Characterization and differentiation of cyclopropylfentanyl from E-crotonylfentanyl, Z-crotonylfentanyl, and 3-butenylfentanyl.

Recently, a sample containing cyclopropylfentanyl was analyzed at this laboratory. Cyclopropylfentanyl began to appear in the United States' illicit drug markets in 2017. Unfortunately, cyclopropylfentanyl presents an analytical challenge due to its mass spectrum being almost identical to that of crotonylfentanyl. There are two possible isomers of crotonylfentanyl, Z- and E- crotonylfentanyl. In order to provide sufficient analytical data to distinguish the two isomers of crotonylfentanyl and cyclopropylfentanyl, crotonylfentanyl was synthesized and fully characterized. Each isomer was analyzed via nuclear magnetic resonance spectroscopy, gas chromatography-mass spectrometry, and Fourier transform infrared spectroscopy. During the synthesis of crotonylfentanyl, an unknown compound was formed. The identification of this compound and the analytical characterization of the two isomers of crotonylfentanyl are presented. Through the comparison of these compounds, it was confirmed that cyclopropylfentanyl can be differentiated from crotonylfentanyl.

Applying the principles of isotope analysis in plant and animal ecology to forensic science in the Americas.

The heart of forensic science is application of the scientific method and analytical approaches to answer questions central to solving a crime: Who, What, When, Where, and How. Forensic practitioners use fundamentals of chemistry and physics to examine evidence and infer its origin. In this regard, ecological researchers have had a significant impact on forensic science through the development and application of a specialized measurement technique-isotope analysis-for examining evidence. Here, we review the utility of isotope analysis in forensic settings from an ecological perspective, concentrating on work from the Americas completed within the last three decades. Our primary focus is on combining plant and animal physiological models with isotope analyses for source inference. Examples of the forensic application of isotopes-including stable isotopes, radiogenic isotopes, and radioisotopes-span from cotton used in counterfeit bills to anthrax shipped through the U.S. Postal Service and from beer adulterated with cheap adjuncts to human remains discovered in shallow graves. Recent methodological developments and the generation of isotope landscapes, or isoscapes, for data interpretation promise that isotope analysis will be a useful tool in ecological and forensic studies for decades to come.

Changes in illicit cocaine hydrochloride processing identified and revealed through multivariate analysis of cocaine signature data.

For nearly 30years, the methods utilized in illicit cocaine hydrochloride production have remained relatively consistent. Cocaine hydrochloride is typically produced one kilogram at a time. As a result, each individual kilogram is unique and distinct from other kilograms in any particular seizure based on the total alkaloid profile, occluded solvent profile, and isotopic signature. Additionally, multi-kilogram cocaine seizures are often comprised of cocaine from several different coca growing regions. There has been a documented shift in this type of processing based on the recent analysis of a large cocaine seizure in the Eastern Pacific. Signature analyses of samples from 21kg randomly selected from a 517kg seizure were virtually identical. Triplicate analyses of each sample via gas chromatography with flame ionization detection, static headspace gas chromatography mass spectrometry, and isotope ratio mass spectrometry were completed. An initial outlier evaluation of the data and an in-depth univariate analysis indicated there was no statistically significant difference among the 21 samples at the 95% confidence interval. Principal components analysis did reveal consistent minor deviations between the samples and known authentic data from the Nariño coca growing region of Colombia. These deviations were only observed on the latter principal components and could be explained by differences in solvent selection during cocaine hydrochloride processing. Chemical analyses in addition to a thorough statistical evaluation suggest a shift in the traditional small-batch method of cocaine processing to a multi-kilogram, high throughput approach.

The isotopic fractionation of carbon, nitrogen, hydrogen, and oxygen during illicit production of cocaine base in South America.

Stable isotope measurements have become a key component in sourcing the origin of illicit cocaine seized within the United States. Therefore, it is imperative to understand the process by which isotopes may be fractionated during illicit cocaine processing. In a controlled observational study, there was apparent isotopic fractionation of carbon, nitrogen, hydrogen, and oxygen. To investigate the potential source of the fractionation, cocaine base was fractionally precipitated from a dilute sulfuric acid solution with dilute ammonium hydroxide. The values of δ13C, δ15N, δ2H, and δ18O for each fraction were measured by isotope ratio mass spectrometry (IRMS). There was an equilibrium fractionation observed in all measured stable isotopes. Early fractions were depleted, and later fractions were enriched, with 15N and 2H being the most affected. The described trend is opposite of the Rayleigh distillation observed for cocaine hydrochloride precipitation.

Characterization and origin of the 'B' and 'C' compounds in the acid/neutral forensic signatures of heroin - products from the acylation of porphyroxine and subsequent hydrolysis.

Two significant compounds often found in the gas chromatographic analysis of the acid/neutral extracts from illicit heroin have remained uncharacterized for 30 years. The unknown compounds are referred to as the 'B' and 'C' compounds. It has been postulated that these compounds arise from acetylation of porphyroxine, a rhoeadine alkaloid found at trace levels in the opium poppy, Papaver somniferum. Porphyroxine was isolated from opium and acetylated to produce N,O8 -diacetylporphyroxine. Mild hydrolysis produced N,O8 -diacetyl-O14 -desmethyl-epi-porphyroxine (the C compound) and N-acetyl-O14 -desmethyl-epi-porphyroxine (the B compound). Both N,O8 -diacetyl-O14 -desmethyl-epi-porphyroxine and N-acetyl-O14 -desmethyl-epi-porphyroxine were determined to be C-14 epimers of porphyroxine and N,O8 -diacetylporphyroxine. The non-epimerized isomers of the B and C compounds were also detected in illicit heroin, but at much lower levels. Chromatographic and spectroscopic data are presented for the aforementioned compounds. The presence/absence and relative concentrations of these compounds is presented for the four types of heroin (Southwest Asian, South American, Southeast Asian, and Mexican). The prevalence of detection for the B and C compounds are Southwest Asian = 92-93%, South American = 64-72%, Southeast Asian = 45-49%, and Mexican ≤ 3%. When detected, the overall trend of relative concentrations of dicaetylporhyroxine, the B-compound, and C-compound is Southwest Asian > South American > Southeast Asian, each by an order of magnitude. These compounds were rarely detected in Mexican heroin. The presence/absence and relative concentrations of these compounds provide pertinent forensic signature characteristics that significantly enhance the final regional classifications. Copyright © 2015 John Wiley & Sons, Ltd.

Geographically Sourcing Cocaine's Origin - Delineation of the Nineteen Major Coca Growing Regions in South America.

Previously, geo-sourcing to five major coca growing regions within South America was accomplished. However, the expansion of coca cultivation throughout South America made sub-regional origin determinations increasingly difficult. The former methodology was recently enhanced with additional stable isotope analyses ((2)H and (18)O) to fully characterize cocaine due to the varying environmental conditions in which the coca was grown. An improved data analysis method was implemented with the combination of machine learning and multivariate statistical analysis methods to provide further partitioning between growing regions. Here, we show how the combination of trace cocaine alkaloids, stable isotopes, and multivariate statistical analyses can be used to classify illicit cocaine as originating from one of 19 growing regions within South America. The data obtained through this approach can be used to describe current coca cultivation and production trends, highlight trafficking routes, as well as identify new coca growing regions.

Headspace-gas chromatographic-mass spectrometric analysis of South American commercial solvents and their use in the illicit conversion of cocaine base to cocaine hydrochloride.

This study presents data that establish the makeup of solvents utilized in illicit cocaine hydrochloride production, as determined via the identification of the occluded solvents in the crystal matrix of the final product. The occluded solvent ratios can differ dramatically from the ratios of the original processing solvents. Additionally, the presented data suggest the diversion of commercial solvents to illicit cocaine hydrochloride laboratories. Thirty-five commercial solvents were obtained from five chemical manufacturing companies in South America. Each solvent was qualitatively and quantitatively analyzed using static headspace-gas chromatography-mass spectrometry (HS-GC-MS). After obtaining the chemical profile for each commercial solvent, solvents and/or solvent mixtures were prepared to be comparable in composition to several of the commercial products. Over 90 individual batches of cocaine hydrochloride were prepared from cocaine base using these solvents or solvent mixtures, which match those most commonly employed in clandestine laboratories. Additionally, a number of unique manufacturing by-products produced from processing solvents were identified, and their significance is discussed.

Differentiation of South American crack and domestic (US) crack cocaine via headspace-gas chromatography/mass spectrometry.

South American 'crack' cocaine, produced directly from coca leaf, can be distinguished from US domestically produced crack on the basis of occluded solvent profiles. In addition, analysis of domestically produced crack indicates the solvents that were used for cocaine hydrochloride (HCl) processing in South America. Samples of cocaine base (N=3) from South America and cocaine from the USA (N=157 base, N=88 HCl) were analyzed by headspace-gas chromatography-mass spectrometry (HS-GC-MS) to determine their solvent profiles. Each cocaine HCl sample was then converted to crack cocaine using the traditional crack production method and re-examined by HS-GC-MS. The resulting occluded solvent profiles were then compared to their original HCl solvent profiles. Analysis of the corresponding crack samples confirmed the same primary processing solvents found in the original HCl samples, but at reduced levels. Domestically seized crack samples also contained reduced levels of base-to-HCl conversion solvents. In contrast, analysis of South American crack samples confirmed the presence of low to high boiling hydrocarbons and no base-to-HCl conversion solvents. The presented study showed analysis of crack cocaine samples provides data on which processing solvents were originally utilized in the production of cocaine HCl in South America, prior to conversion to crack cocaine. Determination of processing solvents provides valuable information to the counter-drug intelligence community and assists the law enforcement community in determining cocaine distribution and trafficking routes throughout the world.

Rapid determination of the isomeric truxillines in illicit cocaine via capillary gas chromatography/flame ionization detection and their use and implication in the determination of cocaine origin and trafficking routes.

The isomeric truxillines are a group of minor alkaloids present in all illicit cocaine samples. The relative amount of truxillines in cocaine is indicative of the variety of coca used for cocaine processing, and thus, is useful in source determination. Previously, the determination of isomeric truxillines in cocaine was performed with a gas chromatography/electron capture detection method. However, due to the tedious sample preparation as well as the expense and maintenance required of electron capture detectors, the protocol was converted to a gas chromatography/flame-ionization detection method. Ten truxilline isomers (alpha-, beta-, delta-, epsilon-, gamma-, omega, zeta-, peri-, neo-, and epi-) were quantified relative to a structurally related internal standard, 4',4″-dimethyl-α-truxillic acid dimethyl ester. The method was shown to have a linear response from 0.001 to 1.00 mg/mL and a lower detection limit of 0.001 mg/mL. In this method, the truxillines are directly reduced with lithium aluminum hydride and then acylated with heptafluorobutyric anhydride prior to analysis. The analysis of more than 100 cocaine hydrochloride samples is presented and compared to data obtained by the previous methodology. Authentic cocaine samples obtained from the source countries of Colombia, Bolivia, and Peru were also analyzed, and comparative data on more than 23,000 samples analyzed over the past 10 years with the previous methodology is presented.

Chemosystematic identification of fifteen new cocaine-bearing Erythroxylum cultigens grown in Colombia for illicit cocaine production.

Colombian coca farmers have historically cultivated three varieties of coca for cocaine production (Erythroxylum novogranatense var. novogranatense, Erythroxylum novogranatense var. truxillense, and Erythroxylum coca var. ipadu). Within the past 13 years, 15 new cultigens of cocaine-bearing Erythroxylum have been propagated by Colombian coca farmers; each with differing physical characteristics, yet producing cocaine alkaloids at similar levels found in the historical and native varieties. Fifteen new cultigens were collected from throughout Colombia and propagated along with the three historical varieties within an experimental field in Colombia. Five plants/cultigen were randomly selected and examined for alkaloid content to determine their varietal characteristics when compared to the three known varieties. Ten cultigens gave classic Erythroxylum coca var. ipadu alkaloid profiles, four cultigens produced alkaloid profiles consistent with a hybridization of Erythroxylum novogranatense and Erythroxylum coca var. ipadu, while one cultigen gave heterogeneous alkaloid profiles that could not be characterized.

Profiling of illicit fentanyl using UHPLC-MS/MS.

Methodology is presented for the profiling of fentanyl in seized drugs using ultra high performance liquid chromatography in combination with tandem mass spectrometry (UHPLC-MS/MS). Target analysis was performed for 40 fentanyl processing impurities, several of which are markers for a specific synthetic route (Siegfried or Janssen). For the separation of these solutes, an Acquity BEH C18 1.7 µm particle column (150 mm × 2.1 mm) with a binary 1% formic acid (pH 2.0)/acetonitrile gradient was used. For MS/MS detection, an atmospheric pressure positive electrospray source was employed with selected reaction monitoring (SRM). The coupling of the high separation power of UHPLC with the highly selective and sensitive detection of MS/MS is amenable to the determination of synthetic route and linking of drug seizures. The technology is also applicable to exhibits containing trace levels of fentanyl in the presence of significantly excess amounts of heroin and/or adulterants.

Determination of phenyltetrahydroimidazothiazole enantiomers (Levamisole/Dexamisole) in illicit cocaine seizures and in the urine of cocaine abusers via chiral capillary gas chromatography-flame-ionization detection: clinical and forensic perspectives.

Illicit cocaine laboratories in South America have been adding phenyltetrahydroimidazothiazole enantiomers (levamisole and/or tetramisole) to refined illicit cocaine for over 8 years. A chiral capillary gas chromatographic methodology is presented for phenyltetrahydroimidazothiazole enantiomer determination in illicit cocaine samples and in the urine of cocaine abusers. Illicit cocaine samples (N = 752) and urine specimens from cocaine abusers (N = 50) that contained phenyltetrahydroimidazothiazole were analyzed for enantiomeric composition. Legitimate commercial preparations of phenyltetrahydroimidazothiazole are either 100% levamisole or a 50:50 mixture of levamisole and dexamisole (tetramisole). Specimens that contain phenyltetrahydroimidazothiazole mixtures that are other than 50:50 preparations will be enhanced in one isomer over the other, and they are referred to as either "levamisole-enhanced" or "dexamisole-enhanced". Cocaine samples were found to contain levamisole (N = 495, 66%), tetramisole (N = 143, 19%), and levamisole-enhanced enrichment (N = 114, 15%); urine specimens contained levamisole (N = 23, 46%), levamisole-enhanced enrichment (N = 10, 20%), and dexamisole-enhanced enrichment (N = 13, 26%). The toxicological and forensic aspects of these findings are discussed.

14C analyses quantify time lag between coca leaf harvest and street-level seizure of cocaine.

Measurements were made on the natural abundance (14)C content (Δ(14)C) of cocaine specimens seized between 2003 and 2009. The objective of this study was to determine the extent to which Δ(14)C analyses could quantify the "age" of recent cocaine seizures. Here "age" of a seized cocaine specimen is defined as the time period between when a coca leaf was harvested in South America and its seizure as cocaine at either the international or domestic street levels. Based on Δ(14)C analyses of seizure specimens, there were no statistically significant differences in the ages of domestic cocaine HCl and cocaine base specimens seized on the streets in different locations across the United States. Between 2007 and 2009, the average age of a street-level cocaine seizure in the United States was 24.6±1.1 months. Cocaine shipment seizures that were in excess of 150 kg during this time period had an average age of 18.2±1.4 months, whereas smaller shipment seizures were significantly older with an average age of 22.3±0.6 months. Analyses of the largest cocaine shipment seizures suggested that these seizures were composed of specimens with different ages, possibly representing accumulations over as much as a 31-month period.

Signature profiling and classification of illicit heroin by GC-MS analysis of acidic and neutral manufacturing impurities.

The illicit manufacture of heroin results in the formation of trace level acidic and neutral impurities. These impurities are detectable in illicit heroin and provide valuable information about the manufacturing process used. The isolation, derivatization, and semiquantitative analysis of neutral and acidic heroin manufacturing impurities by programmed temperature vaporizing injector-gas chromatography-mass spectrometry (PTV-GC-MS) is described. Trace acidic and neutral heroin impurities were isolated from basic fractions using liquid-liquid extraction. Extracted impurities were treated with N-Methyl-N-trimethylsilyltrifluoroacetamide followed by PTV-GC-MS analyses. Semiquantitative data were obtained using full scan mass spectrometry utilizing unique ions or ion combinations for 36 trace impurities found in crude and/or highly refined heroin samples. Minimum detection limits for acidic and neutral impurities were estimated to be at the 10(-7) level relative to total morphine. Over 500 authentic heroin samples from South America, Mexico, Southwest Asia, and Southeast Asia were analyzed. Classification of illicit heroin based on the presence or absence and relative amounts of acidic and neutral impurities is presented.

The effects of ambient solar UV radiation on alkaloid production by Erythroxylum novogranatense var. novogranatense.

Truxillines are alkaloids produced by Erythroxylum species and are thought to be derived from the UV-driven dimerization of cinnamoylcocaines. This study was conducted to determine the effects of ambient UV radiation on the production of truxillines in Erythroxylum novogranatense var. novogranatense. Field plants were grown under shelters covered with plastic filters that were transparent to UV radiation, filtered UV-B, or both filtered UV-B and UV-A radiation. The treatments had no significant effect on plant biomass or specific leaf weight. Absorption values in the UV-C and UV-A region of acidified-methanol leaf extracts were higher for plants exposed to UV radiation compared to the no UV radiation treatment. There was a trend in decreasing levels of trans-cinnamoylcocaine and a statistically significant decrease in levels of cis-cinnamoylcocaine in the leaves of plants exposed to UV radiation compared to the no UV radiation treatment. Truxilline levels increased in leaves from plants exposed to UV radiation compared to the no UV radiation treatment. Most significantly, the ratio of truxillines to total cinnamoylcocaines in the leaves was affected by UV, increasing with increased UV exposure. The results support the hypothesis that UV radiation is involved in the formation of truxillines from cinnamoylcocaines.

Chlorinated opium alkaloid derivatives produced by the use of aqueous sodium hypochlorite during the clandestine manufacture of heroin.

A clandestine chemist was observed producing heroin from crude morphine utilizing a solution of sodium hypochlorite during the process. Numerous chlorinated opium alkaloid derivatives were created when the morphine acetylation reaction was quenched and neutralized with a solution of sodium hypochlorite and ammonium hydroxide. Four of these compounds, 1-chloroheroin, 1-chloroacetylcodeine, 1-chloro-O(6)-monoacetylmorphine, and 2'-chloropapaverine, were characterized via preparative isolation, gas chromatography/mass spectrometry, nuclear magnetic resonance spectroscopy, and independent synthesis. These chlorinated derivatives were formed via electrophilic aromatic substitution with free chlorine during the illicit process. Although no illicit heroin exhibits containing these compounds have been observed in seizures to date, mass spectral data are provided for several of these compounds for their identification should they be seen within future seizures of illicit heroin.

Tropane ethyl esters in illicit cocaine: isolation, detection, and determination of new manufacturing by-products from the clandestine purification of crude cocaine base with ethanol.

Seven ethyl homologues of known tropane esters have recently been detected as impurities in the gas chromatographic signature profiles of authentic Peruvian illicit cocaine base and hydrochloride exhibits. Peruvian cocaine base processors are now known to use ethanol for the purification of crude cocaine base. This process is referred to as the "base lavada" or "washed base" process and is a recent substitute method for the potassium permanganate oxidation purification methodology. Seven ethyl ester homologues were formed in illicit cocaine from the transesterification of known tropane methyl esters or possibly ethyl esterification of their respective tropane C-2 carboxylic acids in the presence of ethanol. Exhibits containing these compounds were subjected to gas chromatographic-mass spectrometric analyses to determine their identity and were subsequently synthesized to verify their structures. Quantitative determinations were obtained from ion-pair chromatography isolation followed by gas chromatography with flame ionization detection. Specifically, hexanoylecgonine ethyl ester, cocaethylene, cis-cinnamoylecgonine ethyl ester, trans-cinnamoylecgonine ethyl ester, 3',4',5'-trimethoxybenzoylecgonine ethyl ester, cis-3',4',5'-trimethoxycinnamoylecgonine ethyl ester, and trans-3',4',5'-trimethoxycinnamoylecgonine ethyl ester were detected and characterized. When present, these compounds were detected at levels ranging from 8.6 x 10(-4) to 9.3 x 10(-1)% relative to cocaine.

Four new illicit cocaine impurities from the oxidation of crude cocaine base: formation and characterization of the diastereomeric 2,3-dihydroxy-3-phenylpropionylecgonine methyl esters from cis- and trans-cinnamoylcocaine.

Four new impurities have recently been detected in the gas chromatographic signature profiles of many illicit cocaine hydrochloride exhibits. These impurities are only seen in exhibits that have been oxidized and are most prominent in samples that have been highly oxidized. Exhibits containing these compounds were subjected to gas and liquid chromatographic-mass spectrometric analyses to determine the identity of the impurities. These impurities were subsequently synthesized to verify their structures. Four diastereomeric diols formed from the oxidation of cis- and trans-cinnamoylcocaine were characterized by nuclear magnetic resonance spectrometry, mass spectrometry, and synthesis. Oxidation of cis-cinnamoylcocaine in neutral conditions yielded (2R,3R)-dihydroxy-3-phenylpropionylecgonine methyl ester and (2S,3S)-dihydroxy-3-phenylpropionylecgonine methyl ester, while trans-cinnamoylcocaine produced (2R,3S)-dihydroxy-3-phenylpropionylecgonine methyl ester and (2S,3R)-dihydroxy-3-phenylpropionylecgonine methyl ester. The recent appearance of these new impurities suggests that some illicit cocaine processors have modified their oxidation procedures of crude cocaine base for transformation into illicit refined cocaine hydrochloride.

Neutral heroin impurities from tetrahydrobenzylisoquinoline alkaloids.

Laudanosine, reticuline, codamine, and laudanine are members of the tetrahydrobenzylisoquinoline family of natural products. These alkaloids are present in the opium poppy, Papaver somniferum, and are subsequently found as impurities in clandestinely processed morphine. Morphine is then synthesized to heroin using hot acetic anhydride. During the course of this study, it was determined that these four tetrahydrobenzylisoquinolines undergo degradation to a series of 18 neutral impurities when subjected to hot acetic anhydride. Based on the degradation pathway, these new impurities were categorized into two sets of impurities called the C1-acetates compounds and the stilbene compounds. Synthesis, isolation, and structural elucidation information is provided for the tetrahydrobenzylisoquinoline alkaloids, and the new neutral impurities have been studied. Several hundred authentic heroin samples were analyzed using an established heroin signature program method. This methodology features the detection of trace neutral impurities present in heroin samples. It was determined that all 18 new impurities were detected in various quantities in four different types of heroin samples. Analytical results featuring these new impurities are reported for South American-, Southwest Asian-, Mexican-, and Southeast Asian-type heroin samples. These new impurities, coupled with other established forensic markers, enhance the ability to classify illicit heroin samples.