The pharmacokinetics of the semisynthetic cannabinoid hexahydrocannabinol (HHC) and its 11-nor-carboxy- as well as 11-hydroxy-metabolites has already been the subject of research in various recent publications. However, further metabolism studies showed the presence of other, especially (side-chain-)hydroxylated metabolites in urine and serum. This study aimed to investigate the pharmacokinetics of these metabolites after oral (25 mg fruit gum) and inhalative (three puffs from vape) consumption, using serum and urine samples from a previous study. Serum (up to 48 h) and urine (up to five days) samples of six participants (three per consumption group) were collected at different time points and analyzed by LC-QqTOF and LC-QqLIT. Three metabolites (M1: side-chain-hydroxylated; M2: hydroxylated on alicyclic moiety; M3: 8-OH-HHC) were further investigated. The peak area ratios of analytes and internal standards were plotted against the time after consumption to obtain pharmacokinetic information. Overall, this study proved the presence of these metabolites in biological samples and provided pharmacokinetic insights for the first time. Differences between serum and urine, oral and inhalative consumption as well as inter-individual differences were observed. The investigated metabolites could partially serve as unambiguous consumption markers for HHC, as they are unlikely to be formed in vivo from corresponding Δ9-tetrahydrocannabinol (THC) metabolites.

The semi-synthetic cannabinoid hexahydrocannabinol (HHC) has become a highly discussed topic in forensic toxicology since 2022 due to its legal availability at this time and its psychoactive effects. This study aimed to investigate the pharmacokinetics, effects, and immunological detectability of HHC after oral (25 mg HHC fruit gum) and inhalative (three puffs from HHC vape) consumption with three participants per group. Serum (up to 48 h), urine (up to five days), and saliva (up to 48 h) samples were collected at different relevant time points and analyzed by HPLC-MS/MS for (9R)/(9S)-HHC, 11-hydroxy-HHC, and (9R)/(9S)-HHC carboxylic acid with a fully validated method. Additionally, immunological detectability was investigated with three different commercially available tests. To address the psychoactive effects, the subjective “high” feeling (scale 0–10) was monitored and different psychophysical tests (e.g. modified Romberg test, walk and turn) were conducted. Overall, the pharmacokinetics and effects of HHC were comparable to tetrahydrocannabinol (THC). However, the route of administration as well as inter-individual factors played a crucial role regarding maximum concentrations, pharmacokinetic profiles, and psychoactive effects.

Im Jahr 2024 wurde in Deutschland der Cannabiskonsum teillegalisiert, ein gesetzlich geltender Tetrahydrocannabinol(THC)-Grenzwert von 3,5 ng/ml Blutserum im Straßenverkehrsgesetz (StVG) verankert sowie das semisynthetische Cannabinoid Hexahydrocannabinol (HHC) in das Neue-psychoaktive-Stoffe-Gesetz (NpSG) aufgenommen. Ziel dieser Studie ist, erste Aussagen über die Auswirkungen dieser Gesetzesänderungen aus forensisch-toxikologischer Sicht zu treffen. Es erfolgte eine retrospektive Datenauswertung der in den entsprechenden Zeiträumen vorliegenden Daten aus der Region Südwestsachsen hinsichtlich der Blutkonzentrationen von THC, 11-Hydroxy-THC (11-OH-THC), THC-Carbonsäure (THC-COOH), (9R)- und (9S)-HHC. Die nachgewiesenen THC-, 11-OH-THC- als auch THC-COOH-Konzentrationen sind seit der Teillegalisierung im Vergleich zum gleichen Zeitraum des Jahres 2023 signifikant höher, was insbesondere in einem höheren Anteil an regelmäßigen Cannabiskonsumenten begründet scheint. Die Erhöhung des THC-Grenzwertes auf 3,5 ng/ml führt zu einer Nichtahndung von Ordnungswidrigkeiten in 27,2

Background In 2024 cannabis consumption was partially legalized in Germany, a tetrahydrocannabinol (THC) limit of 3.5 ng/ml blood serum was established in the German Road Traffic Act (StVG) and the semisynthetic cannabinoid hexahydrocannabinol (HHC) was added to the German New Psychoactive Substances Act (NpSG). The aim of this study was to provide initial insights into the impact of these legislative changes from a forensic toxicological perspective. Methods A retrospective data analysis was conducted evaluating available data from the region of Southwest Saxony during the relevant time periods. The focus was laid on the blood concentrations of THC, 11-hydroxy-THC (11-OH-THC), THC carboxylic acid (THC-COOH), (9R)-HHC and (9S)-HHC. Results Since the partial legalization, the detected concentrations of THC, 11-OH-THC and THC-COOH have been significantly higher compared to the same period in 2023. This appears to be primarily due to a higher proportion of regular cannabis users. The increase of the THC limit to 3.5 ng/ml resulted in the nonprosecution of administrative offenses in 27.2% of cases compared to the previous threshold of 1.0 ng/ml. Since HHC was classified under the NpSG and in connection with the partial legalization of cannabis, its consumption has significantly declined. Discussion These initial findings clearly demonstrate that the legislative changes implemented last year have led to shifts in both cannabis consumption patterns and the prosecution of administrative offenses. Further analyses from other regions in Germany are necessary to potentially provide recommendations for corrective measures.

Introduction: Δ9-tetrahydrocannabinolic acid A (THCA-A) is one of the main ingredients of cannabis plants and is converted to the psychoactive substance Δ9-tetrahydrocannabinol (THC) by decarboxylation during heating above ∼90°C. During the consumption of cannabis, a varying proportion of THCA-A is absorbed into the body. Therefore, the quantification of THCA-A in serum/plasma might provide additional information on consumption behavior in driving under the influence of cannabis cases. Materials and Methods: In this study, an already established gas-chromatography mass-spectrometry (GC-MS) method for the quantification of THC, 11-OH-THC, and THC-COOH in serum and plasma samples was extended to include THCA-A. This validated method was then applied to 1228 routinely achieved serum/plasma samples from drivers suspected of cannabis consumption in Western Saxony. Two different grouping systems for chronic/occasional consumption, one system for acute/subacute consumption, Huestis formulas, and the cannabis influence factor (CIF) were used for evaluation. Results: Method validation showed appropriate results for forensic toxicological routine analysis. Limit of detection and lower limit of quantification (LLOQ) for THCA-A were 0.3 and 1.0 ng/mL, respectively. Reproducibility was <11% and accuracy ranged between 104% and 107%. THCA-A was stable in native samples at least for 2 weeks at room temperature or 4°C as well as 1 month at -20°C. Freeze-thaw stability for three cycles and processed sample stability over 3 days was proven. A total of 865 cases with a THC concentration above the German analytical cutoff of 1 ng/mL as well as the analytical LLOQs of 0.9 and 2.5 ng/mL for 11-OH-THC and THC-COOH, respectively, were included in further statistical analysis. In 407 (47.1%) of these samples, THCA-A was quantifiable. Different statistical analyses indicated a correlation between THCA-A and THC concentrations in cases of chronic and acute consumption. In addition, an increase of chronic and acute cases with increasing THCA-A concentrations was observed. However, no correlation between THCA-A and CIF was found. Discussion: These data show that THCA-A might be an additional indicative marker to provide information about consumption frequency and acuteness. Additional studies with known consumption frequencies and times are required to verify these findings.

Background: Natural organic reduction (NOR), called Reerdigung in Germany, is a new form of burial in which a human body is turned into soil after 40 days. For the scientific forensic evaluation, the Institute of Forensic Medicine Leipzig examined samples from the remains of the process.Methods: Bone, soil and hair samples were taken from two deceased persons and examined using available molecular biological, toxicological, morphological, osteological and pedological methods.Results: The DNA in the bones degraded after 40 days and was no longer detectable by conventional methods in the ground-up state added to the soil. The toxicological detection of premortally ingested drugs was partially possible, depending on the analyte and matrix. Morphologically and osteologically this corresponds to a lying time of 20-50 years. The resulting soil has the characteristics of humus.Discussion: The new form of burial requires a second post-mortem examination before the process begins. From a forensic toxicological point of view, the isolated detectability of active substances tends to be assessed positively, as in contrast to cremation, potentially necessary toxicological follow-up examinations could still be possible in some cases. For the soil samples before and after mixing of soil and bone meal, no DNA quantities suitable for DNA analysis could be obtained. It can therefore be assumed that no human DNA can be obtained from mixed samples after burial of the remains in the event of further decomposition. An identification is not possible. The soil samples show no traces of human soft tissue, neither microscopically nor macroscopically. The result suggests that the transformation to soil takes place within 40 days.The low number of corpses examined in this pilot study is a limitation. Further studies to increase the database are planned.

Zusammenfassung Hintergrund Die Reerdigung stellt eine neue Bestattungsform dar, bei der ein menschlicher Leichnam durch natürliche und mechanische Prozesse in 40 Tagen zu Erdsubstrat umgewandelt wird. Für die forensische Evaluation untersuchte das Institut für Rechtsmedizin Leipzig Proben aus den Überresten des Prozesses. Methodik Knochen‑, Erd- und Haarproben wurden von zwei Verstorbenen entnommen und mit molekularbiologischen, toxikologischen, morphologisch-osteologischen und bodenkundlichen Methoden untersucht. Ergebnisse Die DNA in den Knochen ist nach 40 Tagen degradiert und im zermahlenen, der Erde beigemischten Zustand nicht mehr mit herkömmlichen Methoden nachweisbar. Ein toxikologischer Nachweis von prämortal eingenommenen Medikamenten war analyt- und matrixabhängig teilweise möglich. Morphologisch-osteologisch entsprechen die Knochen einer Liegezeit von 20–50 Jahren. Die entstehende Erde weist die Eigenschaften von Humus auf. Diskussion Die neue Bestattungsform bedarf vor Beginn des Prozesses einer zweiten Leichenschau. Aus forensisch-toxikologischer Sicht ist die vereinzelte Nachweisbarkeit von Wirkstoffen tendenziell positiv zu bewerten, da im Gegensatz zur Kremierung potenziell nötige toxikologische Nachuntersuchungen teilweise noch möglich sein könnten. Für die Erdprobe vor und nach Durchmischung von Erde und Knochenmehl konnten hier keine für eine DNA-Analyse geeigneten DNA-Mengen gewonnen werden. Es ist somit davon auszugehen, dass nach der Bestattung der Überreste bei einer weiteren Zersetzung keine humane DNA aus Mischproben zur Identifizierung gewonnen werden kann. Die Erdproben weisen neben den Knochen mikro- sowie makroskopisch keine Spuren von humanem Weichgewebe auf. Das Resultat spricht dafür, dass innerhalb von 40 Tagen die Umwandlung zu Erde stattfindet. Die geringe Zahl der in dieser Pilotstudie untersuchten Leichen stellt eine Limitation dar. Weitere Untersuchungen zur Vergrößerung der Grundgesamtheit sind geplant.

Hypoxia and low glucose abundance often occur simultaneously at sites of inflammation. In monocytes and macrophages, glucose-oxygen deprivation stimulates the assembly of the NLRP3 inflammasome to generate the proinflammatory cytokine IL-1β. We found that concomitant glucose deprivation and hypoxia activated the NLRP3 inflammasome by constraining the function of HMG-CoA reductase (HMGCR), the rate-limiting enzyme of the mevalonate kinase pathway. HMGCR is involved in the synthesis of geranylgeranyl pyrophosphate (GGPP), which is required for the prenylation and lipid membrane integration of proteins. Under glucose-oxygen deprivation, GGPP synthesis was decreased, leading to reduced prenylation of the small GTPase Rac1, increased binding of nonprenylated Rac1 to the scaffolding protein IQGAP1, and enhanced activation of the NLRP3 inflammasome. In response to restricted oxygen and glucose supply, patient monocytes with a compromised mevalonate pathway due to mevalonate kinase deficiency or Muckle-Wells syndrome released more IL-1β than did control monocytes. Thus, reduced GGPP synthesis due to inhibition of HMGCR under glucose-oxygen deprivation results in proinflammatory innate responses, which are normally kept in check by the prenylation of Rac1. We suggest that this mechanism is also active in inflammatory autoimmune conditions.

The semisynthetic cannabinoid hexahydrocannabinol (HHC) is currently getting a lot of media attention because the legal status in many countries is not clearly specified. In this study, a GC-MS method for the quantification of Δ9-tetrahydrocannabinol (THC), 11-hydroxy-Δ9-tetrahydrocannabinol (11-OH-THC), and 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH) was extended to (9R)- and (9S)-HHC. The applicability was proven by serum/plasma samples from drivers suspected of cannabis consumption. Limit of detection (LOD) and lower limit of quantification (LLOQ) were 0.15 and 0.25 ng/mL, respectively. Within-run imprecision was <6.5% and between-run imprecision was <10.0%. Inter-injection stability, processed sample stability (3 days), freeze-thaw stability (three cycles), and storage stability (1 week room temperature; 1 month 4°C, -20°C) could be proven. Both HHC diastereomers could be detected in 17 (5.3%) out of 321 analyzed samples from traffic controls in Western Saxony. The mean ratio between (9R)- and (9S)-HHC was 1.99 (CV = 14.6%). Quantification resulted in concentrations between <LLOQ and 35.35 ng/mL for (9R)-HHC and <LLOQ and 21.76 ng/mL for (9S)-HHC. Additionally, cross-reactivities of HHC, 11-hydroxy-hexahydrocannabinol (11-OH-HHC), 11-nor-9-carboxy-hexahydrocannabinol (HHC-COOH), hexahydrocannabiphorol (HHC-P), hexahydrocannabinol acetate (HHC-O), and tetrahydrocannabidiol (H4-CBD) were evaluated in five immunological screening tests for serum, urine, and saliva. Urine test strips and ELISA tests for the detection in serum seem to be beneficial to detect HHC consumption in comparison with saliva tests. HHC analogs and H4-CBD showed no cross-reactivity with any of the tests.

ZusammenfassungEin 90-jähriger Mann verstarb durch protrahiertes Verbluten aus einer, mittels Rasierklinge minimal eröffneten, oberflächlichen Beinvene über der Patella und nachfolgender Manipulation. Dabei handelte es sich wahrscheinlich um einen Ast der V. saphena accessoria anterior. Es lag zunächst eine Fehleinschätzung durch den leichenschauhaltenden Arzt vor, der von einer gastrointestinalen Blutung als Todesursache ausging. Diese Einschätzung teilten die Kriminalbeamten aufgrund der Auffindesituation nicht. Begünstigend war eine vorbestehende Varikosis. Die Einnahme von Antikoagulanzien oder eine vorbestehende oder erworbene Gerinnungsstörung konnte nicht nachgewiesen bzw. eruiert werden. Bei Vorlage eines Abschiedsbriefes und Ausschluss fremder Gewalteinwirkungen oder einer Vergiftung kann plausibel von einem ungewöhnlichen suizidalen Ereignis ausgegangen werden. Es wird über die Auffindung sowie die Ergebnisse der Sektion und umfangreicher Zusatzuntersuchungen berichtet.

Background: Human life without sperm is not possible. Therefore, it is alarming that the fertilizing ability of human spermatozoa is continuously decreasing. The reasons for that are widely unknown, but there is hope that metabolomics-based investigations may be able to contribute to overcoming this problem. This review summarizes the attempts made so far. Methods: We will discuss liquid chromatography–mass spectrometry (LC-MS), gas chromatography (GC), infrared (IR) and Raman as well as nuclear magnetic resonance (NMR) spectroscopy. Almost all available studies apply one of these methods. Results: Depending on the methodology used, different compounds can be detected, which is (in combination with sophisticated methods of bioinformatics) helpful to estimate the state of the sperm. Often, but not in all cases, there is a correlation with clinical parameters such as the sperm mobility. Conclusions: LC-MS detects the highest number of metabolites and can be considered as the method of choice. Unfortunately, the reproducibility of some studies is poor, and, thus, further improvements of the study designs are needed to overcome this problem. Additionally, a stronger focus on the biochemical consequences of the altered metabolite concentrations is also required.

A 90-year-old man died due to prolonged bleeding to death from a superficial leg vein at the patella, opened with a razor blade and subsequent manipulation. It was probably a branch of the anterior accessory saphenous vein. There was a misjudgement during the first external examination of the corpse and the physician assumed a gastrointestinal bleeding as the cause of death. Police officers at the scene did not share this opinion. A pre-existing varicosis was beneficial. The use of anticoagulants or hints for a pre-existing or acquired coagulation disorder could not be proven or determined. As a farewell letter was present and no third-party violence or an intoxication was involved, an extraordinary suicidal event can be reasonably assumed. The findings at the scene and autopsy results with extensive additional investigations are reported.

Currently, spermiogram analysis is the most relevant method used to clarify the potential infertility of a couple. However, in some cases, the reasons for infertility remain obscure. Smoking is among the factors that have been described to adversely affect male fertility. Smoking increases oxidative stress and thus promotes various pathological processes. Comparative studies, particularly those on metabolomic changes in sperm and seminal plasma caused by smoking, have not yet been published. Thus, the present pilot study aimed at the mass spectrometric characterization of the metabolomes of specimens from both smoking and nonsmoking subjects and the comparison of the evaluated data in terms of sperm apoptosis and spermiogram parameters. The results provided evidence that the conventional spermiogram is not altered in smokers compared to nonsmokers. However, a more careful investigation of sperm cells by metabolomic profiling reveals profound effects of smoking on sperm: first, nitrogen oxide synthase, a marker of oxidative stress, is activated. Second, the uptake of fatty acids into sperm mitochondria is reduced, leading to an impaired energy supply. Third, phenylalanine hydroxylation and tryptophan degradation, which are both indications of altered tetrahydrobiopterin biosynthesis, are reduced. Moreover, flow cytometry approaches indicated increased sperm caspase-3 activity, a sign of apoptosis. The present study clearly shows the negative effects of smoking on semen quality. Especially for idiopathic cases, metabolomic profiling can help to shed light on male subfertility or infertility.

In 50% of all infertility cases, the male is subfertile or infertile, however, the underlying mechanisms are often unknown. Even when assisted reproductive procedures such as in vitro fertilization and intracytoplasmic sperm injection are performed, the causes of male factor infertility frequently remain elusive. Since the overall activity of cells is closely linked to their metabolic capacity, we analyzed a panel of 180 metabolites in human sperm and seminal plasma and elucidated their associations with spermiogram parameters. Therefore, metabolites from a group of 20 healthy donors were investigated using a targeted LC-MS/MS approach. The correlation analyses of the amino acids, biogenic amines, acylcarnitines, lysophosphatidylcholines, phosphatidylcholines, sphingomyelins and sugars from sperm and seminal plasma with standard spermiogram parameters revealed that metabolites in sperm are closely related to sperm motility, whereas those in seminal plasma are closely related to sperm concentration and morphology. This study provides essential insights into the metabolome of human sperm and seminal plasma and its associations with sperm functions. This metabolomics technique could be a promising screening tool to detect the factors of male infertility in cases where the cause of infertility is unclear.

Mesenchymale Stromalzellen (MSC) konnten zur Therapie der durch MCD-Diät induzierten NASH im Mausmodell erfolgreich angewendet werden. Allerdings ist der Wirkungsmechanismus bisher nicht geklärt. Es sollte daher untersucht werden, inwieweit MSC in der NASH-Leber auf metabolische Veränderungen wirkten.

Im Mausmodell der durch eine Methionin-Cholin-freie Diät induzierten Nicht-alkoholischen Steatohepatitis verminderten mesenchymale Stromalzellen (MSC) die Leberverfettung.