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The author(s) shown below used Federal funds provided by the U.S.Department of Justice and prepared the following final report:Document Title:Forensic Drug Identification by GasChromatography – Infrared SpectroscopyAuthor(s):Robert Shipman, Trisha Conti, Tara Tighe, EricBuelDocument No.:242698Date Received:June 2013Award Number:2008-DN-BX-K161This report has not been published by the U.S. Department of Justice.To provide better customer service, NCJRS has made this Federallyfunded grant report available electronically.Opinions or points of view expressed are thoseof the author(s) and do not necessarily reflectthe official position or policies of the U.S.Department of Justice.

Research and Development in the Area of Controlled SubstancesForensic Drug Identification by Gas Chromatography- Infrared SpectroscopyAward Number: 2008-DN-BX-K161Robert Shipman, Trisha Conti, PhD, Tara Tighe, MS and Eric Buel, PhDABSTRACTThe primary goal of the forensic drug examiner is the unequivocal identification of anycontrolled substance present in a drug exhibit. Most forensic laboratories routinely employ GasChromatography/ Mass Spectrometry (GC/MS) as the preferred method for this examination.The technique provides a rapid, semi-automated analysis of the sample and typically yieldssufficient information to identify the compounds in question. However, the application ofGC/MS for drug analysis does have its limitations.Certain drugs yield minimal mass spectral fragmentation patterns using electron impact MS,while other compounds, such as some diastereomers and positional isomers, are not readilydifferentiated by mass spectroscopy. Infrared spectroscopy (IR, meaning FTIR) provides analternate technique to mass spectroscopy for the identification of organic compounds. Recentimprovements in the hyphenated technique, Gas Chromatography/Infrared Spectroscopy(GC/IR) may provide a simple alternative or supplemental approach to GC/MS for theidentification of certain compounds. A newly introduced instrument collects GC effluent on aliquid nitrogen cooled, IR transparent window that allows the direct analysis of the depositedsolid material. This technique is superior to the IR light pipe in sensitivity, IR spectral quality,and allows direct comparison of the collected spectra to existing IR databases. Our research1This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

developed procedures and protocols for the analysis of drugs and determined the benefits andlimitations of this technology. This research focused on the routine identification of commonlyencountered drugs, designer drugs, closely related drug isomers, as well as the fundamentals ofthe gas chromatography and infrared systems. Statement of purpose: The research wasundertaken to develop this technology into a viable technique for the forensic community.The instrument was studied for repeatability, sensitivity, and selectivity while optimizing foranalysis of a wide range of drug samples. Based upon this work the instrument proved to be apowerful forensic tool providing complimentary data to GC/MS.Acceptable levels ofsensitivity, linearity, and reproducibility were achieved using the GC split-less injection mode.Concern about cross contamination of samples on the collection disk were dispelled as thedeposited GC vapor produced solid “tracks” that were unique to each sample and appropriatelydocumented by the instrument. Analytical methods were developed for the routine analyses ofdrugs and synthetic cannabinoids.Through these studies the instrument was verified forcasework analysis and is presently in operational use in our laboratory.Software limitations hindered research progress, although software and hardware upgradeswere made by the vendor (Spectra Analysis) some of which were driven by feedback providedby staff at the Vermont Forensic Laboratory (VFL).2This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

TABLE OF CONTENTSITEMPAGE(S)ABSTRACT1-2EXECUTIVE SUMMARY4-10MAIN BODY11-62I. Introduction11-18II. Methods19-25III. Results26-37IV. Conclusions38-40V. References41-42VI. Dissemination of Research Findings43-44VII. Appendix 1 Tables45-49VIII. Appendix 2 Figures50-623This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

EXECUTIVE SUMMARYProblemForensic Scientists are required to identify an ever increasing and more complex assortment ofdrugs and related compounds. A particular problem is the increased submission of designer orsynthetic drugs. A variety of compounds appear “on the street” which are designed to avoidexisting laws by making slight modifications to the structure of the controlled substance. Thesedrugs were once an occasional problem, today they have become much more common. Due tothe structural similarity of the specimens encountered by the forensic laboratory, an array ofinstruments is needed to correctly identify these substances. A new GC/IR instrument has beendeveloped which can aid in the forensic analysis of drug samples. An instrument that canprovide additional data to distinguish closely related compounds could be an asset to thecommunity.Samples submitted to the lab may contain complex mixtures of drugs and other compounds. In2011/2012, the Federal and Vermont State governments added many synthetic compounds totemporary schedules or regulated lists (see 17, 18 for recent regulations). These includedcathinones (“bath-salts”), cannabinoids(“K2/Spice”), and additional 2C compounds (2,5dimethoxy phenethylamines) compounds. Identification of these chemicals in combination withother compounds created unexpected demands on Forensic labs, including the Vermont ForensicLab. The final part of the research grant focused on these new synthetic compounds which werebeing submitted to the lab as casework.4This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

PurposeThe purpose of this research was to determine the feasibility of a new type of GC/IR instrumentto aid in the analysis of samples for suspected drugs. Infrared analysis is already a powerfulanalytic tool utilized in most forensic laboratories. Coupling IR detection with a separationtechnique would provide a valuable instrument to forensic labs. This research was undertakenby the VFL to access the assets and limitations of the Spectra Analysis DiscovIR-GC system.Research designOur laboratory received this grant to further study the limitations and benefits of this newinstrument for drug analysis. Feedback to the vendor helped create useful software and hardwareupgrades. The VFL developed protocols to enable successful testing by GC/IR of samples andstandards, and optimized the system for routine casework analysis. Several IR spectral librarieswere used: commercial (available through the vendor), vendor generated and in-house generated.FindingsPreliminary work at the VFL on the GC/IR system (before obtaining the grant) revealed a toolwith a large potential. We examined several drug types which are hard to differentiate usingGC/MS. Diastereoisomers are challenging to analyze. Ephedrine and pseudoephedrine may beresolved by GC (figure 1), but their mass spectra is not determinative (figure 2). The GC/IRspectra for these two compounds are shown in figure 3. Differences in the infrared fingerprintregion allow identification of these diastereoisomers. This research was conducted to determineif the GC/IR could be used as a cutting edge forensic tool to identify complex drug samples.5This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

The working concentration range was determined for two representative drugs, pseudoephedrineand cocaine. The VFL desired the concentration range to be similar to that employed for GC/MSanalysis of drugs. This would allow the transfer of sample vials between instruments withoutadditional dilutions or extractions. The practical limit of detection (PLOD) for these compoundsby GC/IR was 25-50 parts per million (ppm) (weight/volume), which is similar in concentrationto what is routinely used for GC/MS work. Saturation and overloading of the GC/IR instrumentoccurred near the 1000 ppm level for the two compounds.Our work did not includedetermination of saturation or an overload point for the GC/MS. However from running thesame sample extracts on both systems, it is apparent that the GC/IR will show detector saturationat lower concentration levels compared to that observed with the GC/MS. This may be due to awider linear range for the MS, a greater IR sensitivity for some compounds, and/or the use ofsplit injection by the GC/MS system.A split/split-less injection study was undertaken for the GC/IR. The split-less mode was shownto have an increase in sensitivity, and produce acceptable resolution and peak symmetry for thetest synthetic cannabinoid compound JWH-122. Other drugs (heroin, cocaine, and ethcathinone)were shown to have acceptable resolution and peak symmetry using the normal drug scanconditions.Acceptable resolution of a JWH isomer mix (composed of a mix of ortho, meta and paraisomers) was demonstrated using the synthetic cannabinoid drug conditions.Retention times were highly reproducible as ten injections of cocaine and pseudoephedrine hadlow % relative standard deviation (RSD) values when comparing the retention times.It is well known that different crystalline states of a compound will affect the IR spectraobtained from that compound. We noticed differences between the IR spectra for a number of6This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

compounds obtained via the GC/IR to those derived from other instruments and IR libraries. Astudy was conducted to determine if the GC vapor deposited upon the disk yielded an amorphousor crystalline solid, compared to the known solid forms via bench IR preparations. Six drugcompound solutions were tested to determine if the condensate from the GC yielded amorphousor crystalline deposits. Portions of the solutions were dried and produced a solid for analysis onour Perkin Elmer (PE) IR instrument using microscope and attenuated total reflectance (ATR)sampling. The same solutions were run on the GC/IR instrument and compared to the dataobtained from the PE instrument. For the GC/IR analysis, two compounds (cocaine base andpseudoephedrine) appeared to yield an amorphous deposition. Three compounds (amitriptyline,diphenhydramine, and ephedrine) appeared to form crystalline structures, while one compound(3,4-MDMA) the structure was inconclusive. This was based upon comparison of the IR spectraobtained between the bench PE instrument and the GC/IR. There were some concerns if thisamorphous/crystalline structure was reproducible for a given compound.To test for structure reproducibility on the GC/IR, cocaine and pseudoephedrine were analyzedten times and the resulting spectra were compared. This comparison revealed very similar spectraat these given collection conditions, indicating that the nature of the deposition did not changefrom run to run.We also studied the effect of disk temperature on solid formation. This study was inconclusivefor both the VFL and the manufacturer. The vendor stated that actual disk temperature wasdifficult to measure.Another study based upon the amount of drug deposited upon the disk was conducted to assessthe nature of the deposition. Of concern is if the “form” of the deposited material could beconcentration dependent. A study varying the concentration of pseudoephedrine and cocaine7This document is a research report submitted to the U.S. Department of Justice. This report has notbeen published by the Department. Opinions or points of view expressed are those of the author(s)and do not necessarily reflect the official position or policies of the U.S. Department of Justice.

injected appeared to produce no changes in IR spectra; and hence we conclude no difference inthe nature of the deposited material as the amount of material deposited upon the disk isincreased.The salt form of the injected drug did not appear to effect the formation of crystalline oramorphous solids. Some salts of compounds do affect GC performance, in particular retentiontime and GC resolution (this was most pronounced for amphetamine type compounds). Note:drugs that are injected as the salt form (for instance the hydrochloride of cocaine) are eluted asthe base form of the drug and must be compared to a base form for any library or comparativeexamination. Laws that require determination of the salt form of cocaine must be performedusing additional testing to confirm the form of cocaine.The research then f