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.

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.

Multicomponent (n >or= 3) sorption isotherms in reversed-phase liquid chromatography: the effect of immobilized eluent on the retention of analytes.

The experimental technique of tracer pulse chromatography was used to simultaneously measure the uptake of eluent components by a C(18)-bonded reversed-phase liquid chromatography (RPLC) packing and the retention factors for a series of test analytes over the full range of eluent composition for methanol and acetonitrile with water. The primary objective of the research was to determine whether or not the uptake of eluent components immobilized as part of the stationary phase would influence the retention of analyte standards. Both acetonitrile and methanol were absorbed in or adsorbed on the C(18)-bonded phase with the maximum amount of acetonitrile sorbed being about four times that of methanol. The thermodynamic void volume of the column and the excess sorption isotherms of acetonitrile, methanol, and water in binary aqueous/organic mixtures were determined directly from the tracer pulse experiments. The absolute sorption isotherms of the eluent components were indirectly estimated by a combination of techniques. Regression analysis of the nonstationary inflection point of the excess isotherms provided an estimate of the volume of eluent sorbed by the stationary phase but only over a limited eluent composition range. In order to expand the applicable composition range, several commonly used "unretained" probe solutes were tested to determine the accuracy of the assumption that the retention volumes of these solutes provided a viable measure of the kinetic void volume (mobile-phase volume) of the column. The difference between the thermodynamic and kinetic void volumes provided an estimate of the absolute volume of eluent present in the stationary phase. The experimental results showed that some solutes, viz., water and thiourea, did provide an accurate measure of the mobile-phase volume but only over a limited range of eluent composition. Using deuterated water as the unretained dead time marker for water-rich eluents combined with the regression results from excess isotherm data, the absolute volume of eluent sorbed by the stationary phase could be estimated over the full range of eluent composition. The effect of this uptake of eluent on the retention of the test solutes appeared to be minimal for this particular set of test analytes.

Sorption isotherms of ternary eluents in reversed-phase liquid chromatography.

The experimental technique of mass spectrometric tracer pulse chromatography was used to measure the excess volume of each eluent component for binary and ternary mixtures of water, acetonitrile, and methanol on a C(18)-bonded silica RPLC packing over the full composition range. The tracer pulse method allowed the direct measurement of excess volumes of each eluent component without numerical integration, assumed isotherm equation, detector calibration, or off-line analysis of the eluent composition. Absolute isotherms were estimated from the experimental data for excess volumes by use of various strategies for the estimation of the volumes of the stationary and mobile phases in dynamic equilibrium with eluents of varying composition. The results indicate that all three eluent components interacted with the alkane bonded phase. Some components were selectively taken up as part of the stationary phase while other components were selectively excluded so the composition of eluent in or on the stationary phase often significantly differed from the composition of the bulk eluent. The exact composition of the stationary phase (bonded phase plus immobilized eluent) was dependent upon the type and composition of the bulk eluent.

Mass spectrometric tracer pulse chromatographic investigations of eluent sorption with bonded RPLC packings.

The experimental technique of mass spectrometric tracer pulse chromatography was used to investigate the uptake of RPLC eluents by a C 18-bonded packing. The experiments were carried out with eluents consisting of binary aqueous mixtures with acetonitrile, methanol, and tetrahydrofuran over the full range of eluent composition at 25 degrees C. The primary experimental data obtained were excess volumes of sorption for the eluent components. The excess volume data were then used to determine the absolute volume of each eluent component in the stationary phase as a function of composition. The absolute volumes were calculated by utilizing a series of strategies specific to limited eluent composition range. The linear inflection region of the excess volume isotherms was used to calculate the volume and composition of the eluent in the stationary phase for organic-rich eluents. Three different assumptions were used and evaluated for the water-rich eluent compositions. The assumptions were (i) constant volume of the sorbed layer, (ii) constant amount of water sorbed, and (iii) no water sorption. The latter assumption was adopted as the most satisfactory. The calculated void volume data were compared with the retention volume of thiourea and uracil, commonly used dead time markers. Neither thiourea nor uracil proved to be an accurate measure of the kinetic void volume.

Fundamental mathematical relationships for retention volumes in liquid chromatography.

Various issues currently impeding elucidation of the retention mechanism(s) in reversed-phase liquid chromatography are discussed. A generalized derivation of the retention volume equations for both concentration and tracer pulse chromatography are presented. The derivations follow the general direction of multiple, ad hoc derivations presented in the extant literature. The retention volume equations derived herein represent a body of work previously dispersed throughout the chromatographic literature.

Strategies for the determination of the volume and composition of the stationary phase in reversed-phase liquid chromatography.

The fundamental difficulties currently impeding the elucidation of retention mechanisms in RPLC are reviewed. The various definitions and conventions concerning void volume and excess adsorption in liquid-solid systems are compared and contrasted. The utility and integrity of various chromatographic methods for the measurement of excess adsorption isotherms of HPLC eluents are discussed. Finally, literature methods for obtaining absolute adsorption data from excess quantities are reviewed and evaluated.

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.

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.

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.

Comparison of void volume, mobile phase volume and accessible volume determined from retention data for oligomers in reversed-phase liquid chromatographic systems.

The experimental technique of mass spectrometric tracer pulse chromatography was used to determine the void volume, i.e., the total volume of eluent in the column, and the volume of eluent moving freely through the column, i.e., mobile phase volume, for a series of eluents with a C(18)-bonded RPLC column. The interstitial volume of the column was determined by size exclusion chromatography. In order to evaluate the utility of the accessible volumes determined from the retention volumes of homologous solutes, the accessible volume of the column was determined as a function of eluent composition and temperature with polystyrene and polyethylene glycol samples using Martin's Rule. Comparison of these four measured volumes indicated that the experimentally measured accessible volumes did not correspond to either the void volumes, mobile phase volumes or interstitial volumes.

Interconversion of gradient and isocratic retention data in reversed-phase liquid chromatography: effect of the uptake of eluent modifier on the retention of analytes.

The experimental technique of mass spectrometric tracer pulse chromatography was used to study the effect of the sorption of eluent components by a C(18)-bonded silica RPLC packing on the retention of a series of test analytes during isocratic and gradient elution experiments. The analytes of interest were a substituted phenol, a substituted nitroaniline, an anti-malaria drug, tetrahydrofuran, and methanol. The eluent used was a mixture of acetonitrile and water. The solutes and isotopically labeled eluent components were injected at fixed time intervals during each gradient run. The mass specific detector allowed the assignment of individual analyte peaks even when there was overlap in the chromatograms from successive injections. Thus, the retention time of each analyte could be determined as a function of gradient slope and initial eluent composition at the time of each injection. Experimental gradient retention time data were then compared with the calculated results from two theoretical models. The first model assumed the velocity of the mobile phase and eluent were equal. The second and most realistic model assumed the velocity of the eluent was less than the velocity of the mobile phase due to the uptake of eluent by the stationary phase. Gradient retention times predicted by the two models were reasonably accurate with the sorption model giving slightly more accurate values. Inverse calculations, i.e., calculation of isocratic retention factors from gradient elution data were also carried out with very similar results. That is, the model allowing for the uptake of eluent was slightly more accurate than the model assuming no eluent-stationary phase interaction.