) Haddock, J. D.; Landrum, P. F.; Giesy, J. P. Anal. Chem. 1983, 55, 1197-1200. (4) Maybury, R. "Laboratory Manual for Pesticlde Residue Analysis in Agricultural Products", Food Production and Inspection Branch, Agriculture Canada, Revised 1984. (5) Afghan, B. K.; Wilkinson, R. J. "Method for Determination of Polynuclear Aromatic Hydrocarbons in Environmental Samples (HPLC-multidetectlon system); Environment Canada, Manuscrlpt 20-AMD-3-81BKA, 1981. (6) Harrison, F. L.; Bishop, D. J.; Mallon, B. J. Environ. Sci. Techno/. 1985, 19, 186-193.
RECEIVED for review July 15, 1985. Accepted October 24, 1985. This work was supported by the Petroleum Association for Conservation of the Canadian Environment (PACE) and the Great Lakes Water Quality Program.
Determination of Munitions in Water Using Macroreticular Resins J. J. Richard and G. A. Junk*
USDOE,Ames Laboratory, Ames, Iowa 50011
A simple procedure has been developed for the determlnatlon of munltlons present In water at levels as low as 0.1 pg/L. The method uses 60-80 mesh XAD-4 for extraction, followed by elutlon with 20 mL of ethyl acetate, concentration of the eluate, separation by caplllary GC, and detectlon using electron capture. Contrived and fleld samples contalnlng nltrobenzene, 1,3-dlnltrobenzene, 1,3,5-trlnltrobenzene, 2,4-dlnltrotoluene, 2,6dlnltrotduene, 2,4,&trlnltrotoluene (TNT), and 1,3,5-trlnltrohexahydro-1,3,5-trlarlne (RDX) have been tested successfully. The XAD-4 extractlon In the field reduced the cost of transporting water samples to the laboratory where addltlonal savlngs occurred because less solvent was used to elute the resln than was required for solvent extractlon. Adsorptlon onto partlculate matter, assoclatlon with humlc adds, resln capaclty, resln polarity, and Incomplete dlssolutlon were examlned as the most likely causes of the low recoverles for RDX when XAD-2 and XAD-7 resins were used.
Nitrated organic compounds are introduced into the environment from the production, storage, transport, and disposal of munitions. Wastewater from munitions production and contaminated surface water and groundwater from the disposal of munitions are of greatest concern. The nitrated organic compounds in these waters are usually isolated by liquid-liquid extraction (I-@, and the concentrated extracts me analyzed by some form of chromatography. The extraction 0003-2700/86/0358-0723$01 S O / O
procedure is best accomplished in the laboratory; consequently transport of water samples in cooled containers is the normal procedure. The costly transport can be avoided by sorbing the organic compounds onto a macroreticular resin in the field and then transporting the small column of resin at ambient temperature to the laboratory for further processing. This advantage of resins has not been exploited in the reports of their use for the isolation of nitrated organic compounds (6-9). Resin use has been limited to XAD-2 for the cleanup of wastewaters from the manufacture of trinitrotoluene (6))a mixture of XAD-4 and XAD-8 to remove nitrated compounds from river water (7), and XAD-4 and Porapaks, R and S, to extract various explosives from surface waters (8, 9). This paper describes a convenient procedure for the efficient recovery of two munitions, 1,3,5-trinitrohexahydro-l,3,5-triazine (RDX) and 2,4,6-trinitrotoluene (TNT), from water using the macroreticular resin XAD-4. Also discussed are the recovery results for related compounds, 2,4-dinitrotoluene (2,4-DNT),2,6-dinitrotoluene (2,6-DNT),1,3,5-trinitrobenzene (1,3,5-TNB),1,3-dinitrobenzene(1,3-DNB),and nitrobenzene (NB), from both contrived and groundwater samples.
EXPERIMENTAL SECTION Reagents and Chemicals. All solvents were reagent grade and distilled prior to use. The nitrobenzenes and nitrotoluenes were obtained from Chem Service (West Chester, PA). The RDX was obtained from Army Ordinance. The XAD-2, XAD-4, and XAD-7 solid sorbents are macroreticular resins obtained from 0 1986 American Chemical Soclety
