Anal Bioanal Chem (2009) 394:20192028 DOI 10.

1007/s00216-009-2689-9

ORIGINAL PAPER

Characterization of smokeless powders using nanoelectrospray ionization mass spectrometry (nESI-MS)

Gwynyth Scherperel & Gavin E. Reid & Ruth Waddell Smith

Received: 29 December 2008 / Revised: 4 February 2009 / Accepted: 6 February 2009 / Published online: 26 February 2009 # Springer-Verlag 2009

Abstract Smokeless powder is one of the most common types of explosives used in civilian ammunition and, hence, its detection and identification is of great forensic value. Based on comparison of physical properties, extraction yield in methanol, and the spectra obtained using nanoelectrospray ionization and multistage tandem mass spectrometry (MS/MS) in a quadrupole ion trap mass spectrometer, a method was developed to identify and differentiate unburned smokeless powders from different brands of ammunition. The mass spectrometry method was optimized for the simultaneous detection of the organic stabilizers commonly present in smokeless powders: methyl centralite, ethyl centralite, and diphenylamine. All but two of the powders were differentiated; however, the two that were not differentiated were produced by the same manufacturer. Gunshot residue from the cartridges was deposited on cotton cloth and collision-induced dissociation MS/MS was used to identify low levels of ethyl centralite in the residue, despite the presence of contaminants. Keywords Smokeless powder . Gunshot residue . Forensic analysis . Mass spectrometry
G. Scherperel : R. Waddell Smith Forensic Science Program, School of Criminal Justice, Michigan State University, East Lansing, MI 48824, USA G. E. Reid : R. Waddell Smith (*) Department of Chemistry, Michigan State University, East Lansing, MI 48824, USA e-mail: rwsmith@msu.edu G. E. Reid Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI 48824, USA

Introduction Smokeless powder, which is used as a propellant in ammunition cartridges, is commonly encountered in forensic laboratories either in the unburned form or in the burned form as a component of gunshot residue (GSR). Hence, the ability to rapidly identify and characterize smokeless powder samples and their presence in GSR is of great forensic value. The most common method for the forensic analysis of smokeless gunpowder or GSR is to identify inorganic components contained in the primer of the ammunition cartridge using scanning electron microscopy/ energy dispersive spectroscopy [1, 2]. However, with the relatively recent introduction of heavy-metal-free primers, there is a need for the development of a different approach, with recent research focusing on the analysis of organic components in the propellant [2, 3]. Smokeless powder can be widely classified as singlebased, double-based, or triple-based according to the major component(s) of the powder. Single-based powder consists of nitrocellulose, while double-based powder contains nitrocellulose and nitroglycerine, which is added as a plasticizer in order to soften the propellant and raise the energy content. Triple-based powder contains nitroguanidine in addition to nitrocellulose and nitroglycerine. Triplebased smokeless powders, however, are used primarily in rockets and large-caliber military-grade weapons and are therefore difficult to obtain on the open market [4, 5]. Smokeless powders also contain many additives, such as flash suppressants, opacifiers, plasticizers, and stabilizers [6]. Flash suppressants, such as nitroguanidine and dinitrotoluene, help prevent secondary flash at the muzzle of the firearm [1, 2]. Opacifiers, the most common of which is carbon black, enhance reproducibility of the burn rate. Plasticizers, such as dimethyl phthalate, diethyl phthalate,

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and dibutyl phthalate, are mixed with the smokeless powder grains in order to improve flexibility [2, 7]. However, it is stabilizers, which are added to slow the decomposition of nitrocellulose, that are considered to be the most unique organic components of smokeless powder [810]. The three most commonly used stabilizers are methyl centralite (MC, Scheme 1a), ethyl centralite (EC, Scheme 1b), and diphenylamine (DPA, Scheme 1c), while N-nitrosodiphenylamine (N-NO-DPA, Scheme 1d) is a decomposition product of DPA that can also be found in unburned smokeless powders [11]. The thermal instability of most explosives and the need for high sensitivity due to the low concentration of stabilizers in smokeless powders require the use of highly sensitive analytical techniques for their detection [10, 12 14]. Although mass spectrometry (MS) coupled with either gas chromatography (GC) or liquid chromatography (LC) offers the necessary sensitivity and selectivity [2, 12, 15 18], the use of chromatographic techniques can pose problems for the analysis of smokeless powders. GC can cause the thermal degradation of some compounds such as DPA, while LC is limited due to the potentially wide range of polarities of compounds that may be present in a powder. Several studies have demonstrated the successful identification of smokeless powders using MS without any chromatographic separation steps [9, 10]. For example, Wu et al. used a triple quadrupole mass spectrometer with electrospray ionization (ESI) and a flow-injection system to analyze MC standards as well as GSR extracted directly from shooters skin [10]. Direct sample infusion by ESI overcomes limitations associated with the use of chromatographic steps prior to MS analysis, while also minimizing the time required for sample processing prior to analysis. In another study by the same group, ESI coupled with a triple quadrupole mass analyzer was successfully used to analyze standard solutions of DPA and its nitrated derivates along with GSR extractions from skin [9]. These previous ESIMS studies, however, were limited in the number of stabilizers that were detected simultaneously.

A method is reported here for the detection and identification of organic stabilizers in smokeless powders using direct sample infusion by nanoelectrospray ionization (nESI) and multistage tandem mass spectrometry (MS/MS and MSn) analysis in a quadrupole ion trap mass spectrometer. The ability of the ion trap to perform multiple stages of dissociation (MSn) is shown to enable the identification and characterization of unknown components. Instrument parameters for the simultaneous detection of standard solutions of the organic stabilizers commonly found in smokeless powder (MC, EC, and DPA) were optimized. Then, these stabilizers were successfully detected in unburned smokeless powders, as well as in GSR deposited on cloth at distances of 3 and 12 in., using tandem mass spectrometry to enhance sensitivity and selectivity in the presence of contaminating species.

Experimental Materials Methanol (high-performance liquid chromatography grade), 1,3-dimethyl-1,3-diphenylurea (MC), 1,3-diethyl-1,3diphenylurea (EC), and DPA were purchased from SigmaAldrich (St. Louis, MO, USA). Deionized water was purified using a Barnstead nanopure diamond purification system (Dubuque, IA, USA). All reagents were used as supplied without further purification. Smokeless powder samples were obtained by removing the powder from the following cartridges: (1) 9 mm, 124 grain, full metal jacket (FMJ) produced by Federal; (2) 9 mm +P, 135 grain, jacketed hollow point (JHP) produced by Federal; (3) 0.45 automatic colt pistol (ACP), 230 grain, JHP produced by Federal; (4) 0.45 ACP, 230 grain, FMJ produced by Federal; (5) 0.40 Smith and Wesson (S&W), 155 grain, JHP produced by Federal; (6) 0.40 S&W, 155 grain, JHP (Silvertip) produced by Winchester; and (7) 0.357 Magnum, 125 grain, semi-jacketed hollow point produced by Remington-Peters. Cartridges 1, 3, 4, and 5 were provided by the Michigan State University Police Department, while cartridges 2, 6, and 7 were personally provided by a certified firearm instructor employed by the Michigan State University Police Department. All cartridges were stored in a cool dry location prior to use. Stereomicroscopy Smokeless powders from cartridges 17 were photographed using a Nikon SMZ800 stereomicroscope equipped with a Nikon digital DXM1200F camera and Nikon ACT-1 software, version 2.62 (all Nikon Corporation, Tokyo, Japan). Direct lighting (used for the powders from

Scheme 1. Structures of a methyl centralite, MC, b ethyl centralite, EC, c diphenylamine (DPA), and d N-nitrosodiphenylamine (N-NO-DPA)

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cartridges 2, 4, and 7) was supplied by a Schott Fostec light (Schott North America Inc., Elmsford, NY, USA), while oblique lighting (used for the powders from cartridges 1, 3, 5, and 6) was supplied by a standard light bulb. Images were adjusted for contrast, color, and brightness using Adobe PhotoShop, version 8.0 (Adobe Systems Incorporated, San Jose, CA, USA). Preparation of MC, EC, and DPA standards Stock solutions of MC, EC, and DPA were individually prepared in methanol at an initial concentration of 100 pmol/L and stored at 20 C. The stock solutions were then diluted in methanol to yield standard solutions of concentrations 0.000001, 0.00001, 0.0001, 0.001, 0.01, 0.1, 1, and 10 pmol/L. Extraction of organic stabilizers from smokeless powers Five milligrams of each smokeless powder from cartridges 1 7 were weighed out and crushed with the end of a spatula for 1 min. A 1 mL volume of methanol was added and the mixture was then vortexed for 10 s. The sample was then centrifuged; the methanol containing extracted organic material was removed and the powder was dried under vacuum. The powder was weighed before and after extraction in order to determine and compare extraction yields. Further dilution of the methanol extract (with methanol) was performed to obtain solutions with the desired concentration of extracted material (0.02, 0.5, and 0.1 mg/mL). All solutions were centrifuged prior to injection into the mass spectrometer. Preparation of GSR samples Gunshot residue samples were obtained by firing the aforementioned cartridges from the following handguns: (1) Glock Model 19, 9 mm, 4-in. barrel (for cartridges 1 and 2), (2) Sigarms Model P220, 0.45 ACP (for cartridges 3 and 4), (3) Sigarms Model P229, 4-in. barrel (for cartridges 5 and 6), and (4) Smith and Wesson Model 66, 2.5-in. barrel (for cartridge 7). To obtain GSR samples, target paper was fastened to a foam target backer and 1212-in. squares of cloth cut from mens short sleeve white t-shirts (Fruit of the Loom 100% cotton, size XXL, obtained from a local retail store) were pinned to the paper one at a time. Each cloth square was pinned to a different location on the target paper to avoid transfer of GSR from the paper to the cloth. The target paper was changed between each cartridge type and the firearms were extensively cleaned with MP Pro7 gun cleaner (n-methylpyrrolidone and diethylene glycol) between cartridges to minimize the risk of cross contamination between cartridges. Two cartridges of each type were

discharged at a firing distance of 3 in.: one into a square of unwashed cloth and one into a square of cloth that had been extensively washed with deionized water (to remove any potential contaminants) and air-dried. Cartridge 2 (9 mm +P) was also fired into a square of cloth that had been machinewashed and dried. After GSR was deposited, the cloth was placed between two pieces of wax paper, folded, and sealed in a plastic storage bag. This process was repeated at a firing distance of 12 in. for all cartridges. To extract GSR from the cloth for MS analysis, an approximately 1/4 inch by 1/4 inch square of cloth was cut from directly beside the bullet hole where visible gunshot residue was present. The sample cloth was then placed in 1 mL of methanol and vortexed for 30 s. No dilutions were made and all solutions were centrifuged prior to injection into the mass spectrometer. All samples were analyzed on the day of collection. nESI-MS, MS/MS, and MSn analysis All experiments were performed using a Thermo Scientific model LCQ quadrupole ion trap mass spectrometer (San Jose, CA, USA). Solutions were introduced at a flow rate of 1 L/min by nESI. The nESI conditions were optimized to maximize the intensity of the protonated precursor ion and to minimize the appearance of in-source fragmentation peaks. Typical nESI conditions were: spray voltage 2.5 kV, heated capillary temperature 125 C, capillary voltage 0 V, and tube lens voltage 0 V. All mass spectra were acquired using the normal resonance ejection scan mode. The precursor ion accumulation time for each scan was controlled by the automatic gain control function of the instrument in order to maintain an ion target number of 2 107 (arbitrary value). Collision-induced dissociation (CID) MS/MS and MS3 spectra were acquired at an activation q value of 0.25 using isolation widths of 310 Da, normalized collision energies of 1050%, and an activation time of 30 or 300 ms. The values were chosen such that the gentlest conditions were used in order to completely dissociate the selected precursor ion. Full and selected reaction monitoring scan types were used. The MS spectra shown are an average of 100 individual mass analysis scans while the MS/MS and MS3 product ion spectra shown are an average of 60 individual mass analysis scans.

Results and discussion Optimization of nESI-MS for detection and characterization of MC, EC, and DPA Typical nESI-MS conditions were optimized to maximize the intensity of the protonated MC, EC, and DPA ions,

2022 Fig. 1 nESI-MS and CID-MS/ MS of standard solutions of MC, EC, and DPA obtained under optimized conditions. a MS of MC (10 pmol/L), b MS of EC (10 pmol/L), c MS of DPA (100 pmol/L), d MS/MS of the [M+H]+ precursor ion of MC, e MS/MS of the [M+H]+ precursor ion of EC, and f MS/MS of the [M+H]+ precursor ion of DPA

G. Scherperel et al.

while minimizing the appearance of in-source fragmentation peaks. The potentials applied to most of the nESI-MS interface optics (i.e., entrance lens, interoctapole lens, multipoles 1 and 2, etc.) did not greatly affect precursor ion abundances, allowing the use of standard settings. However, the capillary voltage, tube lens voltage, and

heated capillary temperature were all found to have a significant effect on the abundance of the protonated precursor ions and the extent of in-source fragmentation that was observed. The capillary voltage was varied from 0 to 60 V; the tube lens voltage was varied from 0 to 250 V, and the capillary temperature was increased from

Scheme 2. Potential mechanisms for the multistage gas-phase fragmentation reactions of a protonated MC (m/z 241) to yield product ions at m/z 134 and 106, b protonated EC (m/z 269) to yield product ions at m/z 148, 120, and 92, and c protonated DPA (m/z 170) to yield a product ion at m/z 92

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100 to 325 C. At the higher voltages for both the capillary and the tube lens, in-source fragmentation became pronounced. At low capillary temperatures, the signal was lost, but, as temperature increased, in-source fragmentation occurred. Hence, for subsequent simultaneous detection of MC, EC, and DPA, the capillary and tube lens voltages were both maintained at 0 V while the heated capillary temperature was set at 125 C. Analysis of MC, EC, and DPA standards by MS and CIDMS/MS Mass spectra for each of the MC, EC, and DPA standard solutions, ranging in concentration from 0.000001 to 100 pmol/L, were acquired. Mass spectra collected for MC (10 pmol/L), EC (10 pmol/L), and DPA standard solutions (100 pmol/L) using the optimized MS parameters described above are shown in Fig. 1, indicating the presence of relatively abundant [M + H]+, as well as [M + Na] + and [M + K] + , precursor ions. Above concentrations of 1 pmol/L for the MC and EC samples, significant dimeric adducts were observed, while significant dimeric, trimeric, and quadrameric adducts were observed for the DPA sample above a concentration of 100 pmol/L.
Fig. 2 Stereomicroscopic images of seven smokeless powder samples. Powder (1) 9 mm, 124 grain, full metal jacket by Federal; (2) 9 mm +P, 135 grain, jacketed hollow point by Federal; (3) 0.45 automatic colt pistol, 230 grain, JHP by Federal; (4) 0.45 ACP, 230 grain, FMJ by Federal; (5) 0.40 Smith and Wesson, 155 grain, JHP by Federal; (6) 0.40 S&W, 155 grain, JHP (Silvertip) by Winchester; and (7) 0.357 Magnum, 125 grain, semijacketed hollow point by Remington-Peters

Since DPA did not ionize as efficiently as MC and EC, higher concentrations were necessary to achieve a comparable signal-to-noise (S/N) ratio. Due to the ubiquitous presence but variable concentration of Na and K in the samples or the solutions used for analysis, as well as the observation of multimeric adducts at higher concentrations, a linear calibration scale for the quantification of these species was not obtained. Although quantification using an internal standard or isotopic labeling is possible, from a forensic standpoint, quantification is not as important as identification of the stabilizers. Thus, qualitative analysis is sufficient to identify a powder or residue as a smokeless powder and to differentiate smokeless powders from different ammunition types. CID-MS/MS was then used to confirm the identity of the various ions observed in the spectra shown in Fig. 1. Dissociation of the [M + H]+ ion of MC in Fig. 1a (m/z 241) resulted in the formation of a dominant characteristic primary product ion at m/z 134 (Fig. 1d), via the neutral loss of N-methyl aniline. MS3 and MS4 (data not shown) resulted in the loss of CO to yield a secondary product ion at m/z 106. The fragmentation behavior of the protonated precursor ion of EC (m/z 269 in Fig. 1b) was similar to that for MC, consistent with the similarity in the structures of

2024 Table 1 Average extraction yields for seven smokeless powders based on percent organic material extracted Powder 1 2 3 4 5 6 7 Average extraction yield (%) 12.05 53.25 20.05 39.50 25.27 12.48 46.34

G. Scherperel et al. Relative standard deviation (%) 6.67 4.29 2.00 6.68 1.73 11.9 7.15

the two stabilizers. The fragmentation resulted in the formation of a characteristic primary product ion at m/z 148 via the neutral loss of N-ethyl aniline, a secondary product at m/z 120 via the loss of CO (confirmed by MS3), and a tertiary product at m/z 92 via the loss of CH2CH2 (confirmed by MS4; Fig. 1e). CID-MS/MS of the [M+H]+ precursor ion of DPA (m/z 170 in Fig. 1c) resulted in the formation of a primary product ion at m/z 92 via the neutral loss of benzene. A proposed mechanistic pathway for the formation of each of these ions is illustrated in Scheme 2. We note that Tong et al. previously reported the formation of an odd electron product ion at m/z 93 ([NC6H7]+) by

CID-MS/MS in an ESI-triple quadrupole mass spectrometer, rather than the ion at m/z 92 [9]. This discrepancy is likely due to the longer fragmentation time scale of the ion trap mass spectrometer compared to the triple quadrupole, which permits ion rearrangements and H+ transfers, such as would be needed to form the ion at m/z 92. Interestingly, Gilbert-Lopez et al. previously reported seeing both of these ions by in-source CID in an LC/ESI time-of-flight mass spectrometer, by applying different voltages to the fragmentor [19]. Stereomicroscopic examination Stereomicroscopic images of the seven smokeless powders investigated in this study are shown in Fig. 2. Powders 1 (9 mm), 3 (0.45 ACP, JHP), 5 (0.40 S&W, Federal), and 6 (0.40 S&W, Winchester) all have similar morphology: relatively round in shape with slightly rough surfaces and are gray-brown in color. Slight differences among the four

Fig. 3 nESI-MS analysis of methanol extracts (0.1 mg/mL) of a powder 1, b powder 6, c powder 3, d powder 5

Fig. 4 nESI-MS analysis of methanol extracts (0.1 mg/mL) of a powder 2, b powder 4, and c powder 7

Characterization of smokeless powders using nanoelectrospray ionization mass spectrometry (nESI-MS) Table 2 Summary of MS and MSn data for identified peaks in the smokeless powder samples Component Powder number Precursor ion (m/z) 269 291 307 509 537 559 575 602 606 628 CID-MS/MS product ions (m/z; % relative abundance) 120 (14%)

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CID-MS3 product ions (m/z; % relative abundance) 120 (100%) 148 (100%)

EC EC + Na EC + K ECa 2EC 2EC + Na 2EC + K ECa ECa EC + Na

a

1, 2, 3, 4, 5, 6, 7 2, 4, 7 4 2 1, 2, 4, 7 1, 2, 4, 7 4 3 1 1, 2, 4, 7

269 (100%) 269 (100%) 291 (100%) 307 (100%) 269 (100%) 269 (100%) 291 360 457 92 169 198 170 196 149 227 271 299 227 (100%) (82%) (32%) (100%) (100%) (100%) (75%) (100%) (28%) (10%) (100%) (46%) (100%)

120 148 120 148

(5%) (100%) (9%) (100%)

120 (10%) 148 (100%) 120 (8%) 148 (100%)

DPA N-NO-DPA 4-NDPA DPA-like componentb DPA-like componentb

1, 3, 5, 6 1, 5, 6 6 3 3, 5

170 199 215 227 317

65 (45%) 104 168 92 168 170 196 253 271 170 196 149 149 204 149 204 149 204 (100%) (100%) (100%) (100%) (68%) (100%) (40%) (100%) (70%) (100%) (100%) (8%) (100%) (10%) (100%) (14%) (100%)

DPA-like componentb DBP DBPc DBPc DBPc

3 1, 3, 5, 6 5, 6 1 1

453 279 448 575 585

149 (20%) 204 (100%) 279 (100%) 279 (100%) 279 (100%)

a b c

Unidentified EC adduct/complex Unidentified DPA-like component adduct/complex Unidentified DBP adduct/complex

powders were visually apparent. Powder 3 was rougher in texture while powders 1 and 6 were more brown in color than powders 3 and 5. Powders 1 and 6 had particle sizes ranging from approximately 300 to 600 m while powders 3 and 5 had a more uniform size distribution of approximately 600 m. Powders 2 (9 mm +P), 4 (0.45 ACP, FMJ), and 7 (0.357 Magnum) were all circular discs, approxi-

mately 600 m in diameter, and grainy in texture. Again, differences among these three powders were also clearly apparent: powders 2 and 4 were dark gray in color while powder 7 contained a mixture of black and light gray particles. Overall, powders 1, 3, 5, and 6 were visually different from powders 2, 4, and 7. However, powders 1, 3, 5, and 6 were

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sufficiently similar that differentiation among them based solely on physical properties was not possible. Only the surface roughness of powder 3 could potentially allow its distinction from powders 1, 5, and 6. Powders 2 and 4 were also too similar to be differentiated from each other based on physical appearance alone, but powder 7 was easily distinguished based on the presence of particles with two different colors. Hence, further chemical analysis was necessary in order to fully differentiate the seven smokeless powders. Extraction efficiency The average extraction yields (based on total organic components) determined upon triplicate extraction of each smokeless powder from cartridges 17 using the method described previously are detailed in Table 1. Similar to that observed by stereomicroscopic examination, the extraction yields could also be used to classify and differentiate the seven powders. Powders 1 and 6 were grouped based on the lowest concentration of extractable organic components in methanol (less than 15%) while powders 2, 4, and 7 were grouped based on the highest concentration of extractable organic materials (more than 40%). This latter grouping (powders 2, 4, and 7) is similar to the grouping defined previously based on physical appearance. In addition, further differentiation of the previous grouping of powders 1, 3, 5, and 6 was possible since powders 1 and 6 have a lower extraction yield in methanol than powders 3 and 5. nESI-MS analysis of smokeless powders For each of the seven smokeless powders, extracts were obtained as described above, then diluted further with methanol to yield solutions containing 0.1-mg/mL extracted material. Each solution was then analyzed by nESI-MS, using the conditions optimized previously. The resultant spectra for powders 1, 3, 5, and 6 are shown in Fig. 3, while the spectra for powders 2, 4, and 7 are shown in Fig. 4. Each of the major precursor ions observed in these spectra were isolated then subjected to further analysis by

CID-MS/MS and MS3 in order to confirm the presence of expected components within the extracts (e.g., MC, EC, and DPA), as well as to elucidate the identity of unknown species that were present. A summary of the main components in each powder and their product ions is given in Table 2. None of the powders analyzed were found to contain MC, while all contained EC. Powders 2, 4, and 7 contained EC alone while powders 1, 3, 5, and 6 also contained DPA. The presence of EC in powder 3 was only determined upon CID-MS/MS via detection of the characteristic m/z 148 product ion, due to the very low abundance of the precursor ion. Powders 1, 3, 5, and 6 also contained an abundant precursor ion at m/z 279. Upon MS/MS, this precursor ion yielded product ions at m/z 204 and 149, which is consistent with the expected fragmentation behavior of the plasticizer dibutyl phthalate. Powders 1, 5, and 6 each contained an ion at m/z 199 which fragmented to yield a product at m/z 167. MS3 of this product resulted in the formation of m/z 104 and 65, consistent with the expected fragmentation behavior of N-NO-DPA [9]. Powder 6 also contained an ion at m/z 215 which fragmented upon CIDMS/MS to yield a product ion m/z 198. Upon MS3, this product ion further fragmented to yield a product ion at m/z 168, consistent with the expected fragmentation behavior of 4-nitrodiphenylamine (4-NDPA) [9]. Both N-NO-DPA and 4-NDPA are nitrated derivates of DPA [4, 9, 20]. Although DPA has applications in the rubber and food industries, no nitrating agents or processes are involved in these industries such that the presence of nitrated derivatives of DPA are thought to be characteristic of smokeless powders [9, 20]. Powder 3 contained a dominant ion at m/z 227 which was not observed in the mass spectra of the remaining six powders. This ion fragmented to yield product ions at m/z 170 and m/z 196. The product ion at m/z 170 fragmented to yield an ion at m/z 92, similar to the fragmentation pathway seen for DPA, while the product ion at m/z 196 fragmented to yield m/z 168. This would suggest that the ion at m/z 227 is DPA like in structure due to the similar fragmentation pathway. While powder 5 did not contain the ion at m/z

Table 3 Main components detected in seven smokeless powder samples by nESI-MS (X indicates present) Powder 1 2 3 4 5 6 7 EC (m/z 269) X X X X X X X DPA (m/z 170) X X X X DBP (m/z 279) X X X X X X N-NO-DPA (m/z 199) X X X X 4-NDPA (m/z 215) DPA-like component (m/z 227)

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227, an ion at m/z 317 (also seen for powder 3) that fragmented to yield m/z 227 was present. The ion at m/z 227 dissociated to yield ions at m/z 170 (similar to the fragmentation pathway for DPA) and m/z 196, suggesting that the ion at m/z 317 is also DPA like in structure. A summary of the components present that were used to differentiate the smokeless powders is given in Table 3. Thus, based on physical characteristics, extraction efficiency in methanol, and mass spectra, it was possible to differentiate the seven powders, with the exception of powders 2 and 4. These two powders were different brands produced by the same manufacturer and, hence, it is feasible that powders 2 and 4 are in fact the same smokeless powder. Analysis of GSR by nESI-MS and CID-MS/MS Each of the smokeless powders was fired into squares of cotton cloth (unwashed and washed with deionized water and air-dried) at distances of 3 and 12 in. In addition, cartridge #2 (9 mm +P) was also fired into a square of cloth that had been machine-washed and dried. The cloth samples were extracted in methanol then analyzed by nESI-MS using the optimized conditions described above. Examples of spectra obtained for powder 2 (contains EC only) are shown in Figs. 5 and 6 for firing distances of 3 and 12 in., respectively. For unwashed cloth (Figs. 5a and 6a), ions observed at higher m/z were attributed to background. For both spectra, however, the most abundant ion corresponded to the dimeric [2EC + Na]+ precursor ion at m/z 559. As the firing distance increased from 3 to 12 in., less GSR was deposited. Hence,

Fig. 5 Mass spectra of GSR from Powder 2 at a a firing distance of 3 in. on unwashed cloth, b a firing distance of 3 in. on cloth washed with deionized water, and c a firing distance of 3 in. on machine washed cloth

Fig. 6 Mass spectra of GSR from powder 2 at a a firing distance of 12 in. on unwashed cloth, b a firing distance of 12 in. on cloth washed with deionized water, and c a firing distance of 12 in. on machine washed cloth

Fig. 7 Comparison of mass spectra for unwashed cloth with and without gunshot residue. a Mass spectrum of unwashed cloth (blank). The inset to panel a shows the product ion spectrum obtained by MS/ MS of m/z 269 from this sample. b Mass spectrum of gunshot residue from powder 1 on unwashed cloth (firing distance of 12 in.). The inset to panel b shows the product ion spectrum obtained by MS/MS of m/z 269 from this sample

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the abundance of the background ions was similar in both spectra and only the abundance of the EC peaks varied. Figs. 5b and 6b show the spectra for GSR obtained from powder 2 at firing distances of 3 and 12 in., on cloth washed with deionized water. After washing, the ions attributed to background were largely eliminated (Figs. 5b and 6b). With the background ions eliminated, the most abundant peaks in the spectrum at 3 in. were from EC. No additional ions were observed when compared to the spectrum for the unburned powder 2 (Fig. 4a). At 12 in., ions for EC were observed easily, but their abundance was, as expected, significantly lower than at 3 in. such that background ions were observed at higher relative abundance. For machine-washed and dried cloth, the spectra showed background ions that were most likely due to the presence of detergent (Figs. 5c and 6c). At 3 in. (Fig. 5c), EC ions were distinguishable among the other background and detergent ion, but, at 12 in. (Fig. 6c), any EC ions that may be present were below the level of noise and thus were not detectable. The utility of performing CID-MS/MS to improve the limit of detection for the identification of low abundance EC in GSR was demonstrated by analysis of the gunshot residue obtained from powder #1. The mass spectra obtained from unwashed cloth without gunshot residue (blank) and with gunshot residue fired from a distance of 12 in. are shown in Fig. 7a, b, respectively. Both spectra showed abundant detergent ions, the most abundant of which range from m/z 463 to m/z 777, but neither spectra showed an ion at m/z 269 (protonated EC) above the level of noise in the spectra. Upon performing CID-MS/MS at m/ z 269 from the blank sample, only product ions at m/z 187 and 239 (inset to Fig. 7a) were observed, presumably originating from an m/z 269 contaminant in the cloth. The absence of the characteristic product ions of EC at m/z 120 and 148 therefore indicated that the blank did not contain EC. In contrast, the characteristic m/z 120 and 148 product ions were readily observed upon MS/MS of m/z 269 from the cloth containing gunshot residue, in addition to the contaminant product ions at m/z 187 and 239 (inset to Fig. 7b), confirming the presence of EC, albeit at low concentration. This result clearly exemplifies the value of MS/MS and the significantly enhanced sensitivity and specificity associated with the characteristic transition from m/z 269 to 148 upon MS/MS or from 148 to 120 upon MS3 for the detection of EC, even when in the presence of contaminant ions at the same m/z value.

Conclusions nESI-MS was applied for the differentiation of seven smokeless powders in the unburned form. All three organic stabilizers (MC, EC, and DPA), which are considered characteristic of GSR, can be detected simultaneously using this method. Of the seven powders investigated, all but two could be fully distinguished based on a combination of physical properties, extraction yield in methanol, and nESIMS. The two powders that could not be distinguished were different brands from the same manufacturer; hence, it is conceivable that both were the same smokeless powder. For burned powders, CID-MS/MS was used to identify low levels of EC in GSR, despite the presence of contaminants. These results indicate that, with further development and validation, nESI-MS has great potential for extensive applications in the forensic analysis of GSR.
Acknowledgements The authors would like to thank the certified firearms instructor who provided all ammunition and firearms used and who provided us with an unending supply of gunshot residue.

References
1. Schwoeble JA, Exline DL (2000) Current methods in forensic gunshot residue analysis. CRC Press, Boca Raton 2. Meng H, Caddy B (1997) J Forensic Sci 42:553 3. MacCrehan WM, Patierno ER, Duewer DL, Reardon MR (2001) J Forensic Sci 46:57 4. Andrasko J (1992) J Forensic Sci 37:1030 5. Yinon J, Zitrin S (1993) Modern methods and applications in analysis of explosives. Wiley, Chichester 6. Yinon J (1999) Forensic and environmental detection of explosives. Wiley, Chichester 7. Zeichner A (2003) Anal Bioanal Chem 376:1178 8. Mach MH, Pallos A, Jones PF (1978) J Forensic Sci 23:433 9. Tong Y, Wu Z, Yang C et al (2001) The Analyst 126:480 10. Wu Z, Tong Y, Yu J et al (1999) The Analyst 124:1563 11. Reardon MR, MacCrehan WA, Rowe WF (2000) J Forensic Sci 45:1232 12. Berberich DW, Yost RA, Fetterolf DD (1988) J Forensic Sci 33:946 13. King RM (1993) In: Yinon J (ed) Advances in the analysis and detection of explosives. Kluwer Academic, Boston 14. Wu Z, Tong Y, Yu J et al (2001) J Forensic Sci 46:495 15. Druet L, Angers J (1988) Propellants Explos Pyrotech 13:87 16. MacCrehan WA, Bedner M (2006) Forensic Sci Int 163:119 17. MacCrehan WA, Reardon MR (2002) J Forensic Sci 47:996 18. Tamiri T, Zitrin S (1986) J Energetic Mat 4:215 19. Lopez-Gilbert B, Garcia-Reyes JF, Ortega-Barrales P et al (2007) Rapid Commun Mass Spectrom 21:2059 20. Espinoza EON, Thornton JI (1994) Anal Chim Acta 288:57