Gas Chromatography-electron Ionization-mass Spectrometry Research Articles

Improved Discrimination of Mass Spectral Isomers Using the High-Dimensional Consensus Mass Spectral Similarity Algorithm.

This study employs a high-dimensional consensus mass spectral (HDCMS) similarity scoring technique to discriminate isomers collected using an electron ionization mass spectrometer. The HDCMS method was previously introduced and applied to the discrimination of mass spectra of constitutional isomers, methamphetamine and phentermine, collected with direct analysis real-time mass spectrometry (DART-MS). The method formulates the problem of discriminating mass spectra in a mathematical Hilbert space and is hence called "high dimensional." It requires replicate mass spectra to build a Gaussian model and evaluate the inner products between these functions. The resulting measurement variability is used as a signature by which to discriminate spectra. In this work, HDCMS is tested on electron impact ionization (EI) mass spectra of 7 terpene and terpene-related (C10H16 and C10H14) isomers with experimental retention indices that differ by less than 30 and with traditional cosine similarity scores greater than 0.9, on a scale of 0 to 1, when compared with at least one other compound in the test set. Using identical instrument parameters, 15 replicate gas chromatography-electron ionization-mass spectrometry (GC-EI-MS) spectra of each isomer were collected and separated into distinct library and query sets. The HDCMS algorithm discriminated each isomer, indicating the method's potential. Because the method requires replicate measurements, observations from a simple heuristic study of the number of replicates required to discriminate these isomers is presented. The paper concludes with a discussion of compound discrimination using HDCMS and the benefits and drawbacks of applying the method to EI-MS data.

Structural characterization of nitazene analogs using electron ionization-mass spectrometry (EI-MS)

Nitazene analogs are among the most recent and potent additions to the novel synthetic opioid (NSO) market, and new analogs continue to emerge. Seized drug analysis commonly utilizes gas chromatography-electron ionization-mass spectrometry (GC-EI-MS), so it is therefore imperative to understand how nitazene analogs behave under EI-MS conditions, and how substitution at various sites on the molecule may impact the resulting EI mass spectra. This study characterizes the EI fragmentation behavior of 20 representative nitazene analogs that contain differing substitutions and proposes rational mechanisms to explain the observed behavior.A general EI fragmentation pathway for nitazene analogs was proposed, with the most common nitazene fragment ions being observed at m/z 86, m/z 107, m/z 58, and m/z 77. Characteristic ions were determined for different substitution groups, enabling the identification of diethyl, desethyl, pyrrolidine, and piperidine substitutions at the amine moiety, and different alkoxy chain lengths at the aromatic ring of the benzyl group. Mechanisms for the formation of these characteristic ions were proposed with the aid of isotopically labeled standards and high-resolution mass spectrometry measurements. To help with the interpretation of EI mass spectra for nitazene analogs, decision trees were developed that encompass the characteristic fragment ions observed for substitutions to the amine moiety and benzyl group, with additional criteria provided for substitutions to the benzimidazole moiety. This study summarizes the fragmentation patterns and characteristic fragment ions in the EI mass spectra of 20 representative nitazene analogs, which will aid the seized drug community in identifying novel nitazene analogs.

Determination of N,N-Dialkylethanolamines Related to Chemical Warfare Agents as Benzylated Species in an OPCW Proficiency Test Soil Sample by Electron Ionization Gas Chromatography-Mass Spectrometry

In the field of chemical warfare agents (CWAs), ethanolamines are important building blocks for the preparation of nerve agents belonging to the V-series such as VX and VR as well as the nitrogen-based mustards. In this work, the determination of three members in this class of Schedule 2B reportable chemicals, namely N,N-dimethylethanolamine (DMEA), N,N-diethylethanolamine (DEEA), and N,N-diisopropylethanolamine (DIEA) by electron ionization gas chromatography-mass spectrometry (EI-GC-MS) is presented. The three dialkylethanolamines were spiked in a high organic content soil matrix featured during the 45th Organization for the Prohibition of Chemical Weapons (OPCW) proficiency test (PT) at two separate concentrations each (1 and 10 µg/g separately), simulating values often encountered during these tests. The N,N-dialkylethanolamines were O-alkylated using benzyl bromide and sodium carbonate at 55 °C and were detected as their benzylated versions by electron ionization GC-MS. The benzylation of each dialkylethanolamine yielded derivatives with higher molecular weight and therefore different GC-MS profiles such as longer retention times and sharper peak chromatography relative to their unmodified counterparts. These characteristics enable the analytical chemist to detect and confirm these important CWA-related chemicals specifically in instances where they are present in low concentrations (i.e. <10 µg/g) among more abundant interferences.

Investigating the effect of substitution location on fentanyl analog identification for methyl-substituted fentanyl analogs using GC-EI-MS

Fentanyl and fentanyl analogs are key figures at the center of the ongoing opioid epidemic in the United States. Currently, gas chromatography-electron ionization-mass spectrometry (GC-EI-MS) instrumentation is one of the main analytical tools used for the identification of fentanyl analogs in forensic laboratories. However, the resulting retention times and EI mass spectra for many fentanyl analogs are quite similar given that there are only subtle differences in their structures. This research investigates the effect of substitution location on fentanyl analog identification using GC-EI-MS and the National Institute of Standards and Technology (NIST) Simple Similarity Search (SSS) and Hybrid Similarity Search (HSS) algorithms through the analysis of a series of 11 methyl-substituted fentanyl analogs.The 95% confidence interval retention times for each isomer were never more than ±0.009 min and a statistically significant (p < 0.001) difference was observed for all pairwise comparisons. Careful examination of the EI mass spectra revealed several general trends about the influence of substitution location on the observed EI mass spectra. The combination of unique fragmentation patterns, formed through shifts in m/z values due to the location of substitution, and the retention time enabled the identification of the 11 methyl-substituted fentanyl analogs analyzed in this study. The SSS was able to correctly identify all methyl-substituted fentanyl analogs within the top five hits for compounds that were present in the library. In comparison, the HSS was used to identify compounds absent from the mass spectral libraries and provided drastically different results depending on the location of substitution.

The differentiation of N-butyl pentylone isomers using GC-EI-MS and NMR

Forensic laboratories are faced with an ever-expanding seized drug landscape including the increasing prevalence of novel psychoactive substances (NPS), such as synthetic cathinones, that have varying potencies and scheduling. This study demonstrates a combined gas chromatography-electron ionization-mass spectrometry (GC-EI-MS) and nuclear magnetic resonance (NMR) spectroscopy approach for the differentiation of N-butyl pentylone isomers based on distinct retention times, characteristic EI mass spectra, and NMR characterization. Retention time reproducibility was assessed from 60 replicate measurements for each isomer over the course of a month. In addition, the effect of the mass spectrometer tune and the stability of an identified characteristic ion ratio using spectral data from ± 1 scan on either side of the peak apex were also statistically assessed using Welch’s ANOVA testing.The presence of diastereomers for N-sec-butyl pentylone was identified using the developed GC-EI-MS method, which was confirmed using one-dimensional and two-dimensional NMR spectroscopy. The retention time reproducibility of the chromatographic method was ± 0.076% or less over the course of a month. An identified characteristic ion ratio between the abundance of the fragment ion at m/z 128 and the fragment ion at m/z 72 enabled the differentiation of the four N-butyl pentylone isomers, even when accounting for the effect of the mass spectrometer tune and mass spectral scans used to calculate the characteristic ion ratio. The 95% confidence interval mean abundance ratio of the fragment ions at m/z 128 and m/z 72 was 17.14 ± 0.14 for N-butyl pentylone, 6.44 ± 0.05 for N-isobutyl pentylone, 3.38 ± 0.02 for N-sec-butyl pentylone, and 0.75 ± 0.01 for N-tert-butyl pentylone. These results highlight the capabilities of a combined GC-EI-MS and NMR approach for the differentiation and characterization of synthetic cathinone isomers.

Practical benzylation of N,N-substituted ethanolamines related to chemical warfare agents for analysis and detection by electron ionization gas chromatography-mass spectrometry.

The benzylation of three low molecular weight N,N-disubstituted ethanolamines related to chemical warfare agents (CWAs) to furnish derivatives with improved gas chromatography-mass spectrometry (GC-MS) profiles is described. Due to their low molecular weight and polar nature, N,N-disubstituted ethanolamines are notoriously difficult to detect by routine GC-MS analyses during Organisation for the Prohibition of Chemical Weapons (OPCW) proficiency tests (PTs), particularly in scenarios when they are present at low levels (~1-10 ppm) amidst more abundant interferences. Our studies revealed that the optimal derivatization conditions involved the treatment of the ethanolamine with benzyl bromide in the presence of an inorganic base (e.g., Na2 CO3 ) in dichloromethane at 55°C for 2 h. This optimized set of conditions was then successfully applied to the derivatization of N,N-dimethylethanolamine, N,N-diethylethanolamine and N,N-diisopropylethanolamine present separately at 1 and 10 μg/mL concentrations in a glycerol-rich matrix sample featured in the 48th OPCW PT. The benzylated derivatives of the three ethanolamines possessed retention times long enough to clear the massive glycerol-containing matrix interferences. The protocol herein is introduced as an alternative method for derivatization of these CWA and pharmaceutically important species and should find broad applicability in laboratories where routine forensic analysis is carried out.

Detection and confirmation of fentanyls in high clay-content soil by electron ionization gas chromatography-mass spectrometry.

Detection of illicit drugs in the environment, particularly in soils, often suggests the present or past location of a clandestine production center for these substances. Thus, development of efficient methods for the analysis and detection of these chemicals is of paramount importance in the field of chemical forensics. In this work, a method involving the extraction and retrospective confirmation of fentanyl, acetylfentanyl, thiofentanyl, and acetylthiofentanyl using trichloroethoxycarbonylation chemistry in a high clay-content soil is presented. The soil was spiked separately with each fentanyl at two concentrations (1 and 10 μg/g) and their extraction accomplished using ethyl acetate and aqueous NH4 OH (pH ~ 11.4) with extraction recoveries ranging from ~56% to 82% for the high-concentration (10 μg/g) samples while ranging from ~68% to 83% for the low-concentration (1 μg/g) samples. After their extraction, residues containing each fentanyl were reacted with 2,2,2-trichloroethoxycarbonyl chloride (Troc-Cl) to generate two unique and predictable products from each opioid that can be used to retrospectively confirm their presence and identity using EI-GC-MS. The method's limit of detection (MDL/LOD) for Troc-norfentanyl and Troc-noracetylfentanyl were estimated to be 29.4 and 31.8 ng/mL in the organic extracts. In addition, the method's limit of quantitation for Troc-norfentanyl and Troc-noracetylfentanyl were determined to be 88.2 and 95.5 ng/mL, respectively. Collectively, the results presented herein strengthen the use of chloroformate chemistry as an additional chemical tool to confirm the presence of these highly toxic and lethal substances in the environment.

Synthesis, characterisation and quantification of the new psychoactive substance 1-(1,3-benzodioxol-5-yl)-2-(propylamino)butan-1-one (bk-PBDB, putylone)

Synthetic cathinones are a continually evolving family of illicit drugs, with novel analogues frequently being detected. This paper reports the detection of 1-(1,3-benzodioxol-5-yl)-2-(propylamino)butan-1-one, bk-PBDB (putylone), within solid dosage forms (tablets) seized by law enforcement for the first time in the United Kingdom. The identity of the compound was confirmed via the synthesis of a pure bk-PBDB reference standard and direct spectral comparison by 1H NMR and GC-EI-MS analysis. A full analytical profiling of bk-PBDB by nuclear magnetic resonance (NMR), attenuated total reflection Fourier-transform infrared (FTIR) spectroscopy and gas chromatography-electron ionisation-mass spectrometry (GC-EI-MS) is reported and shows good concordance between the seized sample and the reference standard. A validated GC-EI-MS method for the routine quantification of the cathinone in bulk forensic samples (LOD: 0.09 μg/mL, LOQ: 0.26 μg/mL) was also developed and using this method, the seized tablets were determined to contain a mixture of bk-PBDB (130.6–135.5 mg/tablet) and caffeine (40.2–43.4 mg/tablet) respectively.

Novel In Vitro Intestinal Microbiome Model to Study Lipidomic and Metabolomic Adaptations Due to Exposure to Glyphosate, Perfluorooctanoic Acid, and Docusate Sodium.

Cell and animal models have been used to provide insights with regard to physiological changes in intestinal flora due to exposure to drugs and environmental contaminants. Here, a novel in vitro model known as simulator of the human intestinal microbial ecosystem (SHIME) was used to assess the effects of three chemicals of emerging concern, namely glyphosate, perfluorooctanoic acid (PFOA), and docusate sodium (dioctyl sulfosuccinate, DOSS), on the lipidomic and metabolomic profiles of the gut microenvironment in both the proximal and distal colonic compartments. Nontargeted analyses by ultra-high performance liquid chromatography-tandem mass spectrometry and gas chromatography-electron ionization-mass spectrometry revealed minor differences in the lipidomic and metabolomic signatures of the proximal and distal colon following treatment with either glyphosate or PFOA at acceptable human daily intake levels or average daily exposures. However, global dysregulation of lipids and metabolites was observed due to DOSS treatment at conventional prescription doses when indicated as a stool softener. Our findings suggest that the current guidelines for glyphosate and PFOA exposure may be adequate at the level of the lower gut microbiome in healthy adults, but the probable yet uncharacterized off-target effects, safety, and efficacy of long-term DOSS treatment warrants further investigation. Indeed, we highlight the SHIME system as a novel in vitro approach which can be used as a screening tool to assess the impact of drugs and/or chemicals on the gut microbiome, while implementing state-of-the-art and data-driven mass spectrometric workflows to identify toxic lipidomic and metabolomic signatures.

Non-target GC–MS analyses of fecal VOCs in NASH-hepatocellular carcinoma model STAM mice

The increased incidence of obesity in the global population has increased the risk of several chronic inflammation-related diseases, including non-alcoholic steatohepatitis (NASH)-hepatocellular carcinoma (HCC). The progression from NASH to HCC involves a virus-independent liver carcinogenic mechanism; however, we currently lack effective treatment and prevention strategies. Several reports have suggested that fecal volatile organic compounds (VOCs) are strongly associated with NASH-HCC; therefore, we explored the biomarkers involved in its pathogenesis and progression. Fecal samples collected from control and NASH-HCC model STAM mice were subjected to headspace autosampler gas chromatography-electron ionization-mass spectrometry. Non-target profiling analysis identified diacetyl (2,3-butandione) as a fecal VOC that characterizes STAM mice. Although fecal diacetyl levels were correlated with the HCC in STAM mice, diacetyl is known as a cytotoxic/tissue-damaging compound rather than genotoxic or mutagenic; therefore, we examined the effect of bioactivity associated with NASH progression. We observed that diacetyl induced several pro-inflammatory molecules, including tumor necrosis factor-α, cyclooxygenase-2, monocyte chemoattractant protein-1, and transforming growth factor-β, in mouse macrophage RAW264.7 and Kupffer KPU5 cells. Additionally, we observed that diacetyl induced α-smooth muscle actin, one of the hallmarks of fibrosis, in an ex vivo cultured hepatic section, but not in in vitro hepatic stellate TWNT-1 cells. These results suggest that diacetyl would be a potential biomarker of fecal VOC in STAM mice, and its ability to trigger the macrophage-derived inflammation and fibrosis may partly contribute to NASH-HCC carcinogenesis.

Determination of fentanyl and acetylfentanyl in soil in their intact form and orthogonal corroboration of their presence by EI-GC-MS using chloroformate chemistry

In the field of forensic chemistry, methods for identifying and characterizing illicit drugs remain of great interest to government and law enforcement entities. To this end, the unambiguous identification of a drug in the environment (e.g., soil) prompts further investigation into the possibility of a clandestine point of production for such drug. In this report, a method for the analysis of the two common synthetic opioids fentanyl and acetylfentanyl in a soil matrix when present separately at 1 and 10 μg/g by electron ionization gas chromatography-mass spectrometry (EI-GC-MS) is presented. The method involves the initial extraction of the intact opioids followed by their reaction with 2,2,2-trichloroethoxycarbonyl chloride (Troc-Cl) to predictably generate Troc-modified norfentanyl, Troc-noracetylfentanyl and 2-phenylethyl chloride products that can be used to retrospectively corroborate the identity of the opioid by EI-GC-MS. Estimated method detection limits (MDLs) for the Troc-norfentanyl and Troc-noracetylfentanyl species were 15.4 and 13.6 ng/mL respectively from the soil matrix extracts employed (Virginia type A soil) while the calculated method’s LOQ values were 45.7 and 40.4 ng/mL respectively. The protocol presented herein describes for the first time the analysis of these highly toxic and common synthetic opioids in a soil matrix as well as their retrospective confirmation by analyzing the products arising from their reaction with Troc-Cl by EI-GC-MS.

Analytical characterization and differentiation between threo- and erythro-4-fluoroethylphenidate.

Methylphenidate analogs appeared on the drug market during the last years. Its analogs contain two chiral centers and, thus, have potential varying configurations (i.e., threo and erythro forms). This study presents the analytical characterization of 4-fluoroethylphenidate (4-FEP) and its differentiation between threo- and erythro-4-FEP. Analysis of the samples included high-performance liquid chromatography (HPLC), gas chromatography-electron ionization-mass spectrometry (GC-EI-MS), high-resolution mass spectrometry (HRMS) analyses, nuclear magnetic resonance (NMR) spectroscopy and X-ray crystal structure analysis. NMR spectroscopic investigations confirmed the differences between threo- and erythro-4-FEP, and demonstrated that both isomers could be separated using HPLC and GC methods. Two samples obtained from one vendor in 2019 consisted of threo-4-FEP, whereas the other two samples obtained from a different vendor in 2020 consisted of a mixture of threo- and erythro-4-FEP. Several analytical approaches including HPLC, GC-EI-MS, HRMS analyses, NMR spectroscopy and X-ray crystal structure analysis enabled the unambiguous identification of threo- and erythro-4-FEP. The analytical data presented in this article will be useful for identifying threo- and erythro-4-FEP included in illicit products.

Rapid screening of Chemical Weapons Convention-related chemicals in oil matrix by headspace solid-phase microextraction and gas chromatography-mass spectrometry

The Chemical Weapons Convention (CWC) requires verification of a large number of compounds with different types and properties. The results of the verification are of great political and military sensitivity. However, the sources of verification samples are complex and diverse, and the contents of the target compounds in these samples are usually very low. These issues increase the likelihood of missed or false detection. Thus, establishing rapid and effective screening methods for the accurate identification of CWC-related compounds in complex environmental samples are of great importance. In this study, a fast and simple procedure based on headspace solid-phase microextraction (HS-SPME) followed by gas chromatography-electron ionization mass spectrometry (GC-EI/MS) in full-scan mode was developed to determine CWC-related chemicals in oil matrix. A total of 24 CWC-related chemicals with different chemical characteristics were selected to simulate the screening procedure. The selected compounds were divided into three groups based on their properties. The first group included volatile and semi-volatile CWC-related compounds with relatively low polarity, which could be extracted by HS-SPME and directly analyzed by GC-MS. The second group included moderately polar compounds with hydroxyl or amino groups; such compounds are related to nerve, blister, and incapacitating agents. The compounds in the third group included non-volatile CWC-related chemicals with relatively strong polarity, such as alkyl methylphosphonic acids and diphenyl hydroxyacetic acid. These compounds must be derivatized into vaporizable derivatives prior to extraction by HS-SPME and analysis by GC-MS. Variables that influence the SPME process, such as fiber type, extraction temperature and time, desorption time, and derivatization protocol, were optimized to improve the sensitivity of the method. The screening procedure for CWC-related compounds in the oil matrix samples included two main steps. First, low-polarity volatile and semi-volatile compounds (i. e. the first group) were extracted by HS-SPME with divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fibers and analyzed in split-injection mode (split ratio, 10∶1) using GC-MS. The use of a large split ratio can reduce the solvent effect, which is conducive to the detection of low-boiling-point compounds. If necessary, the sample could be extracted once more and analyzed in splitless mode. The derivatization agent bis(trimethylsilyl)trifluoroacetamide (BSTFA) was then added to the sample. Mid- and high-polarity compounds (i. e. the second and third groups) were extracted with polydimethylsiloxane/divinylbenzene (PDMS/DVB) fibers after derivatization and analyzed in splitless mode using GC-MS. The established method exhibited good repeatability and sensitivity. The detection limits for the compounds in the first group ranged from 0.5 ng/mL to 100 ng/mL, whereas the detection limits for the compounds in the second and third groups ranged from 20 ng/mL to 300 ng/mL. Except for compounds with extremely high boiling points and a few compounds that are not suitable for derivatization with BSTFA, the method can be used to analyze most CWC-related compounds in oil matrix samples. In particular, it greatly shortened the preparation time of the oil matrix samples and reduced the loss of low-boiling-point compounds owing to the sample concentration process, thereby avoiding missed detection. The method was successfully applied to the Organization for the Prohibition of Chemical Weapons (OPCW) proficiency tests and proved to be a useful technique for the rapid screening of trace levels of CWC-related chemicals in oil matrix.

Determination of double bond positions in methyl ketones by gas chromatography-mass spectrometry using dimethyl disulfide derivatives.

Methyl ketones are of interest for the application as biofuels. The fatty acid metabolism of different microbes has been rearranged to achieve a sustainable production of methyl ketones. The biofuel properties and possible further chemical modifications of these methyl ketones are influenced by their chain length, as well as their degree of saturation and the corresponding double bond position. A method based on gas chromatography-electron ionization; mass spectrometry (GC-EI-MS) was used to determine the double bond position of methyl ketones. Derivatization using dimethyl disulfide (DMDS) and an iodine catalyst enabled the activation of the double bonds for selective fragmentation during electron ionization. The cleavage led to key fragments in the Orbitrap high-resolution mass spectrum and allowed the unequivocal localization of the double bond position of the respective monounsaturated methyl ketone. The double bond position of several medium chain length methyl ketones originating from the gram-negative bacterium Pseudomonas taiwanensis (P. taiwanensis) VLB120 was determined. The DMDS derivatives of methyl ketones can yield isobaric fragment ions for different possible double bond positions, which can be distinguished only using high-resolution MS. The double bond position of all methyl ketones deriving from P. taiwanensis VLB120 was the same, counting from the end of the aliphatic chain, and was determined as ω-7. The derivatization of medium chain length monounsaturated methyl ketones with DMDS allowed the determination of the corresponding double bond position via GC-EI-MS. High-resolution MS is needed to differentiate possible double bond positions that yield isobaric fragment ions.

Mass calibrants for positive chemical ionization-high resolution mass spectrometry (CI-HRMS) for the identification of unknown compounds using accurate mass measurements.

Gas Chromatography-Electron Ionization-Mass Spectrometry (GC-EI-MS) is still the most routinely performed method for metabolite profiling as compared to other hyphenated techniques. But when it comes to identification of unknown compounds, information on the molecular weight is not readily available because the molecular ion is not always found with electron ionization (EI). Thus, the use of chemical ionization (CI) is envisaged that commonly produces the molecular ion; in combination with accurate mass measurement, this technique would further allow for calculation of sum formulas of those compounds. However, for proper accuracy of analysis, a mass calibrant is needed. We set out to find a commercially available reference material with mass peaks that would qualify the substance as mass calibrant under CI conditions. Six commercially available mass calibrants, FC 43, PFK, Ultramark 1621, Ultramark 3200F, Triton X-100, and PEG 1000, were tested under CI conditions to understand their fragmentation behavior. Our findings indicate that Ultramark 1621 and PFK best fit the expectations of a mass calibrant for HRMS analysis whereby PFK provided a fragmentation pattern similar to EI outcomes thus enabling the use of mass reference tables commonly provided within commercial mass spectrometers. On the other hand, Ultramark 1621 is a mixture of fluorinated phosphazines that shows stable fragment intensities.

Benzyl trichloroacetimidates as derivatizing agents for phosphonic acids related to nerve agents by EI-GC-MS during OPCW proficiency test scenarios.

The use of benzyl trichloroacetimidates for the benzylation of phosphonic acid nerve agent markers under neutral, basic, and slightly acidic conditions is presented. The benzyl-derived phosphonic acids were detected and analyzed by Electron Ionization Gas Chromatography-Mass Spectrometry (EI-GC-MS). The phosphonic acids used in this work included ethyl-, cyclohexyl- and pinacolyl methylphosphonic acid, first pass hydrolysis products from the nerve agents ethylN-2-diisopropylaminoethyl methylphosphonothiolate (VX), cyclosarin (GF) and soman (GD) respectively. Optimization of reaction parameters for the benzylation included reaction time and solvent, temperature and the effect of the absence or presence of catalytic acid. The optimized conditions for the derivatization of the phosphonic acids specifically for their benzylation, included neutral as well as catalytic acid (< 5mol%) and benzyl 2,2,2-trichloroacetimidate in excess coupled to heating the mixture to 60°C in acetonitrile for 4h. While the neutral conditions for the method proved to be efficient for the preparation of the p-methoxybenzyl esters of the phosphonic acids, the acid-catalyzed process appeared to provide much lower yields of the products relative to its benzyl counterpart. The method's efficiency was tested in the successful derivatization and identification of pinacolyl methylphosphonic acid (PMPA) as its benzyl ester when present at a concentration of ~ 5μg/g in a soil matrix featured in the Organisation for the Prohibition of Chemical Weapons (OPCW) 44th proficiency test (PT). Additionally, the protocol was used in the detection and identification of PMPA when spiked at ~ 10μg/mL concentration in a fatty acid-rich liquid matrix featured during the 38th OPCW-PT. The benzyl derivative of PMPA was partially corroborated with the instrument's internal NIST spectral library and the OPCW central analytical database (OCAD v.21_2019) but unambiguously identified through comparison with a synthesized authentic standard. The method's MDL (LOD) values for the benzyl and the p-methoxybenzyl pinacolyl methylphosphonic acids were determined to be 35 and 63ng/mL respectively, while the method's Limit of Quantitation (LOQ) was determined to be 104 and 189ng/mL respectively in the OPCW-PT soil matrix evaluated.

Spectral trends in GC-EI-MS data obtained from the SWGDRUG mass spectral library and literature: A resource for the identification of unknown compounds

Rapid identification of new or emerging psychoactive substances remains a critical challenge in forensic drug chemistry laboratories. Current analytical protocols are well-designed for confirmation of known substances yet struggle when new compounds are encountered. Many laboratories initially attempt to classify new compounds using gas chromatography-electron ionization-mass spectrometry (GC-EI-MS). Though there is a large body of research focused on the analysis of illicit substances with GC-EI-MS, there is little high-level discussion of mass spectral trends for different classes of drugs. This manuscript compiles literature information and performs simple exploratory analyses on evaluated GC-EI-MS data to investigate mass spectral trends for illicit substance classes. Additionally, this work offers other important aspects: brief discussions of how each class of drugs is used; illustrations of EI mass spectra with proposed structures of commonly observed ions; and summaries of mass spectral trends that can help an analyst classify new illicit compounds.

Structural modification of fentanyls for their retrospective identification by gas chromatographic analysis using chloroformate chemistry

The one-step breakdown and derivatization of a panel of nine fentanyls to yield uniquely tagged products that can be detected by Electron Ionization Gas Chromatography-Mass Spectrometry (EI-GC-MS) is presented. The method involves the treatment of the synthetic opioids with 2,2,2-trichloroethoxycarbonyl chloride (TrocCl) at 60 °C for 3 h in dichloromethane and furnishes two products from one fentanyl molecule that can be used to retrospectively identify the original opioid. Parameters that were studied and fully optimized for the method included temperature, solvent, nature of scavenging base and reaction time. One of the two resulting products from the reaction bears the trichloroethoxycarbonyl (Troc) tag attached to the norfentanyl portion of the original opioid and greatly aids in the opioid detection and identification process. The methodology has been applied to the chemical modification of a panel of nine fentanyls and in all cases the molecular ion peak for the Troc-norfentanyl product bearing the distinctive trichloroethyl isotopic signature can be clearly observed. The method’s LLOD was determined to be 10 ng/mL while its LLOQ was found to be 20 ng/mL. This methodology represents the first application of chloroformates in the chemical modification of this class of synthetic opioids that are notoriously inert to common derivatization strategies available for GC–MS analysis.

Analysis of Grape Volatiles Using Atmospheric Pressure Ionization Gas Chromatography Mass Spectrometry-Based Metabolomics.

Analysis of volatile compounds in fruits and plants can be a challenging task as they present in a large amount with structural diversity and high aroma threshold, the information on molecular ion can be very useful for compound identification. Electron ionization gas-chromatography-mass spectrometry (EI-GC-MS) which is widely used for the analysis of plant volatiles has a certain limitation providing the limited capability to characterize novel metabolites in a complex biological matrix due to hard fragmentation level. Atmospheric pressure ionization using APGC source in combination with high-resolution time-of-flight mass spectrometry (TOF-MS) provides an excellent combination of GC with high-resolution mass spectrometry. The APGC-MS approach provides several advantages over the conventional EI and CI based GC-MS techniques in metabolomics studies due to highly reduced fragmentation, which preserves molecular ion, and accurate mass measurement by HRMS allows to deduce the elemental composition of the volatile compounds. Moreover, the use of MSE mode provides spectral similarity to EI in high-energy mode which can be used for the further confirmation of metabolite identity. We describe an APGC-MS-based untargeted metabolomics approach with a case study of grape volatile compounds and the development of a spectral library for metabolite identification.

Synthesis, characterisation, detection and quantification of a novel hexyl-substituted synthetic cannabinoid receptor agonist: (S)-N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-hexyl-1H-indazole-3-carboxamide (ADB-HINACA)

Synthetic cannabinoid receptor agonists (SCRAs) are a continually evolving family of illicit drugs, with novel analogues frequently being detected. This paper reports the detection of a new N-hexyl-1H-indazole derivative, ADB-HINACA, within a herbal sample seized by law enforcement for the first time in the United Kingdom. The identity of the compound was confirmed via the synthesis of a pure ADB-HINACA reference standard and direct spectral comparison by 1H NMR and GC-EI-MS analysis. A full analytical profiling of ADB-HINACA by nuclear magnetic resonance (NMR), attenuated total reflection Fourier-transform infrared (FTIR) spectroscopy and gas chromatography-electron ionisation-mass spectrometry (GC-EI-MS) is reported and shows good concordance between the seized sample and the reference standard. A validated GC-EI-MS method for the routine quantification of the cannabinoid in seized samples (LOD:1.7 μg/mL, LOQ: 5.8 μg/mL) was also developed and using this method, the seized herbal sample was determined to contain 4.9%w/w ADB-HINACA.