Ultrafast, Selective, and Highly Sensitive Nonchromatographic Analysis of Fourteen Cannabinoids in Cannabis Extracts, Δ8-Tetrahydrocannabinol Synthetic Mixtures, and Edibles by Cyclic Ion Mobility Spectrometry-Mass Spectrometry.

The diversity of cannabinoid isomers and complexity of Cannabis products pose significant challenges for analytical methodologies. In this study, we developed a method to analyze 14 different cannabinoid isomers in diverse samples within milliseconds by leveraging the unique adduct-forming behavior of silver ions in advanced cyclic ion mobility spectrometry-mass spectrometry. The developed method achieved the separation of isomers from four groups of cannabinoids: Δ3-tetrahydrocannabinol (THC) (1), Δ8-THC (2), Δ9-THC (3), cannabidiol (CBD) (4), Δ8-iso-THC (5), and Δ(4)8-iso-THC (6) (all MW = 314); 9α-hydroxyhexahydrocannabinol (7), 9ß-hydroxyhexahydrocannabinol (8), and 8-hydroxy-iso-THC (9) (all MW = 332); tetrahydrocannabinolic acid (THCA) (10) and cannabidiolic acid (CBDA) (11) (both MW = 358); Δ8-tetrahydrocannabivarin (THCV) (12), Δ8-iso-THCV (13), and Δ9-THCV (14) (all MW = 286). Moreover, experimental and theoretical traveling wave collision cross section values in nitrogen (TWCCSN2) of cannabinoid-Ag(I) species were obtained for the first time with an average error between experimental and theoretical values of 2.6%. Furthermore, a workflow for the identification of cannabinoid isomers in Cannabis and Cannabis-derived samples was established based on three identification steps (m/z and isotope pattern of Ag(I) adducts, TWCCSN2, and MS/MS fragments). Afterward, calibration curves of three major cannabinoids were established with a linear range of 1-250 ng·ml-1 for Δ8-THC (2) (R2 = 0.9999), 0.1-25 ng·ml-1 for Δ9-THC (3) (R2 = 0.9987), and 0.04-10 ng·ml-1 for CBD (4) (R2 = 0.9986) as well as very low limits of detection (0.008-0.2 ng·ml-1). Finally, relative quantification of Δ8-THC (2), Δ9-THC (3), and CBD (4) in eight complex acid-treated CBD mixtures was achieved without chromatographic separation. The results showed good correspondence (R2 = 0.999) with those obtained by gas chromatography-flame ionization detection/mass spectrometry.

Exploring affinity between organic probes and Prussian Blue Analogues via inverse gas chromatography.

Prussian Blue Analogues (PBAs), which are characterized by their open structure, high stability, and non-toxic properties, have recently been the subject of research for various applications, including their use as electrode precursors for capacitive deionization, gas storage, and environmental purification. These materials can be readily tailored to enhance their affinity towards gases for integration with sensing devices. An improved understanding of PBA-gas interactions is expected to enhance material development and existing sensor deposition schemes greatly. The use of inverse gas chromatography (IGC) is a robust approach for examining the relationship between porous materials and gases. In this study, the adsorption properties of (functionalized) hydrocarbons, i.e., probe molecules, on the copper hexacyanoferrate (CuHCF) lattice were studied via IGC, demonstrating that alkylbenzenes have a higher affinity for this material than n-alkanes. This difference was rationalized by steric hindrance, π-π interactions, and vapour pressure effects. Along the same line, the five isomers of hexane showed decreasing selectivity upon increased steric hindrance. Enthalpy values for n-pentane, n-hexane and n-heptane were lower than that of toluene. The introduction of increased probe masses resulted in a surface coverage of 46% for toluene. For all n-alkane probe molecules this percentage was lower. However, the isotherms of these probes did not show saturation points and the observed linear regime proves beneficial for gas sensing. Our work demonstrates the versatility of CuHCF for gas sensing purposes and the potential of IGC to characterize the adsorption characteristics of such a porous nanomaterial.

Comprehensive cannabinoid profiling of acid-treated CBD samples and Δ8-THC-infused edibles.

Δ8-Tetrahydrocannabinol (Δ8-THC) is increasingly popular as a controversial substitute for Δ9-tetrahydrocannabinol (Δ9-THC) in cannabinoid-infused edibles. Δ8-THC is prepared from cannabidiol (CBD) by treatment with acids. Side products including Δ9-THC and other isomers that might end up in Δ8-THC edibles are less studied. In this paper, three orthogonal methods, namely reversed-phase (RP)-UHPLC-DAD/HRMS, normal-phase/argentation (silica-Ag(I))-HPLC-DAD/MS, and GC-FID/MS were developed for analysis of cannabinoid isomers, namely Δ8-THC, Δ9-THC, CBD, Δ8-iso-THC, Δ(4)8-iso-THC, and hydrated THC isomers. Eight acid-treated CBD mixtures contained various amounts of Δ8-THC (0-89%, w/w%), high levels of Δ9-THC (up to 49%), Δ8-isoTHC (up to 55%), Δ(4)8-iso-THC (up to 17%), and three hydrated THC isomers. Commercial Δ8-THC gummies were also analyzed, and issues like overclaimed Δ8-THC, excessive Δ9-THC, undeclared Δ8-iso-THC, and Δ(4)8-iso-THC were found. These findings highlight the urgency of improving regulations towards converting CBD to Δ8-THC for use as food ingredients.

Rapid Distinction and Semiquantitative Analysis of THC and CBD by Silver-Impregnated Paper Spray Mass Spectrometry.

The control over the amount of psychoactive THC (Δ-9-tetrahydrocannabinol) in commercial cannabidiol (CBD) products has to be strict. A fast and simple semiquantitative Ag(I)-impregnated paper spray mass spectrometric method for differentiating between THC and CBD, which show no difference in standard single-stage or tandem MS, was established. Because of a different binding affinity to Ag(I) ions, quasi-molecular Ag(I) adducts [THC + Ag]+ and [CBD + Ag]+ at m/z 421 and 423 give different fragmentation patterns. The product ions at m/z 313 for THC and m/z 353 and 355 for CBD can be used to distinguish THC and CBD and to determine their ratio. Quantification of THC/CBD ratios in commercial CBD oils was accomplished with a low matrix effect (-2.2 ± 0.4% for THC and -2.0 ± 0.3% for CBD). After simple methanol extraction (recovery of 87.3 ± 1.2% for THC and 92.3 ± 1.4% for CBD), Ag(I)-impregnated paper spray analysis was employed to determine this ratio. A single run can be completed in a few minutes. This method was benchmarked against the UHPLC-UV method. Ag(I)-impregnated paper spray MS had the same working range (THC/CBD = 0.001-1) as UHPLC-UV analysis (R2 = 0.9896 and R2 = 0.9998, respectively), as well as comparable accuracy (-2.7 to 14%) and precision (RSD 1.7-11%). The method was further validated by the analysis of 10 commercial oils by Ag(I)-impregnated paper spray MS and UHPLC-UV analysis. Based on the determined relative concentration ratios of THC/CBD and the declared CBD concentration, 6 out of 10 CBD oils appear to contain more THC than the Dutch legal limit of 0.05%.

Laser Ablation Electrospray Ionization Hydrogen/Deuterium Exchange Ambient Mass Spectrometry Imaging.

Identification and confirmation of known as well as unknown (bio)chemical entities in ambient mass spectrometry (MS) and MS imaging (MSI) mostly involve accurate mass determination, often in combination with MS/MS or MSn work flows. To further improve structural assignment, additional molecular information is required. Here we present an ambient hydrogen/deuterium exchange (HDX) laser ablation electrospray ionization (LAESI) MS method in which, apart from the accurate mass and MS/MS data, the number of exchangeable protons in (un)known molecules is obtained. While eventually presenting ambient HDX-LAESI-MSI, samples were not preincubated with deuterated solvents, but instead HDX occurred following fusion of ablated sample material with microdroplets generated by ESI of deuterated solvents. Therefore, the degree of HDX was first studied following ablation of nondeuterated sample solutions of melamine and monosaccharides. From these experiments, it was concluded that the set-up used could provide meaningful HDX data in support of molecular structure elucidation by significantly reducing the number of structure options from a measured elemental composition. This reduction was demonstrated with an unknown accurate m/z value obtained in the analysis of an orange slice, reducing the possible number of molecular structures having the same elemental composition by 87% due to the number of H/D exchanges observed. Next, deuterated and nondeuterated MS/MS experiments showed the number of exchangeable protons in the substructures from deuterated neutral losses in the product ion spectra, confirming the compound to be arginine. Finally, the potential of ambient HDX-LAESI-MSI was demonstrated by the imaging of (secondary) plant metabolites in a Phalaenopsis petal.

Ambient surface analysis of organic monolayers using direct analysis in real time Orbitrap mass spectrometry.

A better characterization of nanometer-thick organic layers (monolayers) as used for engineering surface properties, biosensing, nanomedicine, and smart materials will widen their application. The aim of this study was to develop direct analysis in real time high-resolution mass spectrometry (DART-HRMS) into a new and complementary analytical tool for characterizing organic monolayers. To assess the scope and formulate general interpretation rules, DART-HRMS was used to analyze a diverse set of monolayers having different chemistries (amides, esters, amines, acids, alcohols, alkanes, ethers, thioethers, polymers, sugars) on five different substrates (Si, Si3N4, glass, Al2O3, Au). The substrate did not play a major role except in the case of gold, for which breaking of the weak Au-S bond that tethers the monolayer to the surface, was observed. For monolayers with stronger covalent interfacial bonds, fragmentation around terminal groups was found. For ester and amide-terminated monolayers, in situ hydrolysis during DART resulted in the detection of ions characteristic of the terminal groups (alcohol, amine, carboxylic acid). For ether and thioether-terminated layers, scission of C-O or C-S bonds also led to the release of the terminal part of the monolayer in a predictable manner. Only the spectra of alkane monolayers could not be interpreted. DART-HRMS allowed for the analysis of and distinction between monolayers containing biologically relevant mono or disaccharides. Overall, DART-HRMS is a promising surface analysis technique that combines detailed structural information on nanomaterials and ultrathin films with fast analyses under ambient conditions.

Rapid and simple neurotoxin-based distinction of Chinese and Japanese star anise by direct plant spray mass spectrometry.

Ingestion of products containing Chinese star anise (Illicium verum) fruits contaminated or adulterated with Japanese star anise (Illicium anisatum) fruits can cause poisoning due to the neurotoxin anisatin that is present in Japanese star anise. Thus a rapid, simple and unambiguous distinction between the morphologically similar Chinese star anise and toxic Japanese star anise fruits is important for guaranteeing food safety. After adding ~200 µL of methanol to one star anise carpel placed at 7-10mm from the inlet of a mass spectrometer and applying a potential of ~5 kV to the carpel, an electrospray is created. The formation of the electrospray is immediate, robust and stable and lasts for at least a minute. The presence or absence of anisatin could be monitored by orbitrap high resolution mass spectrometry (HRMS) in negative mode by observing the [M-H](-) ion at m/z 327.1074 (C15H19O8) or in positive mode the [M+K](+) ion at m/z 367.079 (C15H20KO8). Several parameters like wetting solvent, voltage, distance and set-up were optimised. The anisatin signal was ~250 times higher in Japanese than in Chinese star anise. An existing Direct Analysis in Real Time (DART) HRMS for anisatin was used for benchmarking. Alternatively a linear ion trap mass spectrometer could be used in negative selective reaction monitoring (SRM) mode albeit with lower selectivity than the HRMS method. The transition of the [M-H](-) ion at m/z 327 to the fragment at m/z 265 was monitored. Direct plant spray and DART ionisation are both robust and provided the same yes/no answer in seconds without any prior sample preparation. Compared with the DART-HRMS procedure, the direct plant spray method is simpler in terms of equipment, yields a more stable signal, does not require heating of the sample but is slightly less selective and requires working with high voltages.

Ambient mass spectrometry of covalently bound organic monolayers.

Detailed molecular analysis by Direct Analysis in Real Time High Resolution Mass Spectrometry (DART-HRMS) of ester and amide-terminated monolayers is demonstrated. The structural information obtained allowed monitoring of the progress of a 4-step surface modification.

Rapid control of Chinese star anise fruits and teas for neurotoxic anisatin by Direct Analysis in Real Time high resolution mass spectrometry.

After ingestion, products containing Chinese star anise (Illicium verum) contaminated or adulterated with Japanese star anise (Illicium anisatum) or other Illicium species, can cause epilepsy, hallucinations, and nausea due to the rare neurotoxic sesquiterpene dilactone anisatin that is present in Japanese star anise. Thus a rapid, simple and unambiguous method for distinguishing between the morphologically similar Chinese star anise and toxic Japanese star anise is important for food safety issues. Direct Analysis in Real Time (DART) ambient ionisation coupled with orbitrap high resolution mass spectrometry allowed the recording of mass spectra of anisatin in solid star anise fruits in seconds without any prior sample pretreatment. Spectra could be obtained in both positive ([M+NH(4)](+) at m/z 346.1496, C(15)H(24)NO(8)) and negative mode ([M-H](-) at m/z 327.1074, C(15)H(19)O(8)) and gave the same outcome provided a mass resolution of at least 27,000 is available. The anisatin signal was typically >1000 times larger in Japanese star anise than in Chinese star anise thus allowing an unequivocal qualitative determination. Herbal teas containing star anise fragments too small to be visually recognised, could be analysed by preparing a tea in 6 min and subsequently sampling ∼2 µL of tea on a glass rod. None of the 8 investigated retail teas contained significant quantities of anisatin. Spiking a complex herbal tea containing Chinese star anise with an equally concentrated tea prepared from Japanese star anise provided a linear calibration curve (R(2) ≥ 0.995) after normalising on a native constituent of Chinese star anise (standard addition method). This showed that adulteration down to 1% (w/w) is still measurable. Compared with existing PCR, TLC, GC-MS and HPLC-ESI-MS/MS procedures, the proposed DART-HRMS procedure is faster and simpler and moreover measures the actual biotoxin.

Comprehensive two-dimensional liquid chromatography with ultraviolet, evaporative light scattering and mass spectrometric detection of triacylglycerols in corn oil.

An improved comprehensive two-dimensional (LC x LC) HPLC system for the analysis of triacylglycerols was developed. In the first-dimension, a Ag(I)-coated cation exchanger (250 mm x 2.1 mm, 5 microm) was employed with a gradient from 100% MeOH to 6% MeCN in MeOH at 20 microL/min. Using a 10-way valve with two switching loops, 1 min sections of the first-dimension were introduced in the second-dimension consisting of a 30 mm x 4.6 mm C18 (1.8 microm) column with an isocratic mobile phase of methanol-methyl tert-butyl ether (70:30) at 3.0 mL/min. As the second-dimension solvent was stronger than the first-dimension solvent, focusing in the second-dimension took place, leading to better separations than in previously reported analyses in which hexane was the main constituent of the first-dimension eluent. Compounds differing by 2 in their partition number were baseline separated in the second-dimension. Detection took place by UV at 210 nm, evaporative light scattering and (+)-atmospheric pressure chemical ionisation-MS with the latter giving the best results. Corn oil was investigated and 44 compounds could be detected: 34 triacylglycerols (TAGs), 8 oxygenated TAGs, and 2 TAGs containing a trans double bond. Data manipulation allowed the construction of contour plots and the automated calculation of the first- and second-dimension retention times and peak areas. Quantitative results are compared with a fatty acid methyl ester analysis, and with literature data.

Fungal biotransformation products of dehydroabietic acid.

Dehydroabietic acid (DHA) (1) is one of the main compounds in Scots pine wood responsible for aquatic and microbial toxicity. The degradation of 1 by Trametes versicolor and Phlebiopsis gigantea in liquid stationary cultures was followed by HPLC-DAD-ELSD. Both fungi rapidly degraded DHA relative to a control. More breakdown products were observed for T. versicolor than for P. gigantea. After 13 days, four compounds were identified by means of spectroscopic methods in P. gigantea cultures: 1beta-hydroxy-DHA (2), 1beta,7alpha-dihydroxy-DHA (3), 1beta,16-dihydroxy-DHA (5), and tentatively 1beta-hydroxy-7-oxo-DHA (4). In T. versicolor cultures, 1beta,16-dihydroxy-DHA (5), 7beta,16-dihydroxy-DHA (6), 1beta,7beta,16-trihydroxy-DHA (7), 1beta,16-dihydroxy-7-oxo-DHA (8), 1beta,15-dihydroxy-DHA (9), and 1beta,7alpha,16-trihydroxy-DHA (10) were identified after 9 days of incubation. Thus the biotransformation of 1 by the two fungi was different, with only 5 being produced by both strains. Compounds 3, 7, 8, and 10 are reported for the first time as natural products.

Fungal bio-treatment of spruce wood with Trametes versicolor for pitch control: influence on extractive contents, pulping process parameters, paper quality and effluent toxicity.

Lipophilic low molar-mass constituents in wood chips for the paper industry result in low quality pulp, pitch deposition, and effluent toxicity. New biotechnological solutions such as fungal pre-treatment of wood chips can reduce pitch problems. This laboratory-scale study focuses on the potential and limitations of a fungal bio-treatment of Norway spruce chips with the white-rot fungus Trametes versicolor. Different fungal treatment conditions were compared. A 4-week fungal treatment reduced the concentration of resin acids and triglycerides by 40% and 100%, respectively, but neither lowered the energy requirements of the TMP process nor significantly affected the morphological fiber characteristics and the physical pulp properties. The pre-treatment led to slightly poorer optical properties. The Trametes versicolor fungal treatment contributed to a less toxic effluent and improved the biodegradability. A treatment of 2-3 weeks appears optimal.

Development of an on-line high performance liquid chromatography detection system for human cytochrome P450 1A2 inhibitors in extracts of natural products.

An on-line HPLC screening method for detection of inhibitors of human cytochrome P450 1A2 in extracts was developed. HPLC separation of extracts is connected to a continuous methoxyresorufin-O-demethylation (MROD) assay in which recombinant human P450 1A2 converts methoxyresorufin to its fluorescent metabolite resorufin. The system was tested with three P450 1A2 inhibitors, for which minimum detectable amounts (MDA) ranging from 0.7 to 9.5 ng were obtained. Analysis of a kava kava and a basil extract showed that the on-line system is applicable to complex mixtures, since in both extracts, peaks with P450 1A2 inhibiting activity were observed.