Enhanced Detection of Nitroaromatic Explosive Vapors Combining

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Anal. Chem. 2003, 75, 3137-3144

Enhanced Detection of Nitroaromatic Explosive Vapors Combining Solid-Phase Extraction-Air Sampling, Supercritical Fluid Extraction, and Large-Volume Injection-GC Ramo´n Batlle,*,† Håkan Carlsson,†,‡ Petter Tollba 1 ck,† Anders Colmsjo 1 ,† and Carlo Crescenzi†

Department of Analytical Chemistry, Stockholm University, 106 91 Stockholm, Sweden, and FOI, Swedish Defense Research Agency, Department of Energetic Materials, 147 25 Tumba, Sweden

A complete method for sampling and analyzing of energetic compounds in the atmosphere is described. The method consists of the hyphenation of several techniques: active air sampling using a solid-phase extraction cartridge to collect the analytes, extraction of the sorbed analytes by toluene/methyl tert-butyl ether modified supercritical fluid extraction (SFE), and analysis of the extract by large-volume injection GC-nitrogen/phosphorus detection. The GC system is equipped with a loop-type injection interface with an early solvent vapor exit, a utilizing concurrent solvent evaporation technique. Chemometric approaches, based on a Plackett-Burman screening design and a central composite design for response surface modeling, were used to determine the optimum SFE conditions. The relative standard deviations of the optimized method were determined to be 4.3 to 7.7%, giving raise to method detection limits ranging from 0.06 to 0.36 ng in the sampling cartridge, equivalent to 6.2-36.4 pg/L in the atmosphere, standard sampling volume 10 L. The analytical method was applied to characterize headspace composition above military grade trinitrotoluene (TNT). Results confirm that 2,4-dinitrotoluene (DNT) and 1,3-dinitrobenzene (DNB) constitute the largest vapor flux, but TNT, 2,6-DNT, and trinitrobenzene TNB were also consistently detected in all the samples. The development of analytical methods for the detection of chemical vapors associated with explosives is of interest for a variety of reasons, including the detection of unexploded ordnance (UXO) and landmines.1 Presently, trained dogs are thought to be the most dependable mine detectors.2 However, research is actively being pursued for the detection of explosives by using sensors,3-5 solid-phase microextraction,6 or air sampling with solid sorbents.7,8 Solid-phase extraction (SPE) methods utilizing organic polymer resins or reversed-phase silicas have been used for extraction of * Corresponding author. Fax: + 46 8 156391. E-mail: ramon.batlle@ anchem.su.se. † Stockholm University. ‡ Sweden Defense Research Agency. (1) Yinon, J. Trends Anal. Chem. 2002, 21, 292-301. (2) Furton, K. G.; Myers L. J. Talanta 2001, 54, 487-500. 10.1021/ac0207271 CCC: $25.00 Published on Web 05/08/2003

© 2003 American Chemical Society

explosives in the field.9-12 However, SPE materials have been rarely used as sampling media for the analysis of air samples,13-16 and to the best of our knowledge, none of the published cases has been related to the analysis of energetic material, such as the nitroaromatic congeners. Analytes sorbed onto the SPE materials are desorbed using organic solvents, and the extract obtained usually requires further treatment, for example, a sample cleanup or a solvent reduction step, prior to analysis. These steps are tedious, labor-intensive, and can be error-prone, especially if the analytes are unstable, such as compounds originating from energetic material. Hence, the use of a more selective desorption technique with lower solvent requirements could clearly be very useful to overcome these problems. Supercritical fluid extraction (SFE) is an environmentally friendly and efficient extraction technique that reduces the use of organic solvents, extracts samples quickly, and simplifies concentration and cleaning of the extracted analyte. SFE has also been applied to the analysis of energetic materials,17,18 especially for the extraction of nitroaromatic compounds from native or spiked soils.19-21 (3) Sylvia, J. M.;. Janni, J. A.; Klein, J. D.; Spencer K. M. Anal. Chem. 2000, 74, 5834-5840. (4) Houser, E. J.; Mlsna, T. E.; Nguyen, V. K.; Chung, R.; Mowery, R. L.; McGill, R. A. Talanta 2001, 54, 469-485. (5) Albert, K. J.; Myrick, M. L.; Brown, S. B.; James, D. L.; Milanovich, F. P.; Walt, D. R. Environ. Sci. Technol. 2001, 35, 3193-3200. (6) Jenkins, T. F.; Leggett, D. C.; Miyares, P. H.; Walsh, M. E.; Ranney, T. A.; Cragin, J. H.; George, V. Talanta 2001, 54, 501-513. (7) Sigman, M. E.; Ma, C.; Ilgner, R. H. Anal. Chem. 2001, 73, 792-798. (8) Hable, M. A.; Sutphin, J. B.; Oliver, G. C.; McKenzie, R. M.; Gordon, E. F.; Bishop, R. W. J. Chromatogr. Sci. 2002, 40, 77-82. (9) Jenkins, T. F.; Miyares, P. H.; Myers, K. F.; McCormick, E. F.; Strong, A. B. Anal. Chim. Acta 1994, 289, 69-78. (10) Walsh, M. E.; Ranney, J. J. Chromatogr. Sci. 1998, 36, 406-416. (11) Cassada, D. A.; Monson, S. J.; Snow, D. D.; Spalding, R. F. J. Chromatogr., A 1999, 844, 87-95. (12) Lu, Q.; Collins, G. E.; Smith, M.; Wang, J. Anal. Chim. Acta 2002, 469, 253-260. (13) Markell, C.; Hagen, D. F.; Bunnelle, V. A. LC-GC, 1991, 9, 332-345. (14) Koostra, P. R.; Herbold, H. A. J. Chromatogr., A 1995, 697, 203-211. (15) Thomsen, C.; Leknes, H.; Lundanes, E.; Becher, G. J. Chromatogr., A 2001, 923, 299-304. (16) Ramil, M.; Rodrı´guez, I.; Cela, R. J. Chromatogr., A 2002, 963, 65-71. (17) Teipel, V.; Gerber, P.; Krause, H. H. Propellants Explos. Pyrotech. 1998, 23, 82-85. (18) Ashraf-Khorassani, M.; Taylor, L. T. J. Chem. Ing. Data 1999, 44, 12541258.

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A critical step in the SFE analysis of energetic substances is trapping the analytes after extraction.21,22 In a previous paper, the use of porous graphitic carbon (PGC) to efficiently retain nitroaromatic explosives and the suitability of its mechanical and chemical properties for use with supercritical fluids have been demonstrated.23 PGC has also proved to be compatible with a large range of organic solvents, which allows desorption to be fine-tuned. The concept behind the combination of SFE and PGC is to achieve an increase in selectivity for the whole methodology by taking advantage of SFE well-known selective features and the PGC selective desorption (neutral, acidic, or basic compounds can be sequentially eluted from the trap, depending on the solvent used). The limited injection volumes that can be used with conventional GC injectors, 1-3 µL, allow only a small fraction of the sample to be introduced into the gas chromatographic system. To overcome this limitation, several large-volume injection (LVI) techniques for GC have been developed.24-26 In the study described in this paper, analytes were transferred utilizing fully concurrent solvent evaporation with a loop-type interface.27-29 The paper presents a new combined sampling and analytical method for the detection of nitroaromatic vapors that may evolve from explosives. The method uses a commercial SPE cartridge for air sampling, SFE with toluene/MTBE-modified carbon dioxide for extraction of the sorbed analytes, PGC as the retention trap, and a loop-type large-volume injection (LVI) to introduce the effluent into the GC system. The optimized method has been applied to air samples obtained from the headspace of a desiccator containing military grade 2,4,6-trinitrotoluene (TNT). EXPERIMENTAL SECTION Chemicals and Materials. The nitroaromatic reference substances were obtained from several sources: 1,2- (1,2-DNB, CAS identification number 528-29-0) and 1,3-dinitrobenzene (1,3-DNB, CAS 99-65-0) were purchased from Dr. Ehrenstorfer (Augsburg, Germany). 2-3-, 2,4-, 2,5-, 2,6-Dinitrotoluene (2,3-, 2,4-, 2,5-, and 2,6-DNT, CAS 602-01-7, 121-14-2, 619-15-8, and 606-20-2, respectively), and 2,4,6-trinitrotoluene (TNT, CAS 118-96-7) were obtained from the Swedish Defense Research Agency (Tumba, Sweden). 1,3,5-Trinitrobenzene (TNB, CAS 99-35-4) and 3,5dinitrotoluene (3,5-DNT, CAS 618-85-9) were purchased as 100 µg/mL acetone solutions from Promochem (Wesel, Germany). Methanol, toluene, acetonitrile, acetone, and dichloromethane (Suprasolv quality), as well as tetrahydrofuran (THF; Lichrosolv quality), were from Merck (Darmstadt, Germany). Methyl tertbutyl ether (MTBE) was from Rathburn Chemicals Ltd. (Walkeburn, Scotland). (19) Francis, E. S.; Wu, M.; Farnswoth, P. B.; Lee, M. K. J. Microcolumn Sep. 1995, 7, 23-28. (20) Wujcik, C. E.; Seiber, J. N. J. Environ. Sci. Health, A 1996, 31, 1361-1377. (21) Deuster, R.; Lubahn, N.; Friedrich, C.; Kleibo ¨hmer, W. J. Chromatogr., A 1997, 785, 227-238. (22) Engelhardt, H.; Zapp, J.; Kolla, P. Chromatographia 1991, 32, 527-537. (23) Batlle, R.; Carlsson, H.; Holmgren, E.; Colmsjo ¨, A.; Crescenzi, C. J. Chromatogr., A 2002, 963, 73-82. (24) Grob, K. On-Line Coupled LC-GC; Hu ¨ thig: Heidelberg, 1981. (25) Engewald, W. T.; Teske, J.; Efer, J. J. Chromatogr., A 1999, 842, 143-161. (26) Grob, K. J. Chromatogr., A 2000, 892, 407-420. (27) Grob, K.; Schilling, B. J. High Resolut. Chromatogr., Chromatogr. Commun. 1985, 8, 726-733. (28) Grob, K.; Schmarr, H. G.; Mosandl, A. J. High Resolut. Chromatogr. 1989, 12, 375-382. (29) Tollba¨ck, P.; Carlsson, H.; O ¨ stman, C. HRC J. High Resolut. Chromatogr. 2000, 23, 131-137.

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Preparation of Standards. Reference standard solutions were prepared in MTBE. Working mixture solutions were prepared twice a week in HPLC quality water or daily in THF (external standard). Air Sampling. Air sampling was performed with a modified personal sampler, previously described.30 The sampler consists of an anodized aluminum holder housing a Nexus SPE cartridge (Varian, Harbor City, CA). These SPE cartridges (i.d., 5 mm) containing 30 mg of highly cross-linked ultraclean coplymeric material (surface area 575 m2/g) were used without any conditioning or pretreatment. Air was pumped through the sampler using a MCS 30 air sampling pump (SKC, Eighty Four, PA). The flow rate was set to the maximum possible to maximize sampling volume and, thus, sensitivity while minimizing sampling time. Because of the backpressure of the cartridge, the flow was limited to 3 L/min. Supercritical Fluid Extraction. Supercritical fluid extractions were performed using a 1-mL stainless steel extraction vessel and an Autoprep 44 stand-alone SFE system (Suprex, Pittsburgh, PA). The system also included a MPA-1 solvent modifier pump (Varian, Walnut Creek, CA), which was used for dynamic modifier delivery, and a variable restrictor able to provide supercritical fluid flow rates in the range 0.5-7.0 mL/min. High-purity carbon dioxide (C-50 quality) under helium headspace (pressure, 13.8 MPa) was purchased from AGA (Sundbyberg, Sweden). After extraction, analytes were trapped onto a trap prepared in-house, consisting of a 30 × 1 mm slurry-packed PGC column. Hypercarb PGC material (particle size 10-40 µm) was kindly provided by Thermo Hypersil Ltd. (Cheshire, U.K.). The trap was connected to the exit of the heated restrictor module, and desorbing solvent was directed through it or to waste by an airactuated six-port switching valve (Valco, Houston, TX). Experiments based on a fractional factorial Plackett-Burman design were performed to identify significant experimental parameters that affect the supercritical extraction. The results were used to develop a response surface design (Central Composite Circumscribed, CCC, design) to finally optimize the supercritical extraction. All statistical calculations were performed with Modde 4.0 for Windows by Umetri (Umeå, Sweden). Large Volume Injection/Gas Chromatography. Preliminary air sampling and trap desorption studies were carried out on a Fisons 8000 Series GC with a NPD 800 nitrogen-phosphorus detector equipped with a TS-1-type bead (CE Instruments, Milano, Italy) and a 30 m × 0.25 mm column coated with a 0.15-µm film of (14% cyanopropylphenyl)-methylpolysiloxane (DB-1701, J&W Scientific, Folsom, CA). The column temperature was initially held at 105 °C for 2 min, then raised by 10 °C/min to 170 °C, then 5 °C/min to 280 °C, which was held for 5 min. Large volume injection (LVI) was performed using a loop-type interface with an early solvent exit, utilizing fully concurrent solvent evaporation. It was constructed from a six-port switch valve used to direct the carrier gas to the stainless steel injection loop or directly to the GC retention gap. The injection interface situated inside the GC oven (Varian 3300) consisted of a retention gap (1.5 m × 0.53 mm i.d., deactivated fused-silica, Restek Corp. Bellefonte, PA) coupled in series with a precolumn (1 m × 0.53 (30) O ¨ stman, C.; Carlsson, H.; Bermgard, A.; Colmsjo ¨, A. Polycyclic Aromatic Compounds: Properties, Analytical Measurements, Occurrence and Biological Effects; Gordon and Breach: Yverdon, 1993; pp 485-492.

mm i.d., DB-200, J&W Scientific) coated with a 0.5-µm stationary phase film of 35% trifluoropropyl-methylpolysiloxane. The outlet of the precolumn was connected to both the analytical column and the solvent vapor exit with an all-glass Y-piece press-fit connector. A three-port valve switched the vapor exit between an open line and a restrictor (1.5 m × 0.05 mm i.d., fused-silica, Restek) for purging during analysis. After injection, a carrier gas pressure drop indicates that the solvent has evaporated. The vapor exit is then closed, and the GC-oven temperature program is started to perform an ordinary GC separation. Since the cyano-containing DB-1701 column is not compatible with the NP detector, a 30 m × 0.25 mm DB-200 (J&W Scientific) analytical column, with a film thickness of 0.25 µm was used for all the experimental work involving LVI. The oven temperature was initially held at 108 °C for 2 min and was then raised 25 °C/min to 170 °C, followed by 8 °C/min to 280 °C, which was maintained for 10 min. RESULTS AND DISCUSSION Porous Graphitic Carbon Trap. Two of the key factors of the proposed method are the sorption and desorption of the analytes onto/from the trap following SFE extraction. PGC was selected for this purpose since it has been proven to have the ability to retain the target analytes effectively.23 The suitability of several solvents to desorb the analytes from the trap was evaluated regarding their ability to desorb all of the analytes sufficiently (>85%) in as small a volume as possible, to avoid the necessity of further concentration steps. Five different solvents were evaluated (four replicates each), namely, dichloromethane, MTBE, methanol, toluene, and THF. Acetonitrile, a solvent commonly used with PGC columns31 was not evaluated, since it is not compatible with the NP detector used for the GC analysis. The evaluation procedure was as follows. Using a 6-port switching valve (Valco), 10 mL of standard water solution (containing ∼10 ng/mL of the analytes) was pumped at a flow rate of 2 mL/min through the PGC column. Water was selected as the carrier solvent, since it has been proven that it is not able to elute the analytes from the trap. Then, the valve was switched, and desorption solvent was pumped through the cartridge at a flow rate of 1.0 mL/min. Fractions were collected every 30 s (corresponding to ∼500 µL) for 3 min, the extracts were evaporated to dryness, and analytes were redissolved by adding 0.6 mL of toluene and then analyzed by GC. The best results were obtained with THF, with which all the analytes were desorbed and eluted in the first 500-µL fraction. These results were not unexpected, since earlier studies had shown that THF is a strong solvent when using PGC columns.32,33 Following these evaluations, a new set of experiments was performed in order to assess whether it would be possible to reduce the elution volume. A Varian 9050 multiwavelength UV detector was connected in series to the PGC trap and used to monitor the absorbance of the effluent at 275 nm. After the solvent peak (THF) was detected, fractions were collected every 6 s (∼100 µL) for a period of 1 (31) Albin ˜ana, J.; Carlsson, H.; Crescenzi, C.; Batlle, R. Unpublished work. (32) Vial, J.; Hennion, M. C.; Ferna´mdez, A.; Agu ¨ era, A. J. Chromatogr., A 2001, 937, 21-29. (33) Gaudin, K.; Chaminade, P.; Baillet, J. J. Chromatogr., A 2002, 973, 61-68.

Table 1. Large Volume Injection (LVI) Performance, Relative Standard Deviation (% RSD) of Uncorrected Peak Areas as a Function of the Injected Volumea precision (% RSD) analyte

5 µL

15 µL

50 µL

100 µL

500 µL

2,6-DNT 1,3-DNB + 2,5-DNT 1,2-DNB 2,4-DNT 2,3-DNT 3,5-DNT TNT TNB

4.0 8.1 6.2 7.1 6.0 6.8 2.6 6.5

2.9 4.4 4.1 4.0 4.4 5.0 1.1 4.0

2.7 3.5 3.0 2.6 2.9 3.0 0.4 2.6

3.0 4.5 3.1 3.6 3.0 4.2 0.9 3.9

14.2 16.5 13.0 14.2 14.8 15.2 8.1 10.0

av

5.9

3.7

2.6

3.3

13.3

a

Absolute amount injected: 5 ng of each analyte

min, and the extracts were processed as described above. Most of the analytes were desorbed in the second fraction (6 to 12 s), with the exception of 3,5-DNT, TNT, and TNB, which also appeared in the third fraction. In the final method, a fraction was collected between 6 and 18 s, giving a total volume of 200 µL. Large-Volume Injection. Once desorption from the trap had been evaluated, large-volume GC injection was optimized. First, the effects of injection temperature were investigated. Both injection temperature and pressure influence the performance of the LVI system, and injection should be performed at approximately the same temperature as the pressure-corrected boiling point of the solvent. At this temperature, the injection pressure is equal to that of solvent vapors, giving a stable evaporation site in the retention gap. At too high or too low temperatures, solvent and dissolved analytes will reach the precolumn, resulting in distorted peaks. The pressure was fixed at a value of 160 kPa, temperatures ranging from 70 to 120 °C were evaluated, and the optimum injection temperature was found to be 108 °C. Injection volumes of 15, 50, 100, and 500 µL were evaluated in terms of reproducibility (n ) 6). As shown in Table 1, performance is acceptable up to 100 µL, and consequently, this was the volume selected, providing the best sensitivity with good chromatographic shape. Figure 1a shows a LVI chromatogram obtained from a 100-µL injection of a 0.05 ng/µL nitroaromatic standard mixture. As can be seen, good separation was obtained for the nitroaromatic congeners, except for 1,3-DNB and 2,5-DNT, which were just partially resolved. For them, method performance was calculated jointly. Air Sampling. The performance of Nexus SPE cartridges for air sampling of energetic material was evaluated with respect to the breakthrough volume. For this purpose, two Nexus cartridges were serially connected, and samples were collected at a flow rate set to 3.0 L/min from a desiccator containing 10 g of military grade TNT. Samples were taken over 6, 8, and 10 min, the latter corresponding to 10 times the air volume contained in the desiccator. After sampling, both cartridges were extracted with 0.6 mL of toluene, followed by 2 mL of acetone. Then extracts were concentrated under a gentle nitrogen stream at 40 °C to remove the acetone fraction and were further analyzed according to the conditions listed in the Experimental Section. No traces of nitroaromatic compounds were found in the second cartridge. Analytical Chemistry, Vol. 75, No. 13, July 1, 2003

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Figure 1. Large-volume injection (LVI) chromatograms of nitroaromatic compounds: (a) Standard THF solution; (b) SFE extract of a spiked SPE cartridge. Peak identification: (1) 2,6-DNT, (2) 1,3-DNB, (3) 2,5-DNT, (4) 1,2-DNB, (5) 2,4-DNT, (6) 2,3-DNT, (7) 3,5-DNT, (8) TNT, and (9) TNB.

To confirm these results, a new set of experiments was performed. Five sets of three SPE cartridges were each spiked with 10 ng of nitroaromatic compounds, and air was pumped through them at a flow rate of 3.0 L/min for 45 min. Another set of three cartridges was spiked with the same amount of analytes without pumping an air stream through them. The cartridges were wrapped in aluminum foil and stored at -4 °C. Starting the following day, the set not exposed to air and one of the air-exposed sets were extracted according to the method described above. Then the remaining air-exposed sets were extracted every second day. Figure 2 shows the results obtained for three representative analytes: 1,3-DNB, 2,4-DNT, and TNT. Recoveries ranged from 85 to 105%, indicating that no breakthrough had occurred, and there was only slight degradation of the analytes on the cartridge within the storage time span tested. 3140

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Figure 2. Breakthrough and storage time profiles (spiking level, 10 ng; results expressed as percentage of the signal from extraction of non-air-exposed set) (9, 1,3-DNB; 2, 2,4-DNT; b, TNT) (n ) 3).

Table 2. Factors and Levels Tested (Coded Values in Parentheses) for SFE Screening and CCC Experimental Design

factor

low level (-1)

medium level (0)

Screening (Plackett-Burman) Design pressure (P, atm) 250 temp (T, °C) 40 static time (S, min) 0 dynamic mass of supercritical fluid (D, g) 5 supercritical fluid flow (F, mL/min) 0.5 restrictor temp (R, °C) 40

335 90 5 27.5 2.0 60

Response Surface Modeling (CCC) Design pressure (P, atm) 250 335 temp (T, °C) 40 90 dynamic mass of supercritical fluid (D, g) 5 27.5 static modifier (SM, toluene % v/v) 0 2.5 dynamic modifier (DM, MTBE % v/v) 0 2.5

high level (+1) 425 140 10 50 3.5 80 425 140 50 5 5

SFE Conditions. Since there are a large number of variables that could have a potential influence on the supercritical extraction efficiency, a multivariate optimization scheme is required. Here, a combination of a screening design to identify the most statistically significant factors followed by a response surface design to optimize the parameters was applied. Experiments in these designs involved extracting sets of “aged” spiked SPE cartridges. Cartridges were spiked by drawing through 5 mL of either HPLC-grade water (blank sets) or standard aqueous solution containing ∼1.2 ng/mL of each analyte using a vacuum device, thereby applying 6 ng of each analyte to the cartridge. Then the cartridges were dried under a gentle stream of nitrogen, wrapped in aluminum foil, and allowed to equilibrate for at least 4 days at -4 °C (the “aging” procedure) prior to analysis. In a first step, a Plackett-Burman34,35 experimental design including six experimental variables at two levels was performed, as shown in Table 2. The screening design included eight experiments plus three replicates at the central point. These experiments were performed twice. The response factor was defined as the percentage of signal (absolute area) obtained compared to that generated by a liquid injection of THF standard containing the same amount of the analytes. Figure 3 shows the plot of the coefficients for the response. The data in the figure suggest there were three significant factors: pressure (P), temperature (T) and mass of supercritical fluid in the dynamic extraction step (D) with positive effects, but these conclusions should be treated with caution, since interactions between factors are not included in this model. The other, nonsignificant variables were fixed at their medium values for the following experiments. Since the addition of modifiers to nonpolar CO2 can have a strong influence on extraction efficiency35,36 and its effect depends on several parameters, such as the identity of the modifier, mode of application, and concentration, an independent univariant study to select better combinations was carried out. Five different modifiers (acetonitrile, toluene, THF, MTBE, and methanol) were (34) Plackett, R. L.; Burman, J. P. Biometrika 1946, 33, 305-325. (35) Nerı´n, C.; Batlle, R.; Cacho, J. J. Chromatogr., A 1998, 795, 117-124. (36) Hollender, J.; Shneine, J.; Dott, W.; Heinzel, M.; Hagemann, H. W.; Go¨tz, G. K. E. J. Chromatogr., A 1997, 776, 233-243.

Figure 3. Scaled and centered coefficients for the screening Plackett-Burman designs for all factors under study. For factor identification, see Table 2.

evaluated. They were applied to the matrix in either a static mode, by adding 50 µL of modifier to a 1-mL extraction vessel, or in a dynamic mode, mixing 2 or 5% (v/v) of the modifier with the supercritical fluid. The best results (data not shown) in terms of average recovery were obtained by using 5% toluene as static modifier and 5% MTBE as the dynamic modifier. These findings can be explained as follows. The main role of a static modifier is to disrupt the analytematrix interactions, and it has to be remembered that toluene was the solvent of choice to develop the SPE procedure, as described above. On the other hand, the effects of a dynamic modifier are more related to changes in CO2 polarity, enhancing analyte solubility and, thus, extraction recovery. Nitroaromatic compounds exhibit high solubility in MTBE, so the combination of MTBE with supercritical CO2 would be expected to provide the best extracting phase, as the results proved. As stated above, a Plackett-Burman design does not count for interaction between individual factors. Furthermore, the effect of the organic modifier on the optimum values as well as the effect of using two different modifiers on the extraction performance should be evaluated. Thus, a response surface design (CCC) was constructed, to further assess the three factors derived from the previous screening investigation plus the two modifier-associated variables, within the experimental domain listed in Table 2. The design consisted of 26 randomized experiments, and six replicates in the central point were performed. Two replicates of the design were carried out, with the objective of maximizing the average recovery for all the compounds as compared to liquid standard injection. Since these experiments had to be carried out on three different days, the block effect was evaluated prior to analyzing the design and was found to be negligible at the 95% confidence level. Thus, performing the optimization runs over a number of days introduces no bias. Figure 4 shows the significant factors and their coefficients, as well as the descriptive values, R2 and Q2. The significance of all cross-interaction terms was extremely low, indicating that no interaction between variables occurs. The individual factors were all positively correlated with the response, but the change of sign in the quadratic coefficients P*P, D*D, SM*SM, and DM*DM indicates a curvature in the surface response. Furthermore, the absolute value of the D*D coefficient Analytical Chemistry, Vol. 75, No. 13, July 1, 2003

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Table 3. Optimum Values Found within the Experimental Domaina variable (units)

value a

P (atm)

T (°C)

density (g/mL)

S (min)

SM (% v/v)

D (g CO2)

DM (% v/v)

F (mL/min)

R (°C)

425

140

0.66

5

toluene, 5

35

MTBE, 5

2.0

60

Density, calculated from the optimized temperature (140 °C) and pressure (425 atm), has been added as an informative value.

Figure 4. Scaled and centered coefficients for significant factors in the response surface modeling CCC design. For factor identification, see Table 2.

Figure 5. Dynamic mass of supercritical fluid-temperature response surface for average recovery from SPE cartridge using SFE.

suggests that the optimum value for the dynamic mass of supercritical fluid could be found within the experimental domain. In fact, the response surface shown in Figure 5 demonstrates that the optimum within the experimental range corresponds to 35 g, which was the selected value. However, as can be concluded from the figure, masses as low as 10 g could be used to increase sample output while maintaining recoveries higher than 80%. Optimum values selected within the experimental domain are shown in Table 3. Figure 1b shows a typical LVI chromatogram from a spiked SPE cartridge extracted under optimum conditions. As shown in Figure 4, temperature was the most significant factor. The extraction of all nitroaromatic compounds was favored by higher temperatures. Following this, static and dynamic modifiers were the most significant factors. 3142

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It is of interest to consider more deeply the role of the static modifier. It could be postulated that the effect of the static modifier is not related to SFE per se, but rather, to liquid-solid extraction. To investigate this possibility, two sets (four replicates each) of samples were extracted. In the first set of cases, toluene was added to the vessel, allowed to impregnate the matrix for 10 min, and then extracted directly in dynamic mode. The second set was also impregnated for 10 min, then extraction was performed with an additional 5 min static extraction before the dynamic extraction. Results obtained with the second system (data not shown) were consistently 20 to 30% higher, as compared with the first set. Thus, the static modifier acts in a more efficient way if it is combined with a static extraction step before the dynamic extraction takes place. Because of the presence of supercritical fluid in the vessel, the interaction between the modifier and the matrix occurs in a supercritical medium. This speeds up the mass transfer of the analytes and increases the effectiveness of the extraction. Nevertheless, if this was the only effect, it should be possible to obtain the same recovery by allowing the modifier to interact with the matrix, that is, impregnate, for a longer period of time before exposure to the supercritical system. To check this point, two further sets of three experiments were performed, one with 20min impregnation, and a second with 45-min impregnation without static extraction. Results were improved by using 20 min impregnation instead of 10 min, but no further improvement was obtained when increasing contact time to 45 min, and recoveries were still 15 to 20% lower than those obtained when using a static extraction step. This can be explained by taking into account the fact that by filling the vessel with CO2, the system can also be regarded as a pressurized solvent extraction (PSE) basic setup unit. Therefore, the extraction efficiency is enhanced, since the solvent/matrix system is exposed to high temperatures and pressure.37 It is worth noting that even if the static extraction time is not a significant variable (as suggested by the data in Figure 3), it is necessary to test its effects with a static modifier, especially if the interactions between the analyte and the matrix are strong. Analytical Performance. To investigate the precision of the analytical method, SPE cartridges were spiked in quadruplicate with energetic compound standard solutions at three concentration levels ranging from 1 to 50 ng/cartridge. The experimental sets (including a blank set, prepared as explained above) were analyzed in randomized order for a period of 3 days. As would be expected, the precision decreased when the concentration of the analytes approached the method detection limit (MDL), resulting in the largest RSD values given in Table 4. (37) Ramos, L.; Kristenson, E. M.; Brinkman, U. A. Th. J. Chromatogr., A 2002, 975, 3-29.

Figure 6. Headspace sampling of military grade TNT. Peak identification: (1) 2,6-DNT, (2) 1,3-DNB, (5) 2,4-DNT, (8) TNT, and (9) TNB. Table 4. Method Performance precision (% RSD) 1 ng 25 ng 50 ng

analyte

method detection limit,a pg

2,6-DNT 1,3-DNB + 2,5-DNT 1,2-DNB 2,4-DNT 2,3-DNT 3,5-DNT TNT TNB

83.0 364 220 163 320 177 62.0 178

15.1 14.0 10.0 7.9 15.0 11.7 10.4 12.8

7.7 7.1 6.6 4.3 7.6 6.7 6.1 6.1

5.6 6.5 7.0 3.9 7.1 6.0 4.1 6.2

av

195

12.2

6.6

5.8

a

Expressed as picograms of analyte on cartridge to be extracted. Calculated on a 10-replicates basis.

To determine the MDL, 10 cartridges were spiked with 5 ng of each analyte, and MDLs were calculated as the product of the standard deviation of the 10 replicates and the two-tailed t-value for nine degrees of freedom at the 95% confidence level (2.26).38,39 The resulting values are shown in Table 4. These method detection limits compared well with corresponding figures reported in the literature,3,7,40 and could be improved by using longer sampling times. Finally, the method was applied to the analysis of real samples obtained from a desiccator containing 10 g of military grade (99% purity) TNT at 20 °C. Samples were collected in intervals between 1 and 12 min (3-min step) at a flow rate of 3.0 L/min. Figure 6 shows a LVI chromatogram obtained following 1-min sampling. TNT explosives contain small amounts of impurities formed during manufacture, where the symmetrical isomer, 2,4,6-TNT is the desired product. Different isomers of dinitrotoluene, DNT, as well as TNT isomers, are present in military grade TNT as a consequence of incomplete yield in the industrial process. These impurities can constitute up to 1% (w/w) of the final product. (38) Currie, L. A. Pure Appl. Chem. 1995, 67, 1699-1723. (39) Miller, J. C.; Miller, J. N. Statistics for Analytical Chemistry, 2nd. ed.; Ellis Horwood: Chichester, 1988; Appendix 2. (40) Albert, K. J.; Walt, D. R. Anal. Chem. 2000, 72, 1947-1955.

Moreover, 2,4- and 2,6-DNT exhibit higher vapor pressures (1.5 and 5.7 × 10-4 mmHg, respectively, as compared to 8 × 10-6 mmHg for TNT), indicating that the probability of detecting these impurities in the vapor phase of the explosive is higher than that of detecting TNT. As expected according to these volatility criteria explained above, 2,4-DNT constitutes the major component found in the headspace, but 2,6-DNT and TNT were also consistently detected in all the analyzed samples. Another typical group of byproducts consists of the benzene derivatives, dinitrobenzene(s), and trinitrobenzene. The origin of the trinitrobenzene can be also be attributed to photodecomposition of TNT.41 As can be seen in Figure 6, TNB and 1,3-DNB have also been detected in the analyzed samples. The high volatility of 1,3-DNB (9 × 10-4 mmHg, 20 °C) makes this compound one of the major constituents in the vapor phase above TNT. The presence of these compounds in TNT profiles has been previously reported.42,43 Moreover, their distribution, together with that of the DNT and TNT isomers, has been used successfully as a tool to characterize and identify the origin of different TNT samples.43 CONCLUSIONS In this paper, the successful hyphenation of three different techniques is described. The reported method shows promise for nitroaromatic vapor detection because of its high reproducibility, low method detection limit, and overall robustness. It can be applied in the simultaneous detection of several nitroaromatic congeners and could be used as a fingerprint characterization tool for unexploded ordnance. Furthermore, detection limits compared favorably to those reported earlier and can be lowered, if necessary, simply by increasing sampling time, because of the excellent retention capabilities of the SPE cartridges. (41) Yinon, J. Forensic and Environmental Detection of Explosives; John Wiley & Sons: New York, 1999. (42) Ravikrishna, R.; Yost, S. L.; Price, C. B.; Valsaraj, K. T.; Brannon, J. M.; Miyares, P. H. Environ. Toxicol. Chem. 2002, 21, 2020-2026. (43) Zhao, X.; Yinon, J. J. Chromatogr., A 2002, 946, 125-132.

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The use of SFE to desorb SPE cartridges reduces the amount of solvent necessary for the whole procedure to ∼250 µL of THF and eliminates the need of further cleaning or concentration steps. The use of PGC to retain the analytes suggests it would be possible to apply the method for extraction of more complex matrixes, such as soil or human fluids, because by combining SFE and selective desorption from PGC, clean and concentrated extracts have been obtained. To fully validate the method, it will be necessary to study the influence in the field of environmental factors such as interfering compounds, humidity, wind, and soil characteristics.

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ACKNOWLEDGMENT This work was supported by the Swedish Defense Research Agency (Tumba, Sweden) and the Swedish Rescue Service Agency (Karlstad, Sweden). The authors thank Luisa Pereira (Thermo Hypersyl, Cheshire, U.K.) for providing Hypercarb PGC material.

Received for review November 25, 2002. Accepted March 7, 2003. AC0207271