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Forensic analysis of hallucinogenic mushrooms and khat (Catha edulis FORSK) using cation-exchange liquid chromatography
Abstract
Hallucinogenic mushrooms (e.g. Psilocybe and Panaeolus species) as well as leaves and young shoots of the khat tree (Catha edulis Forsk) are illicit drugs in many countries. The exact concentration of the hallucinogenic alkaloids psilocin and psilocybin in mushrooms and the sympathomimetic alkaloids cathinone and cathine in khat is usually essential for jurisdiction. Facing an increasing number of mushroom and khat seizures by German customs authorities, a convenient comprehensive quantitative HPLC method based on cation-exchange liquid chromatography for these rather "exotic" drugs has been developed which avoids time-consuming multi-step sample preparation or chemical derivatization procedures. Using this method a number of different hallucinogenic fungi species and products that are mainly distributed via the internet have been analysed (dried and fresh Psilocybe cubensis Singer as well as P. cubensis collected from "grow boxes", Panaeolus cyanescens Berkeley and Broome and so-called "philosopher stones" (sclerotia of Psilocybe species)). Highest total amounts of psilocin have been detected in dried P. cyanescens reaching up to 3.00+/-0.24 mg per 100 mg. The distribution of khat alkaloids in different parts of the khat shoots has been studied. High concentrations of cathinone have not only been detected in leaves but also in green parts and barks of stalks. Additionally, the sample treatment for fresh mushroom and khat samples has been optimised. Highest amounts of alkaloids were found when fresh material was freeze-dried.
... Many forensic toxicology laboratories use gas chromatography (GC) or GC-mass spectrometry (MS) to detect psilocin and psilocybin in hallucinogenic mushroom [12], blood [13] or urine [14] samples after derivatization. High-performance liquid chromatography (HPLC)-ultraviolet (UV) detection has also been used to analyse psilocin and psilocybin in hallucinogenic mushrooms [7,15] and plasma [9]. In addition, some validated liquid chromatography (LC)-MS/MS methods were used to analyse psilocin and psilocybin in hallucinogenic mushroom [16], urine [17] and blood [6,11] samples. ...
... In recent years, hallucinogenic mushrooms have often been sold in their dried form, ground powder or capsules. At present, there have been some reports in the literature using LC-MS or GC-MS to analyse the content and distribution of psilocin and psilocybin in hallucinogenic mushrooms [15]. The psilocin content in six hallucinogenic mushrooms was determined to be lower than 12.67 ng/mg by HPLC combined with the fluorescence derivatization method [4]. ...
... At the next study, we have studied the distribution of psilocin and psilocybin in the canopies, stems and hyphae of hallucinogenic mushrooms themselves. Since such studies had been well conducted [12,15,16], we compared our results with the published data as shown in Tables 5 and 6. Our results and those in the previous studies [7,12] equally indicated that the concentration of psilocybin in the stems and hyphae of hallucinogenic mushrooms are lower than in the canopy. ...
Purpose
In this study, we developed a very sensitive method for quantitative analysis of psilocin and psilocybin in hair samples of magic mushroom consumers.
Methods
The analyses were performed with pretreatments of samples, followed by ultra-high pressure liquid chromatography (LC) connected to a Q-Trap type tandem mass spectrometry (MS/MS). For LC, mobile phase (A) consisted of 0.1% formic acid in water, and mobile phase (B) was acetonitrile for gradient elution using a Acquity™ UPLC HSS T3 column. For MS/MS, electrospray ionization measurements in positive selected reaction monitoring mode were used.
Results
The calibration curves were linear from 5 to 500 pg/mg ( r > 0.99) and no selectivity problems occurred. The limit of detection was 1 pg/mg, and the lower limit of quantitation was 5 pg/mg. The ranges of the matrix effects and recovery rates were 90.4–107% and 76.0–102%, respectively.
Conclusions
The concentrations of psilocin in two authentic hair were 161 and 150 pg/mg, respectively, and psilocybin was not detected from both samples. This method was also used to analyze the distribution of psilocin and psilocybin in seven hallucinogenic mushrooms. To our knowledge, this is the first demonstration of psilocin concentrations in hair samples of hallucinogenic mushroom consumers, and also our method is most sensitive for quantitative analysis of psilocin and psilocybin in hair samples.
... Parmi les champignons hallucinogènes confisqués en Allemagne, les spécimens appartenant à cette espèce possèdent la plus forte concentration en psilocybine, soit jusque 3 mg pour 100 mg de matière sèche dans certains échantillons (Musshoff et al, 2000 ;Laussmann et Meier-Giebing, 2010). ...
... Les champignons contenant de la psilocybine sont les plus consommés parmi les champignons hallucinogènes (Koçak et al, 2010). Parmi eux, le genre Psilocybe est le plus utilisé (Laussmann et Meier-Giebing, 2010). ...
... Dans les pays proches de la France comme l'Allemagne, d'autres espèces sont aussi consommées comme Psilocybe cubensis ou Stropharia coronilla, ou encore d'autres Psilocybe (Musshoff et al, 2000 ;Sticht et Kaferstein, 2000). Panaeolus cyanescens est aussi apparu sur le marché allemand (Laussmann et Meier-Giebing, 2010). Il contient le plus fort taux de substances psychoactives parmi les espèces confisquées, comme cela a été dit plus haut (Musshoff et al, 2000). ...
Panaeolus cinctulus et Panaeolus cyanescens sont les deux champignons possédant un caractère hallucinogène certain et poussant en Normandie. Rarement trouvés, ils contiennent de la sérotonine, mais surtout de la psilocybine et de la psilocine leur conférant des propriétés hallucinogènes. Ces composés appartenant à la famille des alcaloïdes indoliques sont dosés par diverses méthodes chromatographiques. Les panéoles bleuissants possèdent d’ailleurs de forts taux de psilocybine. Après absorption, cette molécule est transformée en psilocine, psychodysleptique. Elle induit des perturbations sensorielles, mais aussi des modifications profondes de la pensée et de l’humeur dont certaines demeurent après l’élimination de la psilocine. Le mécanisme d’action de la psilocine, agoniste des récepteurs sérotoninergiques 5-HT2A principalement, n’est que partiellement élucidé. La psilocine agit notamment au niveau des circuits corticaux et module la transmission de nombreux neuromédiateurs. Certaines études portent sur l’utilisation de la psilocybine pour explorer et soigner certaines pathologies psychiatriques et sur son intérêt dans la prise en charge de certaines addictions. Tous ces développements thérapeutiques potentiels sont limités par la difficulté d’entreprendre des études cliniques mesurant son efficacité dans de tels domaines. La psilocybine est consommée dans un cadre festif. Les champignons hallucinogènes, placés sous la réglementation des substances stupéfiantes, sont souvent considérés comme une drogue « naturelle » et donc inoffensive, mais des accidents mortels ont été reportés. Les confusions avec des espèces plus toxiques présentent des risques plus grands encore.
... In fact it was demonstrated that cathinone is stable for years in the dried khat, but it undergoes rapid decomposition in the fresh or frozen vegetable material [9]. Several analytical methods were developed for the determination of khat alkaloids either involving GC and HPLC techniques [10][11][12][13][14]; in the last years, the Tribunal of Busto Arsizio and Bergamo gave us the task to analyze all the materials suspected to contain khat active principles, coming from the seizures at the Malpensa and Orio al Serio airports in northern Italy. Thus, due to the high number of samples to be analyzed, we developed a fast, effective and reliable GC analytical procedure able to simultaneously detect, cathine and PPA, whose discrimination was mandatory because PPA is not a controlled psychoactive agent, exploiting a derivatization protocol which allowed an effective separation of the two components without dramatically increasing the time of analyses [15]. ...
... From the analyses of the active principles in the various parts of the vegetable material it could be noticed that in the leaves all the three alkaloids were detected, whereas in the stems only cathinone and cathine were present, being PPA detectable only in traces. This finding confirmed what was reported in the literature [11,15]. In the woody stems only low amounts of cathinone were found confirming the hypothesis that the most active alkaloid is enzymatically converted into cathine and PPA in the older parts of the plant ( Table 1). ...
... In bundle 1 the quantity of the active principles were very low, perhaps due to a bad conservation of the vegetable material before coming to our laboratory. In fact generally dried khat contains from 0.9 to 4.5 mg/g of cathinone [1,[9][10][11]. In bundles 2 and 3 the content of the active principles was superior and followed in the range reported in the literature. These bundles were also analyzed by means of GC/FID technique, confirming the quantity found by CE [19]. ...
... It showed that the concentration of cathinone was highest in the leaves and in the younger parts of the plant. [21] The main active principles of C. edulis have been determined in various seized samples using GC-MS after derivatization of the alkaloids. [22] The crude methanolic extracts of fresh khat leaves have been investigated by LC-MS/MS, and this study resulted in the identification of 62 alkaloids derived from cathedulin. ...
... However, according to our results, this effect is due to the higher level of cathinone in stem and not in leaf as mentioned in some previous studies. [21,30] Beside cathinone and cathine which are the main active components in khat, several other compounds were characterized by HR DART-ToF-MS in the different fresh samples. The peak at m/z 177 could correspond to merucathine with the formula C 11 H 15 NO which was mentioned in several previous papers, as well as N-formylnorephedrine (C 10 H 13 NO 2 ) observed at m/z 180. ...
... The most developed and utilized hallucinogenic toxin would be psilocybin which is also the main psychedelic ingredient of hallucinogenic mushrooms (Guzm an et al., 1998;Laussmann and Meier-Giebing, 2010). Since it was first isolated and identified in the 1960s, psilocybin was widely used for experimental research of mental disorders and psychotherapy. ...
Mushrooms have long been considered as delicacies as well as used as important dietary supplements and food. However, there are major concerns with poisonous mushrooms as these pose threats to public health and safety. In this paper, we provide a review focusing on poisonous mushrooms, their toxins, symptoms and utilizations. In addition, this paper establishes a poisonous mushroom list which includes 643 species from two phyla, 16 orders, 51 families and 148 genera. The toxicity of all these species was verified and 332 species were ranked as P1 signifying that these species have toxic studies and or clinical poisoning case records and 311 species were P2 meaning they had previously been recorded as poisonous in other studies. Furthermore, we discuss advances in technology including how genomic studies could be used as a breakthrough tool in the field of toxic mushrooms. With this comprehensive review, we aim to promote public awareness of poisonous mushrooms, including how to avoid mushroom poisoning, and how to better utilize poisonous mushroom resources.
... In this regard, MAE achieved in our study a maximum yield of 1.609 ± 0.081 mg/g relative alkaloids (1.255 ± 0.098 mg/g psilocin and 0.354 ± 0.085 mg/g psilocybin). This extraction yield is greater than the mean yields reported by previous studies where UAE was the extraction method employed; Gotvaldová et al. obtained 0.54 mg/g psilocybin and 0.25 mg/g psilocin by vortexing dried mushroom samples [28], Laussmann et al. reported 1.151 ± 0.228 mg/g psilocybin and 0.126 ± 0.066 mg/g psilocin by UAE [35] and Tsujikawa et al. reported even lower yields of 0.835 ± 0.465 mg/g psilocybin and 0.280 ± 0.140 mg/g psilocin [36] when two UAE cycles were employed. ...
Hallucinogenic fungi, mainly those from the Psilocybe genus, are being increasingly consumed even though there is no control on their culture conditions. Due to the therapeutic potential as antidepressants and anxiolytics of the alkaloids that they produce (psilocin and psilocybin), some form of control on their production would be highly recommended. Prior to identifying their optimal culture condition, a methodology that allows their study is required. Microwave-assisted extraction method (MAE) is a technique that has proven its efficiency to extract different compounds from solid matrices. For this reason, this study intends to optimize a MAE method to extract the alkaloids found in Psylocibe cubensis. A surface-response Box–Behnken design has been employed to optimize such extraction method and significantly reduce time and other resources in the extraction process. Based on the Box–Behnken design, 50 °C temperature, 60% methanol as extraction solvent, 0.6 g:10 mL sample mass:solvent ratio and 5 min extraction time, were established as optimal conditions. These mild conditions, combined with a rapid and efficient UHPLC analysis result in a practical and economical methodology for the extraction of psilocin and psilocybin from Psylocibe cubensis.
... The total amount of psilocybin truffles taken by each participant was recorded, and a sample of the truffles was taken to determine concentrations of psilocybin and its metabolite, psilocin. The German Central Customs Authority determined the contents of psilocin and psilocybin after freezedrying the truffles using a previously described HPLC method (Laussmann and Meier-Giebing 2010). ...
Anxiety disorders are the most common type of psychiatric disorders among Western countries. Evidence-based treatment modalities including pharmacological and cognitive-behavioral therapy result in relatively low response rates (average range: 51 - 58%). Historical and recent research suggests psychedelic drugs may be efficacious in alleviating anxiety-related symptoms among healthy and clinical populations. The main aim of the present study was investigation of the effects of psilocybin-containing truffles, when taken in a supportive group setting, on ratings of state and trait anxiety across self-reported healthy volunteers. Attendees of psilocybin ceremonies were asked to complete a test battery at three separate occasions: before the ceremony (baseline), the morning after, and one week after the ceremony. The test battery included questionnaires assessing state and trait anxiety (State-Trait Anxiety Inventory), mindfulness capacities (Five Facet Mindfulness Questionnaire), and personality (Big Five Inventory). Additionally, the psychedelic experience was quantified with the Persisting Effects Questionnaire and the Ego Dissolution Inventory. The total amount of psilocybin-containing truffles consumed by each participant was recorded, and a sample of the truffles was analyzed to determine psilocin concentrations. Fifty-two attendees (males= 25; females= 25; others= 2) completed parts of the baseline assessment, 46 (males= 21; females= 24; others= 1) completed assessments the morning after the ceremony, and 23 (males= 10; females= 13) completed assessments at the one-week follow-up. Average psilocin consumption across individuals was 27.1 mg. We observed medium to large reductions in anxiety measures (both state and trait) compared to baseline which persisted over a one-week period post-ceremony. At one week post-ceremony, the non-judging facet of the mindfulness scale was increased, while the personality trait neuroticism decreased, when compared to baseline. Additionally, we found neuroticism and ratings of ego dissolution to be the strongest predictors of reductions in trait and state anxiety, respectively. In sum, results indicate rapid and persisting (up to one week) anxiolytic effects in psilocybin retreat attendees, which are related to the acute experience of ego dissolution, as well as lasting changes in trait neuroticism. To understand whether these effects extend to wider populations suffering from heightened anxiety, and the mechanisms involved, further experimental research is needed.
... However, gas chromatography is not recommended for the quantification of tryptamines because of their poor volatility and heat lability. [28][29][30] In our study, ultra-high performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry (UHPLC-MS/MS) was employed to investigate the stability of five tryptamine alkaloids in the biomass of cultivated fruiting bodies of the psychoactive species Psilocybe cubensis. [31][32][33] 2 | MATERIALS AND METHODS ...
Psilocybin, psilocin, baeocystin, norbaeocystin, and aeruginascin are tryptamines structurally similar to the neurotransmitter serotonin. Psilocybin and its pharmacologically active metabolite psilocin in particular are known for their psychoactive effects.
These substances typically occur in most species of the genus Psilocybe (Fungi, Strophariaceae). Even the sclerotia of some of these fungi known as “magic truffles” are of growing interest in microdosing due to them improving cognitive function studies. In addition to microdosing studies, psilocybin has also been applied in clinical
studies, but only its pure form has been administrated so far. Moreover, the determination of tryptamine alkaloids is used in forensic analysis.
In this study, freshly cultivated fruit bodies of Psilocybe cubensis were used for monitoring stability (including storage and processing conditions of fruiting bodies). Furthermore, mycelium and the individual parts of the fruiting bodies (caps, stipes,
and basidiospores) were also examined. The concentration of tryptamines in final extracts was analyzed using ultra-high-performance liquid chromatography coupled with mass spectrometry. No tryptamines were detected in the basidiospores, and only psilocin was present at 0.47 wt.% in the mycelium. The stipes contained approximately half the amount of tryptamine alkaloids (0.52 wt.%) than the caps (1.03 wt.%); however, these results were not statistically significant, as the concentration of tryptamines in individual fruiting bodies is highly variable. The storage conditions showed that the highest degradation of tryptamines was seen in fresh mushrooms
stored at −80°C, and the lowest decay was seen in dried biomass stored in the dark at room temperature.
... Scientific literature reports a series of analytical methodologies to measure khat active principles in plant and in biological matrices, some of these methods are listed in the Table 1 and are discussed below. Laussmann et al. [79] developed an HPLC method based on cation-exchange liquid chromatography to analyse alkaloids in khat. Before the extraction, they applied different methods of sample pretreatment to prevent degradation and find high amount of alkaloids. ...
New psychoactive substances (NPS) can be divided into two main groups: synthetic molecules and active principles of natural origin. With respect to this latter group, a wide range of alkaloids contained in plants, mainly from Asia and South America, can be included in the class of NPS of natural origin. The majority NPS of natural origin presents stimulant and/or hallucinogenic effects (e.g. Catha edulis and Ayahuasca, respectively) while few of them show sedative and relaxing properties (e.g. kratom). Few information is available in relation to the analytical identification of psychoactive principles contained in the plant material. Moreover, to our knowledge, scarce data are present in literature, about the characterization and quantification of the parent drug in biological matrices from intoxication and fatality cases. In addition, the metabolism of natural active principles has not been yet fully investigated for most of the psychoactive substances from plant material. Consequently, their identification is not frequently performed and produced metabolites are often unknown. To fill this gap, we reviewed the currently available analytical methodologies for the identification and quantification of NPS of natural origin in plant material and, whenever possible, in conventional and non-conventional biological matrices of intoxicated and dead subjects. The psychoactive principles contained in the following plants were investigated: Areca catechu, Argyreia nervosa, Ayahuasca, Catha edulis, Ipomoea violacea, Mandragora officinarum, Mitragyna speciosa, Pausinystalia yohimbe, Piper methisticum, Psilocybe, Rivea corymbosa, Salvia divinorum, Sceletium tortuosum, Lactuca virosa. From the results obtained, it can be evidenced that although several analytical methods for the simultaneous quantification of different molecules from the same plants have been developed and validated, a comprehensive method to detect active compounds from different natural specimens both in biological and non-biological matrices is still lacking.
... Somemes, bodies are buried at remote locaons to hide Table 3: Chromatographic and spectroscopic methods used to detect various fungal species from the forensic (mushroom) samples. [34] [53][54][55][56][57][58][59][60][61][62][63][64] [65][66][67][68] [21,66] [44] [69] [70][71][72][73][74][75] ...
... The total amount of psilocybin truffles taken by each participant was recorded, and a sample of the truffles was taken to determine concentrations of psilocybin and its metabolite, psilocin. The German Central Customs Authority determined the contents of psilocin and psilocybin after freeze-drying the truffles using a previously described HPLC method (Laussmann and Meier-Giebing 2010). ...
Creative thinking and empathy are crucial for everyday interactions and subjective well-being. This is emphasized by studies showing a reduction in these skills in populations where social interaction and subjective well-being are significantly compromised (e.g., depression). Anecdotal reports and recent studies suggest that a single administration of psilocybin can enhance such processes and could therefore be a potential treatment. However, it has yet to be assessed whether effects outlast acute intoxication. The present study aimed to assess the sub-acute effects of psilocybin on creative thinking, empathy, and well-being. Participants attending a psilocybin retreat completed tests of creative (convergent and divergent) thinking and empathy, and the satisfaction with life scale on three occasions: before ingesting psilocybin (N = 55), the morning after (N = 50), and seven days after (N = 22). Results indicated that psilocybin enhanced divergent thinking and emotional empathy the morning after use. Enhancements in convergent thinking, valence-specific emotional empathy, and well-being persisted seven days after use. Sub-acute changes in empathy correlated with changes in well-being. The study demonstrates that a single administration of psilocybin in a social setting may be associated with sub-acute enhancement of creative thinking, empathy, and subjective well-being. Future research should test whether these effects contribute to the therapeutic effects in clinical populations.
... Psilocybin is the main psychedelic ingredient of mushrooms of the genus Psilocybe [38] that has been used both in the lay scene, as in various studies for the treatment of anxiety and obsessive compulsive disorder [32,39]. Ritual use of psilocybin- containing mushrooms across the globe dates back millennia and is still regionally practiced today. ...
... Somemes, bodies are buried at remote locaons to hide Table 3: Chromatographic and spectroscopic methods used to detect various fungal species from the forensic (mushroom) samples. [34] [53][54][55][56][57][58][59][60][61][62][63][64] [65][66][67][68] [21,66] [44] [69] [70][71][72][73][74][75] ...
... Psilocybin is the main psychedelic ingredient of mushrooms of the genus Psilocybe [38] that has been used both in the lay scene, as in various studies for the treatment of anxiety and obsessive compulsive disorder [32,39]. Ritual use of psilocybin-250 containing mushrooms across the globe dates back millennia and is still regionally practiced today. ...
Introduction:
Evidence based treatment for Substance use disorders (SUD) includes psychotherapy and pharmacotherapy. However, these are only partially effective. Hallucinogens, such as psilocybin, may represent potential new treatment options for SUD. This review provides a summary of (human) studies on the putative therapeutic effects of psilocybin, and discusses the receptor systems, brain regions and cognitive and emotional processes mediating psilocybin's effects. Psilocybin's chemical structure is similar to that of serotonin. Dysregulations in the serotonin system are associated with alterations in stress hormones, such as cortisol, and mood disorders. After psilocybin administration cortisol levels spike and activate the executive control network, with subsequent increased control over emotional processes, and relief of negative thinking and persistent negative emotions. Preliminary data of ongoing alcohol and smoking addiction studies in humans shows promising effects of psilocybin administration on substance use. Importantly, psilocybin has a low risk of toxicity and dependence and can be used safely under controlled clinical conditions.
Areas covered:
This paper is a narrative review based on the search terms: psilocybin, substance use disorder, addiction, depression, serotonin. Literature on potential efficacy and mechanisms of action of psilocybin in SUD is discussed. Expert commentary: Recent positive findings with psilocybin need confirmation in well-designed placebo controlled randomized trials employing a large sample size.
... These techniques have been applied in pharmacological, phytochemical, forensic and law enforcement environments and were preceded by extensive sample preparation protocols. Owing to the complexity of the Khat composition, tedious procedures involving several liquid-liquid extractions (LLE) have been repeatedly employed for the determination of Khat alkaloids in the plant extracts (Gambaro et al., 2012;Geisshusler and Brenneisen, 1987;Krizevski et al., 2007Krizevski et al., , 2008Laussmann and Meier-Giebing, 2010;Ripani et al., 1996;Roda et al., 2013). Cleanup is generally conducted using organic solvents under alkaline conditions. ...
A molecularly imprinted polymer (MIP) was prepared using (-)-norephedrine as the template, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the cross-linker and chloroform as the porogen. The MIP was used as a selective sorbent in the molecularly imprinted solid-phase extraction (MIP-SPE) of the psychoactive phenylpropylamino alkaloids, norephedrine and its analogues, cathinone and cathine, from Khat (Catha edulis Vahl. Endl.) leaf extracts prior to HPLC-DAD analysis. The MIP was able to selectively extract the alkaloids from the aqueous extracts of Khat. Loading, washing and elution of the alkaloids bound to the MIP were evaluated under different conditions. The clean baseline of the Khat extract obtained after MIP-SPE confirmed that a selective and efficient sample clean-up was achieved. Good recoveries (90.0-107%) and precision (RSDs 2.3-3.2%) were obtained in the validation of the MIP-SPE-HPLC procedure. The content of the three alkaloids in Khat samples determined after treatment with MIP-SPE and a commercial Isolute C18 (EC) SPE cartridge were in good agreement. These findings indicate that MIP-SPE is a reliable method that can be used for sample pre-treatment for the determination of Khat alkaloids in plant extracts or similar matrices and could be applicable in pharmaceutical, forensic and biomedical laboratories. This article is protected by copyright. All rights reserved.
... β-blockers, using a mobile phase containing methanol (MeOH) and ammonium formate with trifluoroacetic acid at pH 2.45 (Law and Appleby, 1996), trazodone in a mobile phase system containing acetonitrile (MeCN) and ammonium phosphate adjusted to pH 6.0 (Li-Boa et al., 2014), MeOH with sodium phosphate buffer adjusted to pH 4.8 with triethylamine (Ghosheh et al., 2000), MeOH and ammonium perchlorate (Morgan et al., 2003); often mixtures of MeCN and ammonium formate or acetate as additives to mobile phase were used ( Jiang et al., 2011;Koseki et al., 2005). Ephedrine alkaloids in dietary supplements were determined on SCX column with mobile phase containing MeCN, sodium phosphate buffer at pH 3.0 (Niemann and Gay, 2003) and alkaloids from hallucinogenic mushrooms with mobile phase containing EtOH and phosphate buffer with the addition of NaCl (Laussmann and Meier-Giebing, 2010). ...
Ion exchange chromatography, an alternative to reversed-phase (RP) chromatography, is described in this paper. We aimed to obtain optimal conditions for the separation of basic drugs because silica-based RP stationary phases show silanol effect and make the analysis of basic analytes hardly possible. The retention, separation selectivity, symmetry of peaks and system efficiency were examined in different eluent systems containing different types of buffers at acidic pH and with the addition of organic modifiers: methanol and acetonitrile. The obtained results reveal a large influence of the salt cation used for buffer preparation and the type of organic modifier on the retention behavior of the analytes. These results were also compared with those obtained on an XBridge C18 column. The obtained results demonstrated that SCX stationary phases can be successfully used as alternatives to C18 stationary phases in the separation of basic compounds. The most selective and efficient chromatographic systems were applied for the quantification of some psychotropic drugs in fortified human serum samples. Copyright © 2015 John Wiley & Sons, Ltd.
Copyright © 2015 John Wiley & Sons, Ltd.
... Convention on Psychotropic Substances [14][15][16]. The stimulant effect of khat is more potent (because of cathinone) when the leaves are still fresh, and 48 h after being harvested, the stimulant elements in khat convert into a less active potent (cathine), which also has an amphetamine-like action [3,[17][18][19]. ...
The chewing of Khat leaves, a natural psychoactive substance is widely chewed in countries of East Africa and the southern Arabian Peninsula, and is reported to be associated with a range of unfavorable health outcomes including khat dependence. The impact of Khat chewing on Health Related Quality of Life is yet to be explored. Aims: to measure and compare the quality of life of the khat chewers and non-khat chewers using a short form health survey (SF36), and to assess factors associated with Khat chewing using SF36 in a sample of adult population in Jazan region, Kingdom of Saudi Arabia.
A total of 630 participants from two independent male populations of khat chewers and non-khat chewers were recruited into a cross-sectional survey study. A self administrative survey based on the SF-36 questionnaire was used to collect data on measures of health-related quality of life (HRQoL). Socioeconomic data of the respondents were also collected for detailed analysis. Data analysis include: descriptive statistics, reliability tests (Cronbach's alpha and intraclass correlation coefficient), and bivariate analysis (Chi square and Mann-Whitney U-test) to compare HRQoL of Khat chewers and non-Khat chewers.
The odds of being a khat chewer were higher in respondents with a lower socioeconomic status. The SF-36 scores were significantly lower in all domains for respondents with khat chewing, indicating that non-khat chewers had higher health perceptions compared with those chewing khat. The overall mean score of HRQoL for non-khat chewers was 92.7% (SD 5.53) compared with 63.5% (SD 21.73) for the khat chewing group. The study had shown good internal consistency and reproducibility across the eight subscales of SF-36 questionnaire (α 0.74-0.95). The Mann-Whitney U-test showed a significant difference between khat chewers and non-khat chewers (P < 0.001).
This study measured and compared the quality of life of khat chewers and non-khat chewers using a generic health survey (SF36). The study had shown that khat chewing is associated with lower quality of life (HRQoL) and lower socioeconomic status. However in future a more refined SF36 developed especially for Khat chewers can provide more useful information.
... Convention on Psychotropic Substances141516. The stimulant effect of khat is more potent (because of cathinone) when the leaves are still fresh, and 48 h after being harvested, the stimulant elements in khat convert into a less active potent (cathine), which also has an amphetamine-like action [3,171819. ...
Abstract
Background
The chewing of Khat leaves, a natural psychoactive substance is widely chewed in countries of East Africa and the southern Arabian Peninsula, and is reported to be associated with a range of unfavorable health outcomes including khat dependence. The impact of Khat chewing on Health Related Quality of Life is yet to be explored. Aims: to measure and compare the quality of life of the khat chewers and non-khat chewers using a short form health survey (SF36), and to assess factors associated with Khat chewing using SF36 in a sample of adult population in Jazan region, Kingdom of Saudi Arabia.
Methods
A total of 630 participants from two independent male populations of khat chewers and non-khat chewers were recruited into a cross-sectional survey study. A self administrative survey based on the SF-36 questionnaire was used to collect data on measures of health-related quality of life (HRQoL). Socioeconomic data of the respondents were also collected for detailed analysis. Data analysis include: descriptive statistics, reliability tests (Cronbach's alpha and intraclass correlation coefficient), and bivariate analysis (Chi square and Mann-Whitney U-test) to compare HRQoL of Khat chewers and non-Khat chewers.
Results
The odds of being a khat chewer were higher in respondents with a lower socioeconomic status. The SF-36 scores were significantly lower in all domains for respondents with khat chewing, indicating that non-khat chewers had higher health perceptions compared with those chewing khat. The overall mean score of HRQoL for non-khat chewers was 92.7% (SD 5.53) compared with 63.5% (SD 21.73) for the khat chewing group. The study had shown good internal consistency and reproducibility across the eight subscales of SF-36 questionnaire (alpha 0.74-0.95). The Mann-Whitney U-test showed a significant difference between khat chewers and non-khat chewers (P < 0.001).
Conclusions
This study measured and compared the quality of life of khat chewers and non-khat chewers using a generic health survey (SF36). The study had shown that khat chewing is associated with lower quality of life (HRQoL) and lower socioeconomic status. However in future a more refined SF36 developed especially for Khat chewers can provide more useful information.
... The extraction procedures [16] are described in detail in the supplementary information S1. ...
This case report describes the analysis of AM-2201 in plant material and its metabolites in human urine obtained from an operator of a motor vehicle in the United States. The samples were taken from the driver because of his illegal driving activities and, his subsequent erratic behaviour. The AM-2201 was extracted from a sample of plant material by sonicating it in methanol, after which an aliquot was taken and diluted with aqueous phosphate buffer (pH 6) and extracted by solid-phase extraction using a C8/aminopropyl SPE cartridge. The cartridges were washed, dried, and eluted with ethyl acetate- methanol solvent system. The urine sample was hydrolyzed with β-glucuronidase at pH 6.8 before being diluted with aqueous phosphate buffer (pH 6) and extracted with the same type of SPE cartridge. After evaporation of the eluates, the samples were dissolved in mobile phase for analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Analysis of the plant material determined the concentration to be 0.05% (AM2201) by mass of dry material. The concentration of AM-2201 (AM2201-N-(4-hydroxypentyl ) metabolite in the urine was found to be 3.1 ng/ml. The urine also contained 109 ng/ml of delta-9-tetrahydrocannabinol carboxylic acid but no other drugs including JWH-018 metabolites. Copyright © 2013 John Wiley & Sons, Ltd.
... Psilocybin and psilocin, the main psychedelic ingredients of hallucinogenic mushrooms (Guzman et al., 1998;Laussmann and Meier-Giebing, 2010) (Table 1), have recently been given a lot of attention as a research tools (Geyer and Vollenweider, 2008) as well as a potential therapeutic agents (Grob et al., 2011;Moreno et al., 2006;Sewell et al., 2006). History of the ritual use of hallucinogenic mushrooms dates back 3000 years in Mexico and regionally its use is still conventional practice today (Carod-Artal, 2011;Hofmann, 2005). ...
Psilocybin, a psychoactive alkaloid contained in hallucinogenic mushrooms, is nowadays given a lot of attention in the scientific community as a research tool for modeling psychosis as well as due to its potential therapeutic effects. However, it is also a very popular and frequently abused natural hallucinogen. This review summarizes all the past and recent knowledge on psilocybin. It briefly deals with its history, discusses the pharmacokinetics and pharmacodynamics, and compares its action in humans and animals. It attempts to describe the mechanism of psychedelic effects and objectify its action using modern imaging and psychometric methods. Finally, it describes its therapeutic and abuse potential.
... Thus, due to the high number of samples, in the past a fast, effective and reliable analytical procedure able to detect cathinone directly after maceration and cathine and PPA, after derivatization was developed [11]. To this end, HPLC and cation-exchange liquid chromatography methods have been reported in the literature [12,13], but because of our expertise in applying GC techniques, we adopted a GC/MS method for qualitative determinations and a GC/FID combined system for quantitative analysis, which are endowed with high specificity and sensitivity in the characterization of drugs of abuse. On the other hand, this method suffered from some limitations: the derivatizing agent, i.e. cyclohexanone [14,15] was effective only for the determination of cathine and PPA, while cathinone was directly determined on the methanol extraction solution. ...
The GC method previously reported by our research group for the analysis of the active principles of Catha edulis, i.e. cathine, cathinone and phenylpropanolamine, was considerably improved. N-methyl-N-trimethylsilyl-trifluoroacetamide (MSTFA) as derivatizing agent was employed, thus allowing an accurate determination of the analytes and a suitable internal standard for quantitative analyses (nicotinamide) was introduced. Moreover the chromatographic conditions were carefully studied to improve the separation of the alkaloids and sensitivity. To this end different chromatographic capillary columns and temperature gradients were investigated. The optimized GC method was validated and resulted adequate for the application in forensic analysis. Finally on behalf of the Tribunal, C. edulis vegetable material seized by the police in northern Italy was analyzed, the quantity of cathine ranging from 0.095 to 0.29%, the quantity of PPA from 0.010 to 0.21% and the quantity of cathinone from 0.025 to 0.374% of the weight of the vegetable material.
Esta revisión bibliográfica explora el papel de los hongos alucinógenos, específicamente el género Psilocybe de la familia Hymenogastraceae, en el desarrollo evolutivo de la conciencia humana. A través de un enfoque multidisciplinario que abarca biología, etnobotánica y neurociencia, se examina la influencia de compuestos como la psilocibina y la psilocina en los consumidores, revelando su potencial para desencadenar efectos neurológicos y psicológicos significativos. Se explora el impacto en la conciencia humana, señalando cambios perceptuales, cognitivos y emocionales. Se analiza el desarrollo evolutivo y antropológico de los seres humanos, explorando cómo la disponibilidad de alimentos, el uso de la psilocibina y las adaptaciones fisiológicas han podido influir en el éxito reproductivo y la supervivencia a lo largo del tiempo.
In forensic case work, fungal growth is occasionally observed on decomposed cadavers, however, the isolation of these organisms is not always carried out routinely. The usefulness of investigating fungi isolated from corpses in forensic settings, has been reported, such as providing trace evidence, estimating the time since death, and ascertaining the time of deposition. A 32-year-old male missing for 4 months, was found in a car in a forest far from his home and had died from blood loss, having been stabbed with a kitchen knife in the left side of his chest. Whitish or white greenish colonies ranging 5 cm to 8 cm in size were observed over his whole body. Penicillium commune and Cladosporium cladosporioides were isolated from the colonies. A 49-year-old male missing for one month, and was found dead at 500 m elevation in the mountains. An autopsy was conducted, but the cause of death could not be determined. Dark greenish or whitish colonies were found on his face and neck, and Epicoccum nigrum, Mucor sp.. Cladosporium sp., and Pestalotiopsis sp. were isolated. Penicillium sp. and Cladosporium sp. are major indoor fungi, and Epicoccum sp. and Pestalotiopsis sp. are plant pathogens found in outdoor environments. There was no discrepancy between the police statement and findings of the corpse such as place discovered, types of isolated fungi, and estimated time since death. Identifying fungi on cadavers may aid in forensic casework, and further studies are needed to develop for the use of fungi as a forensic tool.
Fungi with hallucinogenic properties and neurotoxicity have been listed as prohibited drugs in recent years, but there is a lack of in situ quantification of psilocybin and analogues in these samples to avoid the decomposition of these psychoactive tryptamines in time-consuming sample preparation. In this study, matrix-assisted laser-desorption/ionization (MALDI)-Fourier transform ion cyclotron resonance (FT ICR) mass spectrometric imaging (MSI) was used to analyze the distribution of psilocybin and its analogues in hallucinogenic Psilocybe mushrooms. A cesium chloride (CsCl)-coated target plate was prepared to improve the detection sensitivity and reduce the interference of other compounds or decomposition products with very similar m/z values in MALDI-FT ICR MS analysis. Psilocybin and other tryptamines with structurally similar compounds, including psilocin, baeocystin, tryptophan, tryptamine, and aeruginascin, were identified and imaged in the psilocybe tissue section; the semiquantitative analysis of the distribution of psilocybin was also investigated using a homemade 75-well CsCl-coated plate; and the target plate can be placed on the mass spectrometry target carrier along with the indium-tin oxide (ITO) conductive slide, which can simultaneously carry out matrix vapor deposition, thus ensuring the parallelism between the standards and samples in the pretreatment experiment and MSI. The contents of psilocybin and its analogues in the psilocybe tissue section can be evaluated from the color changes corresponding to different concentration standard curves. Furthermore, a comprehensive comparison between MALDI-FT ICR MS and ultra-performance liquid chromatography-quadrupole time of flight mass spectrometry (UPLC-Q/TOF MS) analysis was performed for quantification and validation. This study reduces the decomposition in time-consuming sample pretreatment and provides a powerful tool for drug abuse control and forensic analysis.
Anxiety disorders are the most common type of psychiatric disorders among Western countries. Evidence-based treatment modalities including pharmacological and cognitive-behavioral therapy result in deficient treatment responses. Historical and recent research suggests psychedelic drugs may be efficacious in alleviating anxiety-related symptoms among healthy and clinical populations. The main aim of the present study was investigation of the effects of psilocybin-containing truffles, when taken in a supportive group setting, on ratings of state and trait anxiety across self-reported healthy volunteers. Attendees of psilocybin ceremonies were asked to complete a test battery at three separate occasions: before the ceremony (baseline), the morning after, and 1 week after the ceremony. The test battery included questionnaires assessing state and trait anxiety (State-Trait Anxiety Inventory), mindfulness capacities (Five Facet Mindfulness Questionnaire), and personality (Big Five Inventory). Additionally, the psychedelic experience was quantified with the Persisting Effects Questionnaire and the Ego Dissolution Inventory. The total amount of psilocybin-containing truffles consumed by each participant was recorded, and a sample of the truffles was analyzed to determine psilocin concentrations. Fifty-two attendees (males = 25; females = 25; others = 2) completed parts of the baseline assessment, 46 (males = 21; females = 24; others = 1) completed assessments the morning after the ceremony, and 23 (males = 10; females = 13) completed assessments at the 1-week follow-up. Average psilocin consumption across individuals was 27.1 mg. The morning after the ceremony, we observed medium reductions in anxiety measures (both state and trait) compared to baseline (d¯ = 6.4; p < 0.001 and d¯ = 6; p = 0.014, respectively), which persisted over a 1-week period post-ceremony (d¯ = 6.7; p = 0.001 and d¯ = 8.6; p = 0.004, respectively). At 1 week post-ceremony, the non-judging facet of the mindfulness scale was increased (d¯ = 1.5; p = 0.03), while the personality trait neuroticism decreased (d¯ = 5.2; p = 0.005), when compared to baseline. Additionally, we found ratings of ego dissolution (mean: 59.7, SD: 28.3) and changes in neuroticism to be the strongest predictors of reductions in state and trait anxiety, respectively. In sum, results suggest rapid and persisting (up to 1 week) anxiolytic effects in individuals with sub-clinical anxiety symptoms, which are related to the acute experience of ego dissolution, as well as lasting changes in trait neuroticism. Results also add support to the feasibility and potential efficacy of group sessions with psychedelics. To understand whether these effects extend to wider populations suffering from heightened anxiety, and the mechanisms involved, further experimental research is needed.
Unusual forms of hallucinogenic mushrooms are emerging, which may delay diagnosis and compromise optimal management. Here, we discuss clinical and biological findings in a case of “magic mushroom” intoxication in a 20-year-old man who was brought to hospital emergency department in a state of euphoria, disorientation, intense visual hallucinations, episodic amnesia and agitation requiring physical restraint. The patient’s family brought intact specimens of the consumed product, without any health professional being able to identify it. Toxicological screening detected psilocin in urine and in the unknown mushrooms, allowing guiding the mushroom identification towards the Psilocybes genus. Ten hours post-ingestion, the patient was discharged asymptomatic. We believe that recognition of the unusual form of hallucinogenic mushroom presented here by emergency physicians might contribute to better diagnosis and subsequent optimal management, especially since hallucinogenic substances are not routinely tested in toxicological analyses.
Contemporary wildlife trade is massively facilitated by the Internet. By design, the dark web is one layer of the Internet that is difficult to monitor and lacks thorough investigation. Here, we accessed a comprehensive database of dark web marketplaces to search across c. 2 million dark web advertisements over 5 years using c. 7k wildlife trade-related search terms. We found 153 species traded in 3,332 advertisements (c. 600 advertisements per year). We characterized a highly specialized wildlife trade market, where c. 90% of dark-web wildlife advertisements were for recreational drugs. We verified that 68 species contained chemicals with drug properties. Species advertised as drugs mostly comprised of plant species, however, fungi and animals were also traded as drugs. Most species with drug properties were psychedelics (45 species), including one genera of fungi, Psilocybe, with 19 species traded on the dark web. The native distribution of plants with drug properties were clustered in Central and South America. A smaller proportion of trade was for purported medicinal properties of wildlife, clothing, decoration, and as pets. Our results greatly expand on what species are currently traded on the dark web and provide a baseline to track future changes. Given the low number of advertisements, we assume current conservation and biosecurity risks of the dark web are low. While wildlife trade is rampant on other layers of the Internet, particularly on e-commerce and social media sites, trade on the dark web may increase if these popular platforms are rendered less accessible to traders (e.g., via an increase in enforcement). We recommend focussing on surveillance of e-commerce and social media sites, but we encourage continued monitoring of the dark web periodically, to evaluate potential shifts in wildlife trade across this more occluded layer of the Internet.
Introduction For ages; people in Faifa Mountains (Jazan region, Kingdome of Saudi Arabia) cultivate Khat tree and the men chew their leaves. Many adverse effects are documented among men chewing Khat for a long time especially on the central nervous system, heart and liver due to the action of sympathetic-like substances that Khat contains. Till today in Faifa; there is no effective intervention that could replace Khat-chewing habit and leads to its withdrawal. Objectives: The goal of this study is to assess the competencies of women in their families to help men in withdrawal of Khat-chewing habit. This will be examined through the specific objectives to investigate the knowledge, attitudes and behaviours of women towards Khat-chewing habit that practiced by men in their families. Methodology: This research is designed to assess the competencies; knowledge, attitudes and behaviours of women in Faifa mountains towards Khat-chewing habit among the men in their families and its effect on the family social and economic status. It is performed according to qualitative research principles using in-depth interviews through semi-structured questionnaire. This study is performed on a purposive sample composed of 20 women above 15 years old having men chewing Khat in their families and residing in Faifa Mountains. Results: The most prominent findings in this study are that 65% of the women believe that Chewing Khat is not forbidden. However, 95% of them are sure that there are adverse effects due to this habit summarized as liver and heart disease. Although 85% of the investigated women are looking for withdrawing this habit and described some interventions to stop this habit; 80% of them feel that they do not have a role in stopping this habit. Seventy percent of women believe that this habit neither affects the work, religious behaviour of men, the financial status of the family nor the social relationships. Ninety-five percent of women do not exert any effort to stop this habit for their men expecting the resistance of men. On the other side, 95% of them educate their children about the adverse effect of this habit as it is wrong to be practiced. Conclusion: Although women realize the adverse effects of Khat chewing habit, they do not recognize it has any negative effect on their lives either socially or economically. However, they have a will to stop this habit among their men but do not exert a considerable effort for that, except giving some information to their children to avoid this habit. This may give place for further studies about how to stop the Khat chewing habit rather than influence of women on men in the Faifa Mountains
The misuse of psychoactive substances is attracting a great deal of attention from the general public. An increase use of psychoactive substances is observed among young people who do not have enough awareness of the harmful effects of these substances. Easy access to illicit drugs at low cost and lack of effective means of routine screening for new psychoactive substances (NPS) have contributed to the rapid increase in their use. New research and evidence suggest that drug use can cause a variety of adverse psychological and physiological effects on human health (anxiety, panic, paranoia, psychosis, and seizures). We describe different classes of these NPS drugs with emphasis on the methods used to identify them and the identification of their metabolites in biological specimens. This is the first review that thoroughly gives the literature on both natural and synthetic illegal drugs with old known data and very hot new topics and investigations, which enables the researcher to use it as a starting point in the literature exploration and planning of the own research. For the first time, the conformational analysis was done for selected illegal drugs, giving rise to the search of the biologically active conformations both theoretically and using lab experiments.
Psilocin (4‐hydroxy‐N,N‐dimethyltryptamine, 4‐HO‐DMT) and bufotenine (5‐hydroxy‐N,N‐dimethyltryptamine, 5‐HO‐DMT), which are both naturally occurring compounds, are classified as controlled substances in numerous countries due to their pharmacological activities and recreational usage. There are two other benzene ring regioisomers, 6‐hydroxy‐N,N‐dimethyltryptamine (6‐HO‐DMT) and 7‐hydroxy‐N,N‐dimethyltryptamine (7‐HO‐DMT), which are not classified by name as controlled substances, and which were synthesized for this current work. The four isomers were analyzed using routine methodologies employed by the Israel's Police Division of Identification and Forensic Science (DIFS) Laboratory, namely thin layer chromatography (TLC), Fourier transform infrared spectroscopy (FTIR), and gas chromatography mass spectroscopy (GC‐MS). It was found possible to differentiate the four isomers. Forensic specimens that were suspected to be psilocybe mushrooms were examined, confirming that it is now possible to unequivocally identify the presence of psilocin and rule out the presence of its other isomers.
The main psychoactive compound in Khat is cathinone which consists of two enantiomers, S-(-)-cathinone being more stimulant than its R antipode. This study aimed to the enantioseparation and determination of these two stereoisomers in different parts of fresh Catha edulis. The samples were solvent extracted and cathinone was derivatized with menthyl chloroformate. The separation of the two diastereomeric derivatives was carried out by gas chromatography and showed an excellent resolution, while their structure was confirmed by mass spectrometry. The quantitative determination of both enantiomers showed a different distribution in various investigated parts of the plant, as shown in their enantiomeric excess. Unlike the results published in some previous articles, the current study confirmed the presence of both S and R cathinone in all parts of the fresh plant. The concentration of S-cathinone was higher in stems while its values were lower in leaves. The obtained concentrations were in the ranges 0.081-0.290 and 0.087-0.211 mg/g for S and R antipodes, respectively. Also, S-cathinone which is the most psychoactive stereoisomer showed an increasing concentration from lower to upper stems of the plant. The present study is the first quantitative investigation of the two cathinone enantiomers in different parts of fresh Catha edulis.
Management of mushroom intoxications requires tremendous effort from clinicians and toxicologists. Management is challenging because of difficult identification, the potential lethality of some species, and because there are few efficacy data on different therapies. Presumptive diagnoses are established by positive identification of the suspect mushroom along with the occurrence of consistent clinical signs and clinicopathological changes. If the animal dies, pathologic lesions may suggest exposure to certain mushroom toxins, but lesions are often nonspecific. Toxicology testing is only available for a limited number of mushroom toxins; thus, many cases are not confirmed.
Psilocybe cubensis, or “magic mushroom,” is the most common species of fungus with psychedelic characteristics. Two primer sets were designed to target Psilocybe DNA using web-based software and NBCI gene sequences. DNA was extracted from eighteen samples, including twelve mushroom species, using the Qiagen DNeasy® Plant Mini Kit. The DNA was amplified by the polymerase chain reaction (PCR) using the primers and a master mix containing either a SYBR® Green I, Radiant™ Green, or LCGreen Plus® intercalating dye; amplicon size was determined using agarose gel electrophoresis. The PCR assays were tested for amplifiability, specificity, reproducibility, robustness, sensitivity, and multiplexing with primers that target marijuana. The observed high resolution melt (HRM) temperatures for primer sets 1 and 7 were 78.85 ± 0.31°C and 73.22 ± 0.61°C, respectively, using SYBR® Green I dye and 81.67 ± 0.06°C and 76.04 ± 0.11°C, respectively, using Radiant™ Green dye.
Background:
Khat (Catha edulis Forsk) is an evergreen shrub of the Celastraceae family. It is widely cultivated in Yemen and East Africa, where its fresh leaves are habitually chewed for their momentary pleasures and stimulation as amphetamine-like effects. The main psychostimulant constituents of khat are the phenylpropylamino alkaloids: cathinone, cathine and norephedrine.
Results:
In this study, simple procedures based on preparative HPLC and salting-out assisted liquid-liquid extraction (SALLE) based methods were developed respectively for large scale isolation and the extraction of psychoactive phenylpropylamino alkaloids; cathinone, cathine and norephedrine, from khat (Catha edulis Forsk) chewing leaves, a stimulant and drug of abuse plant. The three khat alkaloids were directly isolated from the crude oxalate salt by preparative HPLC-DAD method with purity > 98%. In addition, a modified (SALLE) method has been developed and evaluated for the extraction efficiency of psychoactive phenylpropylamino alkaloids from khat (Catha edulis Forsk) chewing leaves. An in situ two steps extraction protocol was followed without dispersive SPE clean up. The method involves extraction of the samples with 1% HAc and QuEChERS salt (1.0 g of CH3COONa and 6.0 g of MgSO4) followed by subsequent in situ liquid-liquid partitioning by adding ethyl acetate and NaOH solution. The optimized method allowed recoveries of 80-86% for the three alkaloids from khat sample with relative standard deviation (RSD) values less than 15% and limits of detection (0.85-1.9 μg/mL).
Conclusion:
The method was found to be simple, cost-effective and provides cleaner chromatogram with good selectivity and reproducibility. The SALLE based protocol provided as good results as the conventional extraction method (ultrasonic assisted extraction followed by solid phase extraction, UAE-SPE) and hence the method can be applicable in forensic and biomedical sectors.
Khat chewing is deeply rooted in the culture and tradition of communities in khat belt countries, and its consumption is spread to other countries through the suitcase trade. The aim of this article is to review current knowledge on the chemistry, social, pharmacology and toxicology of khat and its use. Khat produces effect invariably in every system, which is harmful or beneficial in some instances. Harmful effects are observed in heavy users, although firm evidence is lacking. Chewing khat acutely elicits states of euphoria, which is followed by low mood. Khat contains alkaloids with psychostimulant properties, but the effect cannot be totally explained by these alkaloids. It is also not clear whether the effect produced in some organs like liver could be attributed to khat or pesticides sprayed during farming. Although the evidence indicates that khat has adverse effects in most organs, our understanding of the complex interaction between use and effect is incomplete, and causal relationships have not yet been described. Moreover, khat has positioned itself well in the social, economic and political arena. Thus, a multidisciplinary research is required to understand the different dimensions and come up with ways that maximize the benefit while minimizing the risk. Copyright © 2017 John Wiley & Sons, Ltd.
The main representatives of narcotic substances of natural origin are considered. Methodological solutions used in their screening, identification, and quantitative determination in raw materials, dosage forms, and biological materials are discussed. Prospects of application of state-of-the art hybrid methods combining chromatographic separation with mass spectrometric determination are noted.
ILLANA-ESTEBAN, C. (2011). Hallucinogenic truffles. Bol. Soc. Micol. Madrid 35: 187-190. The marketing of the sclerotia of some species of the genus Psilocybe, which are called hallucinogenic truffles is commented. Its cultivation and production began in Holland a few years ago after authorities banned the marketing of fresh mushrooms of the genus Psilocybe.
History Identifying Characteristics Exposure Dose Effect Toxicokinetics Histopathology and Pathophysiology Clinical Response Diagnostic Testing Treatment References
Alkaloids - Secrets of Life: Alkaloid Chemistry, Biological Significance, Applications and Ecological Role, Second Edition provides knowledge on structural typology, biosynthesis and metabolism in relation to recent research work on alkaloids, considering an organic chemistry approach to alkaloids using biological and ecological explanation. The book approaches several questions and unresearched areas that persist in this field of research. It provides a beneficial text for academics, professionals or anyone who is interested in the fascinating subject of alkaloids. Each chapter features an abstract. Appendices, a listing of alkaloids, and plants containing alkaloids are all included, as are basic protocols of alkaloid analysis.
The aim of our chapter is to review recent developments in a group of medicinally important natural products-alkaloids, with reference to the structure-activity studies in respect of certain diseases. Alkaloids covered by our review come from mushrooms called "hallucinogenic." Hallucinogenic compounds have been chemically identified in mushrooms belonging to various genera, e.g., Agrocybe, Amanita, Conocybe, Galerina, Gymnopilus, Hypholoma, Inocybe, Panaeolus, Psilocybe, Pholiotina, Pluteus, and Weraroa [J.W. Allen, Ethnomycol. J. Sacred Mushroom Stud. 9 (2012) 130-175]. One of the largest classes of alkaloids is indole alkaloids. Indoles are probably the most widely distributed heterocyclic compounds in nature having medicinal importance [K.N. Kumar et al., Molecules 18 (2013) 6620-6662]. Two of simple indole alkaloids: psilocin (3-[2 (dimethylamino)ethyl]-4-indolol) and psilocybin ([3-(2-dimethylaminoethyl)-1H-indol-4-yl] dihydrogen phosphate) are present in most psychedelic mushrooms. Psilocin is a serotonin agonist - psilocybin/psilocin caused effects are thought to be mediated mainly by activation of 5-HT2A receptor. Ligands for the 5-HT2A receptor may be extremely useful tools for future cognitive neuroscience research [D.E. Nichols, Pharmacol. Ther. 101 (2004) 131-181]. They are also other analogs of psilocybin: baeocystin, norbaeocystin, bufotenin, and aeruginascin that are found in hallucinogenic mushrooms. Bufotenin occur also in some animal species (genus Bufo) and plants. Some of the mushroom belonging to genera Gymnopilus and Pholiota were shown to possess bisnoryangonin and hispidin, alkaloids with antimicrobial [K. Shinto et al., J. Home Econ. Jpn. 58 (9) (2007) 563-568] and antioxidant [Lee In-K et al., Mycobiology, 36 (1) (2008) 55-59] activity. Also psychoactive species of genus Amanita, contain the alkaloids (muscimol, ibotenic acid, and muscazone) that react with neurotransmitter receptors in the central nervous system. Isoxazoles, to which these alkaloids belong, have been found to inhibit voltage-gated sodium channels to control pain, enable the construction of tetracycline antibiotic derivatives, and as treatments for depression. The information within this review is intended to serve as a reference tool to better enable future research into important and fascinating area of pharmacognostic science as well as other parts of medical science.
The study demonstrated that by freezing or drying under different conditions, cathinone could be detected in the khat (Catha edulis Forsk) samples that have been harvested for more than four months. Freeze drying or freezing in the refrigerator (-20 oC) yielded more than 73% of cathinone. Air drying, sun drying or oven drying at 60 oC resulted in a cathinone composition of 57%, 42% and 36%, respectively. The study confirmed that, freshly harvested tender leaves of khat contained exclusively of cathinone but upon sun drying, part of it decomposes to cathine and norephedrine leaving cathinone as a major component. Thus it is wrong to assume that sun drying the leaves can result complete conversion of cathinone to cathine and norephedrine. Furthermore, the results of the study confirmed that it was possible to preserve khat samples for longer periods after freeze drying or deep freezing without significant loss of cathinone. KEY WORDS: Khat, Alkaloids, Cathinone, Cathine, Norephedrine, Storage Bull. Chem. Soc. Ethiop. 2013, 27(3), 347-358.DOI: http://dx.doi.org/10.4314/bcse.v27i3.3
Background:
Khat chewing, which has many adverse health and social consequences, is highly prevalent and socially accepted in the Jazan region of the Kingdom of Saudi Arabia.
Methods:
A qualitative study was conducted with 47 adult male former khat users regarding their khat initiation, continuance, and cessation, the amounts of khat they had used, and the health and social consequences of their use and cessation of use.
Results:
Participants noted a desire to show maturity, ease of availability of khat, and peer pressure as reasons for initiating khat chewing. Many noted long leisure times with little to do as a reason for continuing use. Negative consequences of khat use were seen in economic, health, familial, and sexual areas of their lives. After quitting khat use, participants saw improvements in all of these areas.
Conclusions:
A comprehensive community development program (CCDP) tackling, among other issues, the normalization of khat use, substantial leisure times with few positive activities, and misinformation about the "benefits" of khat use, as well as developing peer and family training programs to help prevent or stop khat use, would be useful to reduce khat chewing in this community.
A fast and simple procedure based on matrix solid-phase dispersion was developed for the extraction of psychoactive phenylpropylamino alkaloids; cathinone, cathine and norephedrine, from khat (Catha edulis Forsk) chewing leaves, a stimulant and drug of abuse plant. Determination of the alkaloids was carried out by high performance liquid chromatography with diode array detection. Several extraction parameters, such as type of dispersant, type and volume of elution solvent and the ratio of sample to sorbent material were evaluated and optimized. Mean recoveries ranging from 89 to 92 % with relative SD of less than 6 % were obtained. A marked diversity in the phenylpropylamino alkaloid content and composition was found in seventeen different cultivars of Ethiopian khat. ANOVA results showed the existence of significant differences between the alkaloids profiles among samples of different varieties from different geographical locations in Ethiopia. The proposed method is simpler, faster and comparably more efficient than the frequently reported maceration followed by liquid–liquid extraction but as good and efficient as ultrasonic assisted extraction followed by solid-phase extraction.
A HPLC procedure on strong cation exchange column (SCX) has been developed for the analysis of selected alkaloids from different chemical groups. The retention, separation selectivity, symmetry of peaks and system efficiency were examined in different eluent systems containing different types or concentrations of buffers at various pH and the addition of organic modifiers: methanol (MeOH), acetonitrile (CH3CN), tetrahydrofuran (THF) or dioxane (Dx). The retention factors as the function of the concentration of buffers, the mobile phase pH and the percentage of modifier in the eluents were investigated. More symmetrical peaks and the highest theoretical plate number were obtained in eluents containing acetonitrile or tetrahydrofuran. In most cases, the increase of buffer concentration caused the decrease of alkaloids' retention, the improvement of peaks' symmetry and the increase of theoretical plate number. The improved peak symmetry and the efficiency of system for most investigated alkaloids were observed in the systems containing buffers at strongly acidic pH. The obtained results also reveal a large influence of salt cation used for buffer preparation. The results obtained on SCX column were compared with those obtained on a C18 column. The most efficient and selective systems were used for the separation of alkaloid standard mixture.
: Psychoactive drugs of fungal origin, psilocin, ibotenic acid, and muscimol among them have been proposed for recreational use and popularized since the 1960s, XX century. Despite their well-documented neurotoxicity, they reached reputation of being safe and nonaddictive. Scientific efforts to find any medical application for these hallucinogens in psychiatry, psychotherapy, and even for religious rituals support are highly controversial. Even if they show any healing potential, their usage in psychotherapy is in some cases inadequate and may additionally harm seriously suffering patients. Hallucinogens are thought to reduce cognitive functions. However, in case of indolealkylamines, such as psilocin, some recent findings suggest their ability to improve perception and mental skills, what would motivate the consumption of "magic mushrooms." The present article offers an opportunity to find out what are the main symptoms of intoxication with mushrooms containing psilocybin/psilocin, muscimol, and ibotenic acid. The progress in analytical methods for detection of them in fungal material, food, and body fluids is reviewed. Findings on the mechanisms of their biologic activity are summarized. Additionally, therapeutic potential of these fungal psychoactive compounds and health risk associated with their abuse are discussed.
Rapid quantification of psilocybin and psilocin in extracts of wild mushrooms is accomplished by reversed-phase high-performance liquid chromatography with paired-ion reagents. Nine solvent systems and three solid supports are evaluated for their efficiency in separating psilocybin, psilocin and other components of crude mushroom extracts by thin-layer chromatography.
High-performance liquid chromatography (HPLC) was used for the analysis of the minor consistuents psilocybin and psilocin in fungi of the genus Psilocybe. The separation and determination of these compounds was carried out on a stationary phase of LiChrosorb RP-18. The analytical column (A) and semipreparative column (B) were eluted isocratically with water—ethanol—acetic acid (79.2:20:0.8) at flow-rates of 20 ml/h (A) and 180 ml/h (B). The compounds were detected with a UV detector at 267 nm and a fluorometric detector (excitation, 280 nm; emission, 360 nm). The UV detection limit of psilocybin was 20–40 ng (267 nm) and several ng could be detected fluorometrically. The identity of the compounds was verified by HPLC and thin-layer chromatography and by mass spectrometry and UV spectroscopy. The compounds were determined by means of a direct calibration method and by means of the method of internal normalization. The standard deviation of the determination was ±3.4% (relative). The above methods were used to determine these compounds in extracts of fruit bodies of two species of the genus Psilocybe growing at various places in Czechoslovakia, and found to contain 0.25–1.15% of psilocybin and 0.02–0.16% psilocin per dry mass.
This paper describes a procedure for the determination of psilocin and psilocybin in mushroom extracts using high-performance liquid chromatography with postcolumn chemiluminescence detection. A number of extraction methods for psilocin and psilocybin in hallucinogenic mushrooms were investigated, with a simple methanolic extraction being found to be most effective. Psilocin and psilocybin were extracted from a variety of hallucinogenic mushrooms using methanol. The analytes were separated on a C12 column using a (95:5% v/v) methanol:10 mM ammonium formate, pH 3.5 mobile phase with a run time of 5 min. Detection was realized through a dual reagent chemiluminescence detection system of acidic potassium permanganate and tris(2,2′-bipyridyl)ruthenium(II). The chemiluminescence detection system gave improved detectability when compared with UV absorption at 269 nm, with detection limits of 1.2 × 10−8 and 3.5 × 10−9 mol/L being obtained for psilocin and psilocybin, respectively. The procedure was applied to the determination of psilocin and psilocybin in three Australian species of hallucinogenic mushroom.
Khat, the leaves of Catha edulis Forsk., is widely used as a “natural amphetamine” in East Africa and the Arab Peninsula. In recent years, air-transported khat appeared on local khat markets in Europe (e.g. France, Italy, Great Britain). Due to increasing seizures of khat imported by ethnic communities and emigrants the psychoactive drug has been banned in 1992 by the Swiss Health Authorities. The demand for monitoring the psychotropic potential of the illicit drug made it necessary to establish specific, sensitive and rapid methods for the determination of the alkaloids, mainly the psychoactive principle S-(−)-cathinone. The S-(−)-cathinone concentration measured by high-performance liquid chromatography with photodiode array detection (HPLC-DAD) and alternatively by instrumental thin-layer chromatography (TLC) varied between 2182 and 4506 μg/g dried material. It is typical for khat originating from Kenya and demonstrates its high psychoactivity. The S,S-(+)-norpseudoephedrine and R,S-(−)-norephedrine content ranged from 2114 to 4094 and 192 to 1345 μg/g, respectively.
An efficient analytical and isolation method was elaborated for biologically active tryptamines using a computer-aided liquid chromatographic-gas chromatographic system. The separation method includes a new efficient extraction procedure, optimization programmed for high-performance liquid chromatographic separation, identification by diode-array detection and a spectrometric and electrochemical assay. The identification of indole alkaloids was confirmed by thin-layer and gas chromatography and mass spectrometry. The method was used for analysis and isolation of psychotropic substances in extracts from the fruit bodies of hallucinogenic fungi of genera Psilocybe, Inocybe and Amanita and in mycelial extracts from the species Psilocybe bohemica.
Khat, the leaves of Catha edulis Forsk. (Celastraceae), is widely used as a stimulant in East Africa and the Arab Peninsula. Samples from the most important markets of Ethiopia, Kenya, North Yemen and Madagascar were analysed with high-performance liquid chromatography (HPLC) concerning their khatamine content (phenylpropyl and phenylpentenylamines). The variability of khatamines depending on origin, type and quality is discussed. In many samples a good correlation between the amount of cathinone, the main CNS-active compound, and quality estimation (price) of dealers and consumers was found.
This paper presents a review of literature on the chemical composition of khat (Catha edulis Forsk., Celastraceae). The effect of chewing fresh khat could not be explained satisfactorily by the action of d-norpseudoephedrine which was, for a long time, believed to be the only stimulant in khat. A comprehensive study on the chemical composition of khat was undertaken at the United Nations narcotics Laboratory with the aim of isolating and characterizing the principles of the fresh plant active on the central nervous system. This work resulted in the detection and isolation of cathinone, a phenylalkylamine characterized as (-)-alpha-aminopropiophenone. It is the main phenylalkylamine component of fresh khat, and pharmacological studies indicate that it may be the compound responsible for the characteristic stimulant activity and abuse potential of the plant. Some of its "decomposition" or transformation products, such as norpseudoephedrine, norephedrine, 3,6-dimethyl-2,5-diphenylpyrazine, and 1-phenyl-1,2-propanedione, have also been isolated and characterized.
The occurrence and extraction of indole derivatives in six species from four genera of higher fungi were investigated. By using pure methanol for extraction of the mushrooms analysis revealed the highest concentrations of psilocybin and baeocystin. The psilocin content of the species was higher by using aqueous solutions of alcohols than with methanol alone but was an artificial phenomenon caused by enzymatic destruction of psilocybin. The extraction with dilute acetic acid yielded better results than with the water containing alcohols.
The simple one-step procedure with methanol for the quantitative extraction is still the safest method to obtain the genuine alkaloids from fungal biomass.
'Magic mushrooms' is the name most commonly given to psychoactive fungi containing the hallucinogenic components psilocybin and psilocin. Material confiscated by local authorities was examined using morphologic, microscopic, microchemical, and toxicological methods. Psilocybe cubensis was the most popular mushroom in the sample collective, followed by Psilocybe semilanceata, Panaeolus cyanescens and Psilocybe tampanensis. The alkaloid content was determined with <0.003-1.15% of psilocybin and 0.01-0.90% psilocin. Panaeolus cyanescens was the mushroom with highest levels of psilocybin and psilocin.
The stimulating herbal drug kath is uncommon in most countries, and information on its detection and interpretation of analytical results is limited. Therefore, a study with kath was carried out to compare the efficiencies of different analytical techniques used to detect drug use.
Four volunteers chewed kath leaves for 1 h; urine samples were collected up to 80 h afterward and analyzed by the Abbott fluorescence polarization immunoassay (FPIA), the Mahsan-AMP(300) on-site immunoassay, the Bio-Rad Remedi HS HPLC system with photodiode array detection (DAD), and gas chromatography-mass spectrometry (GC-MS).
FPIA gave negative results, whereas positive results were obtained with the Mahsan test during the first day. With HPLC, one peak could be observed up to 50 h, but its DAD spectrum could not be identified by the system. Further investigations indicated that the kath alkaloids coeluted and produced a mixed DAD spectrum. With GC-MS, the specific kath ingredient cathinone was detected up to 26 h, whereas cathine and norephedrine were still detectable in the last samples. Maximum concentrations of cathinone, cathine, and norephedrine in urine samples from the study were 2.5, 20, and 30 mg/L, respectively, whereas in authentic cases the concentrations were much higher.
GC-MS is superior to the screening techniques Mahsan-AMP(300) and Remedi with respect to specificity and sensitivity for the detection of kath use in urine.
A sensitive and reproducible method for the quantitative determination of cathinone (CTN), norpseudoephedrine (NPE, cathine) and norephedrine (NE) from hair was developed. The compounds were extracted for 4 hours with phosphate buffer pH 2.0, followed by a standard solid phase extraction procedure on a mixed phase column, derivatization with heptafluorobutyric anhydride (HFBA) and GC-MS separation and quantification using D(3)-ephedrine (D(3)-E) and alpha-aminoacetophenone (AAP) as the internal standards. The diastereomers NPE and NE were satisfactorily separated. In the validation, the limits of detection and of quantification were determined at 0.03-0.08 ng/mg and 0.10-0.24 ng/mg, respectively and the interday standard deviation was between 10 and 15%. The method was applied to hair samples of 24 Yemenite khat chewers. All three compounds were detected in 23 of these cases. The concentrations ranged from 0.57 to 23.9 ng/mg for NPE, 0.19-25.0 ng/mg for NE and 0.11-22.7ng/mg for CTN. A highly significant correlation was found between the self-reported data about the khat consumption habits of the volunteers (4-56h chewing per week) and the concentrations of norephedrine and norpseudoephedrine in hair.
A GC-MS method has been developed for the detection of amphetamine, methamphetamine, and the ephedrines, in seizures and the urine, based on on-GC condensation (derivatization) with cyclohexanone. The method is simple: the dried seizure material or the urine extract was mixed with cyclohexanone and injected into the GC-MS. The method was found to be superior to the methods based on acyl and trimethylsilyl (TMS) derivatization. Unlike for the acyl and TMS derivatives, the molecular and fragment ions of the cyclohexanone condensation products (cyclohexanone derivatives) were of substantial abundance, a useful property in unambiguous compound characterization. Furthermore, the high stability of the "derivatizing" reagent, cyclohexanone, compared with acyl and TMS derivatizing reagents, is a useful property in method development. The present method has proved selective and, tentatively, sensitive enough in the following areas (where methods based on acyl and TMS derivatization, as tested in this laboratory, have failed): (a) detection of amphetamine as a metabolite of methamphetamine; (b) detection of norpseudoephedrine as a metabolite of pseudoephedrine; (c) detection of amphetamine as an impurity of methamphetamine; (d) detection of cathine (norephedrine) as a constituent of Khat leaves; and (e) differentiation of Khat use from phenylpropanolamine use.
Morphological and toxicological analyses were performed on hallucinogenic mushrooms that are currently circulated in Japan. Scanning electron microscope (SEM) indicated a three-dimensional microstructures in the mushrooms. The complementary use of SEM with an optical microscope was effective for observing characteristic tissues, such as basidiomycetes, spores, cystidia and basidia. Hallucinogenic alkaloids were extracted with methanol and determined by high performance liquid chromatography (HPLC) with a UV detector set at 220 nm. The psilocin/psilocybin contents in Psilocybe cubensis were in the range of 0.14-0.42%/0.37-1.30% in the whole mushroom (0.17-0.78%/0.44-1.35% in the cap and 0.09-0.30%/0.05-1.27% in the stem), respectively. The hallucinogenic alkaloids in Copelandia were 0.43-0.76%/0.08-0.22% in the whole mushroom (0.64-0.74%/0.02-0.22% in the cap and 0.31-0.78%/0.01-0.39% in the stem). It thus appears that P. cubensis is psilocybin-rich, whereas Copelandia is psilocin-rich.
The herbal stimulants Ephedra species, Catha edulis (khat), and Lophophora williamsii (peyote) have been abused for a long time. In recent years, the herbal drug market has grown owing to publicity on the Internet. Some ingredients of these plants are also ingredients of cold remedies. The aim of the presented study is to develop a multianalyte procedure for detection and validated quantification of the phenalkylamines ephedrine, pseudoephedrine, norephedrine, norpseudoephedrine, methylephedrine, methylpseudoephedrine, cathinone, mescaline, synephrine (oxedrine), and methcathinone in plasma. After mixed-mode solid-phase extraction of 1 ml of plasma, the analytes were separated using a strong cation exchange separation column and gradient elution. They were detected using a Q-Trap LC-ESI-MS/MS system (MRM mode). Calibration curves were used for quantification using norephedrine-d3, ephedrine-d3, and mescaline-d9 as internal standards. The method was validated according to international guidelines. The assay was selective for the tested compounds. It was linear from 10 to 1000 ng/ml for all analytes. The recoveries were generally higher than 70%. Accuracy ranged from - 0.8 to 20.0%, repeatability from 2.5 to 12.3%, and intermediate precision from 4.6 to 20.0%. The lower limit of quantification was 10 ng/ml for all analytes. No instability was observed after repeated freezing and thawing or in processed samples. The applicability of the assay was tested by analysis of authentic plasma samples after ingestion of different cold medications containing ephedrine or pseudoephedrine, and after ingestion of an aqueous extract of Herba Ephedra. After ingestion of the cold medications, only the corresponding single alkaloids were detected in human plasma, whereas after ingestion of the herb extract, all six ephedrines contained in the plant were detected. The presented LC-MS/MS assay was found applicable for sensitive detection and accurate and precise quantification of all studied analytes in plasma.