Analysis of TNT, DNA Methylation, and Hair Pigmentation Via Gas Chromatography-Mass Spectrometry and Spectroscopic Techniques

Jacqueline Ruchti, Purdue University

Abstract

Gas chromatography-mass spectrometry (GC-MS) is a commonly used analytical technique that efficiently separates and identifies various volatile organic compounds. Here GCMS was used with new methodologies for detecting 2,4,6-trinitrotoluene (TNT), deoxyribonucleic acid (DNA) methylation, and hair melanins. Spectroscopic techniques including FourierTransform Infrared Spectroscopy (FTIR), Microspectrophotometry (MSP), and Raman Spectroscopy were also used for detecting hair melanins. These methodologies would determine the following: 1) if TNT could be simultaneously collected and extracted with DNA; 2) if DNA methylation could determine the aging of DNA in blowflies; and 3) if an objective metric for hair pigmentation can be established. For the first study, it was hypothesized that simultaneous analysis of post-blast debris for DNA and explosive compounds could improve explosives investigations. Hence, DNA and explosive residues would need to be separated via an extraction method. In preliminary work, GCMS analysis via immersion solid phase microextraction (SPME) was optimized to directly analyze Microbial-Vac System (M-Vac) buffer extracts for explosives. In this work, the PCIA extraction method was then applied to liquid samples rather than the traditional swab samplings. To maintain good layer partitioning and TNT peak resolution, the volume ratio of PCIA to Lysis Buffer to chloroform solution of TNT was set to 1:1:1. However, PCIA extraction was later deemed unnecessary for the chemical extraction of TNT. Instead, simpler methods of liquid-liquid extraction, charcoal strip extraction, and filtration were devised for extracting TNT prior to traditional DNA extraction methods. The partitioning of TNT, a common military explosive, was studied for the following four aqueous/organic solutions: water/chloroform, Lysis Buffer/chloroform, Lysis Buffer with proteinase K (incubated)/chloroform, and Butterfield’s Buffer/chloroform. A liquid injection method was developed for the chloroform layer. An immersion SPME method was developed for the aqueous layer. Since the Log P value for TNT in an octanol/water solution is 1.6, TNT was expected to partition by approximately 40:1 between octanol and water. However, the partitioning of TNT between Chloroform and Water was inconclusive. TNT partitioned by approximately 3:5 between Chloroform and Lysis Buffer, 1:2 between Chloroform and Lysis Buffer with proteinase K (incubated), and 35:2 between Chloroform and Butterfield’s Buffer. Due to the ability of the M-Vac to collect DNA from rough and porous surfaces, the device was tested for its effectiveness in also collecting TNT. Evidence collected via the M-Vac device was analyzed by GC-MS after extraction. Samples prepared via the chloroform partition method or charcoal strip extraction method were run via liquid injection while samples prepared via the filtration method were run via liquid immersion SPME. Following the application of a 50 ppm chloroform solution of TNT to pieces of a white t-shirt, TNT was successfully detected via the chloroform partition method. To simulate samples from an explosives investigation, a backpack containing a pipe bomb filled with TNT was detonated in a secured field. However, TNT was not successfully detected in any post-blast backpack debris samples prepared via the three extraction methods. Upon performing a direct liquid extraction with other post-blast backpack debris samples, it was determined that the amount of TNT on the post-blast backpack debris was too low. Hence, the M-Vac’s efficiency for collecting TNT from post-blast debris is inconclusive. Therefore, other M-Vac buffer solutions, concealment materials, and explosive compounds should be explored.

Degree

M.Sc.

Advisors

Goodpaster, Purdue University.

Subject Area

Analytical chemistry|Biochemistry|Chemistry|Materials science|Organic chemistry

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