Forensic investigators analyze gunshot and explosive residues not only to identify persons involved in crimes but also for intelligence purposes (i.e., in analyzing the methods of terrorist and criminal organizations).
Two types of component are important for this purpose: inorganic constituents and organic residues. However, despite a great deal of effort worldwide, using the most sophisticated (and expensive) instrumentation, including neutron activation analysis, mass spectrometry, X-ray and infrared techniques, and a variety of chromatographic techniques, problems persist in the development of methods for routine applications.
HPCE offers the unique possibility of analyzing both organic and inorganic compounds because the separation mechanisms are unique and because a minimal amount of sample is needed for analysis. In an early paper (if not the first) reporting the use of HPCE in this field, Northrop et al. (1991) discussed the performance of MEKC in the separation and determination of some of the constituents of gunshot and explosive materials. Their results demonstrated the possibilities of HPCE for identification of residues. Un-coated silica capillaries 67 cm long with i.d. ranging from 50 to 150 mm were used in experiments aimed at optimizing the separation and at studying the effect of different conditions. The buffer was 2.5 or 5.0 mM in borate, in the pH range 7.8-8.9, and contained SDS concentrations from 10 to 50 mM. The applied potential was 20 kV for the separation, but 5 kV, for 2 seconds, for electrokinetic injections. Detection was by U V absorption. After optimization, the authors decided on 100 mm i.d., 2.5 mM borate, 25 mM SDS, and a detection wavelength of 250 nm (200 nm for nitroglycerin). As many as 11 components of gunshot residues (test mixture), namely, nitroguanidine, nitroglycerin, 2,4-dinitrotoluene (DNT), 2,6-DNT, 3,4-DNT, 2,3-DNT, diphenyl-amine (DPA), /V-nitroso-DPA, 2-nitro-DPA, ethylcentralite, and dibutyl phthalate were all resolved in 10 minutes, with efficiencies between 200,000 and 400,000 theoretical plates and mass detection limits of fractions of nanograms. The results from HPCE were reportedly superior to those from supercritical fluid chromatography.
The same MEKC system was also used for the separation of a test mixture of 15 compounds of interest in high explosive analysis, including nitroguanidine, ethylene glycol dinitrate, diethylene glycol dinitrate, l,3,5-trinitro-l,3,5-triazacyclohexane (RDX), nitroglycerin, 2,4,6-trinitrotoluene (TNT), penta-erythritol tetranitrate, and picric acid, with excellent resolution except for the overlapping of 1,5- and 1,8-isomers of dinitronaphthalene. Also, separation of all the components (26) of the two sets of standards was attempted with extremely limited coelutions.
In addition, since many compounds have characteristic UV absorption profiles, multiwavelength analysis helped their identification. Because the instrument used was not capable of peak spectra acquisition, these authors had to perform multiple runs of the same sample at different wavelengths and draw absorbance profiles from the comparison of the peak heights in the individual electropherograms. More modern instrumentation fitted with diode array or fast-scanning UV detectors would facilitate analyses of this type.
Application to forensic cases concerned the investigation of spent ammunition casings, reloading powders (IMR 700X, W-W 452AA, HRD, W-W 296,
W-W 748, IMR 4831), and plastic explosives (Semtex, C4, Detasheet, Tovex). Compositional differences among reloading powders were found in products from different manufacturers, but also between powders from the same producers. The reasons for these differences were discussed by the authors.
In a subsequent paper, the same authors (Northrop and MacCrehan, 1992) optimized the sample collection and preparation procedures for the MEKC analysis of gunshot residues. Instead of the widely used swabbing techniques, the authors suggested sample collection with masking adhesive tape (1-inch-square sections). The sample area of interest was, as usual in case of users of revolvers and pistols, the back of the shooting hand along the thumb and forefinger and the webbing between these fingers. The tape "lifts" were stored sealed and refrigerated. Each lift was examined with a binocular stereoscope for gunshot residue particles, which were collected with tweezers and placed in a glass microvial. The particles were added with 50 mL of ethanol and extracted for 30 minutes in an ultrasonic bath. Ethylene glycol (1 mL) was then added to the vial, the solvent was evaporated under a nitrogen stream, and finally the residue was reconstituted with 25 mL of MEKC buffer. Alternatively, sections of the tape lifts were extracted with ethanol, observing the procedure just described.
According to the authors, adhesive tape lifts were clearly superior to swabbing, which was discarded because of unacceptable losses of analytes and high recovery of interferences from the skin. Adhesive tape lifts provided a much cleaner material, without interfering compounds (fats and oils) from the skin. Because of the small sample size required for HPCE, a single particle collected from the skin surface could be subjected to analysis. However, the quality of the adhesive tape was fundamental for the success of the assay; a test of several tapes comparing adhesive character, resistance to extraction solvents, and lack of coeluting interferent peaks showed that masking tape was the most suitable for MEKC analysis.
Another problem the authors addressed was the loss of analytes during evaporation of the extraction solvent. For this purpose, the addition of a small percentage of ethylene glycol, a nonvolatile "keeper," according to the authors, prevented the sample from going to dryness and also avoided adsorption losses. An additional advantage was that ethylene glycol, with a relatively high viscosity, acted to equalize the slight viscosity differences between standard solutions and real samples, which produced some nonrepro-ducibility in migration times. Quantitative results were improved by using an internal standard (/3-naphthol). In firing range experiments with subjects who fired different weapons, characteristic gunshot residue constituents were found on each adhesive tape lift on which they were expected to be present, but not on blanks.
HPCE was also applied in the analysis of low explosive ionic residues and compared to ion chromatography (Hargadon and McCord, 1992). Anions and cations left behind from the blast represent useful pieces of evidence from which to infer the type and source of the explosive material used. Although ion chromatography is the ideal tool for this purpose, it suffers from the lack of an adequate complementary technique for peak confirmation. Again, HPCE, which is based on completely different separation and detection principles, gives a unique opportunity to confirm ion analysis results.
To this aim, Hargadon and McCord used CZE (65 cm X 75 mm i.d. capillary) in borate buffer (2 m M borate, 40 m Ai boric acid) containing 1 m M diethylenetriamine as EOF modifier, at the final pH of 7.8. The applied potential was 20 kV, with reversed polarity. The addition to the running buffer of a dichromate chromophore (1.8 mM) permitted the use of indirect UV detection at 280 nm.
The most important ions present at the blast residue are chloride, nitrite, nitrate, sulfate, sulfide, chlorate, carbonate, hydrogen carbonate, cyanate, thio-cyanate, and perchlorate, which are susceptible of CZE determination in a single run. A comparison CZE with ion chromatography showed extensive differences between the relative separation patterns, with a clear advantage of CZE in efficiency and of ion chromatography as far as capacity is concerned. The reproducibility of the elution times in CZE was evaluated over 2 months with the same capillary, with resulting RSD better than 1%, which could be improved by adding bromide as a marker for calculating relative migration times.
It was also observed that peak identity could be checked by comparing the electropherograms recorded at 280 nm with other electrophoretic profiles recorded at 205 nm, a wavelength at which nitrite, nitrate, and thiocyanate absorb, generating positive peaks. Anions that do not absorb also at this wavelength produce negative peaks as at 280 nm. The sensitivity of CZE, with a signal-to-noise ratio of 3, was 0.5 mg/mL, whereas for ion chromatography it was 2 mg/mL; the dynamic range was up to 50 mg/mL for CZE and up to 200 mg/mL for ion chromatography.
The practical consequences of these studies was the discovery that solutions suitable for ion chromatography must be diluted before CZE. This analytical strategy was tested with pipe bomb fragments experimentally prepared with different explosive mixtures (potassium chlorate-Vaseline, black powder, smokeless powder, a mixture of black and smokeless powders) and detonated. Fragments from bombs were extracted with water, filtered, and analyzed. Dual-ion chromatography and CZE analysis were carried out with concordant results.
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