Hashish Research Articles

Acetylcholinesterase, butyrylcholinesterase and paraoxonase 1 activities in rats treated with cannabis, tramadol or both.

To investigate the effect of Cannabis sativa resin and/or tramadol, two commonly drugs of abuse on acetylcholinesterase and butyrylcholinesterase activities as a possible cholinergic biomarkers of neurotoxicity induced by these agents. Rats were treated with cannabis resin (5, 10 or 20mg/kg) (equivalent to the active constituent Δ9-tetrahydrocannabinol), tramadol (5, 10 and 20mg/kg) or tramadol (10mg/kg) combined with cannabis resin (5, 10 and 20mg/kg) subcutaneously daily for 6 weeks. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activities were measured in brain and serum. We also measured the activity of paraoxonase-1 (PON1) in serum of rats treated with these agents. (i) AChE activity in brain increased after 10-20mg/kg cannabis resin (by 16.3-36.5%). AChE activity in brain did not change after treatment with 5-20mg/kg tramadol. The administration of both cannabis resin (5, 10 or 20mg/kg) and tramadol (10mg/kg) resulted in decreased brain AChE activity by 14.1%, 12.9% and 13.6%, respectively; (ii) BChE activity in serum was markedly and dose-dependently inhibited by cannabis resin (by 60.9-76.9%). BChE activity also decreased by 17.6-36.5% by 10-20mg/kg tramadol and by 57.2-63.9% by the cannabis resin/tramadol combined treatment; (iii) Cannabis resin at doses of 20mg/kg increased serum PON1 activity by 25.7%. In contrast, tramadol given at 5, 10 and 20mg/kg resulted in a dose-dependent decrease in serum PON1 activity by 19%, 36.7%, and 46.1%, respectively. Meanwhile, treatment with cannabis resin plus tramadol resulted in 40.2%, 35.8%, 30.7% inhibition of PON1 activity compared to the saline group. These data suggest that cannabis resin exerts different effects on AChE and BChE activities which could contribute to the memory problems and the decline in cognitive function in chronic users.

The main cannabinoids content in hashish samples seized in Israel and Czech Republic

Cannabis, both herbal and resin, has been the most popular illicit drug in Israel in recent years. Until 2005, the main sources of cannabis resin (known also as hashish) to the Israeli drug market were Lebanon and India. Hashish from these sources can be distinguished by its external appearance. The aim of this study was to determine if there is any difference in the quality of the hashish from each source. For this purpose, we quantified the main cannabinoids, cannabidiol (CBD), Δ9-tetrahydrocannabinol (Δ9-THC), and cannabinol (CBN) of hashish from different police seizures of known origins, Lebanon, India and Morocco, that had been submitted to the Analytical Chemistry Laboratory of the Division of Identification and Forensic Science at the Israeli National Police Headquarters and subsequently to the Hebrew University for quantitative analysis. The results, based on many different seizures, showed that the average CBD content of hashish from Lebanon varied from 5.69% to 12.79% (an average 8.98 ± 0.59%), THC of hashish from Lebanon varied from 0.93% to 4.20% (an average of 2.38 ± 0.27 %), CBD of hashish from Morocco varied from 1.52% to 5.14% (an average of 3.72 ± 0.19%), THC of hashish from Morocco varied from 5.08% to 13.41% (an average of 9.21 ± 0.40%), CBD of hashish from India varied from 0.78% to 13.13% (an average of 4.59 ± 1.07%), and THC of hashish from India varied from 0.53% to 16.45% (an average of 6.35 ± 1.50%), At the same time, several other cannabinoids present in the samples in lower amounts were identified (cannabidivarol, CBDV; cannabicitran; Δ9-tetrahydrocannabivarol, Δ9-THCV; cannabivarol, CBV; cannabicyclol, CBL; cannabichromene, CBC; cannabielsoin, CBE; Δ8-THC; and cannabigerol, CBG). The samples, predominantly from Lebanon, Morocco, and India, were evaluated for chemical phenotype (drug type and fiber type) to determine the geographical origin of these samples.Indian hashish, seized by Czech Republic authorities, was analyzed qualitatively for many cannabinoids and terpenes. In this sample three new cannabinoids were identified: cannabiorcochromene, cis-Δ9-tetrahydrocannabivarol, and cannabinerol.

Results of an international drug testing service for cryptomarket users

IntroductionUser surveys indicate that expectations of higher drug purity are a key reason for cryptomarket use. In 2014–2015, Spain's NGO Energy Control conducted a 1-year pilot project to provide a testing service to cryptomarket drug users using the Transnational European Drug Information (TEDI) guidelines. In this paper, we present content and purity data from the trial. Methods219 samples were analyzed by gas chromatography associated with mass spectrometry (GC/MS). Users were asked to report what substance they allegedly purchased. Results40 different advertised substances were reported, although 77.6% were common recreational drugs (cocaine, MDMA, amphetamines, LSD, ketamine, cannabis). In 200 samples (91.3%), the main result of analysis matched the advertised substance. Where the advertised compound was detected, purity levels (m±SD) were: cocaine 71.6±19.4%; MDMA (crystal) 88.3±1.4%; MDMA (pills) 133.3±38.4mg; Amphetamine (speed) 51.3±33.9%; LSD 123.6±40.5μg; Cannabis resin THC: 16.5±7.5% CBD: 3.4±1.5%; Ketamine 71.3±38.4%. 39.8% of cocaine samples contained the adulterant levamisole (11.6±8%). No adulterants were found in MDMA and LSD samples. DiscussionThe largest collection of test results from drug samples delivered from cryptomarkets are reported in this study. Most substances contained the advertised ingredient and most samples were of high purity. The representativeness of these results is unknown.

Synthetic cannabinoids 2015: An update for pediatricians in clinical practice.

Synthetic cannabinoids are a group of substances in the world of designer drugs that have become increasingly popular over the past few years. Synthetic cannabinoids are a chemically diverse group of compounds functionally similar to THC. Since first appearing on the world market a few years ago these compounds have evolved rapidly. Newer more potent analogues have been developed. Identifying youth who abuse these substances can be difficult. Newer forms of consumption have also evolved. These products are now manufactured in products that look like natural cannabis resin and in liquid cartridges used in electronic cigarettes. Synthetic cannabinoids appear to be associated with potentially dangerous health effects that are more severe than that of marijuana. Some synthetic cannabinoid compounds have been associated with serious physical consequences, such as, seizures, myocardial infarction and renal damage. In addition, psychoactive effects, such as aggression, confusion, anxiety and psychosis have also been reported. The diagnosis remains primarily clinical with toxicological confirmation difficult due to manufacturers constantly developing new analogues to avoid detection. Pediatricians are urged to familiarize themselves with these drugs and the typical presentations of patients who use them.

Proximate composition, phytochemical screening, GC-MS studies of biologically active cannabinoids and antimicrobial activities of Cannabis indica

ABSTRACT Objective To investigate the proximate composition, minerals analysis, phytochemical screening, gas chromatography-mass spectrometry (GC-MS) studies of active cannabinoids and antimicrobial activities of Cannabis indica ( C. indica ) leaves, stems, and seeds. Methods Standard qualitative protocols of phytochemical screening were accomplished for the identification of biologically active phytochemicals. Minerals in plant samples were analyzed by using atomic absorption spectrophotometer. The resins of C. indica were analyzed for medicinally active cannabinoid compounds by GC-MS. The sample for GC-MS study was mixed with small quantity of n -hexane and 30 mL of acetonitrile solution for the identification of cannabinoids. Agar well diffusion method was used for antibacterial activity. For antifungal activity, the tested fungal strains were sub-cultured on Sabouraud's dextrose agar at 28 °C. Results Mineral analysis revealed the presence of sodium, potassium, magnesium and some other minerals in all parts of C. indica . Phytochemical investigation showed the presence of alkaloids, saponins, tannins, flavonoids, sterols and terpenoids. C. indica divulged wide spectrum of antibacterial activities against Staphylococcus aureus, Bacillus cereus, Klebsiella pneumoniae , and Proteus mirabilis . The extracts of plant leaves, seeds and stems showed significant antifungal activities against Aspergillus niger, Aspergillus parasiticus , and Aspergillus oryzae . The biologically active cannabinoids of delta-9-tetrahydrocannabinol (25.040%) and cannabidiol (resorcinol, 2-p-mentha-1,8-dien-4-yl-5-pentyl) (50.077%) were found in Cannabis resin in high percentage. Conclusions The findings of the study suggested that the existence of biologically active remedial cannabinoids in elevated concentrations and antimicrobial bioassays of C. indica make it a treasured source to be used in herbal preparation for various ailments.

Potency trends of Δ9-tetrahydrocannabinol, cannabidiol and cannabinol in cannabis in the Netherlands: 2005-15.

Between 2000 and 2005 the average percentage of Δ(9) -tetrahydrocannabinol (THC) in marijuana as sold in Dutch coffeeshops has increased substantially; the potency of domestic products (Nederwiet and Nederhasj) has particularly increased. In contrast with imported marijuana, Nederwiet hardly contained any cannabidiol (CBD), a cannabinoid that is thought to offset some of the adverse effects of THC. In 2005, the THC content in Nederwiet was significantly lower than in 2004. This study investigates the further decrease or increase of cannabinoids in these cannabis products. From 2005 to 2015 five different cannabis products were bought anonymously in 50 coffeeshops that were selected randomly each year from all coffeeshops in the Netherlands. A total of 2126 cannabis samples were bought, consisting of 664 Nederwiet samples (most popular), 537 Nederwiet samples (supposed strongest varieties), 183 imported herbal cannabis samples, 140 samples of cannabis resin made of Nederwiet and 602 samples of imported cannabis resin. All samples were analysed chemically for their THC, CBD and cannabinol (CBN) content. Between 2005 and 2015, the mean potencies of the most popular and the strongest Nederwiet and of imported cannabis resin were 16.0±4.0%, 17.0±3,9% and 16.5±6.3%, respectively. Imported herbal cannabis (6.5±3.5%) and cannabis resin made from Nederwiet (30.2±16.4%) contained, respectively, less (β=-10.0, P<0.001) and more (β=13.7, P<0.001) THC than imported cannabis resin. Linear regression models were used to study the trends in THC of the different cannabis products over time. A marginal, but significant (P<0.001), overall decline of THC per year of 0.22% was found in all cannabis products. However, no significant difference was found between the five products in the THC linear trajectories across time. Of all the cannabis products, only imported cannabis resin contained a relatively high CBD/THC ratio (median 0.42). The average tetrahydrocannabinol (THC) content of the most popular herbal cannabis products in the Netherlands has decreased slightly since 2005. The popular Nederwiet type still has a relatively high THC to cannabidiol ratio.

False hashish without cannabis resin

A substance that looked like hashish, seized in Israel and the Czech Republic was analyzed with a GC/MS for the identification of the main cannabinoids expected in cannabis (Cannabis sativa). No compounds typical of cannabis were identified in either of the two sets of samples, but compounds typical of coniferous plants, most probably from pine, and henna (Lawsonia inermis), were identified (various alcohols and organic acids).

Some physical properties of novel Cannabis suppositories formulated with theobroma oil

As part of development efforts for a suitable dosage form, crude Cannabis resin was formulated into suppository dosage form using theobroma oil and the physical properties of the suppositories were evaluated. The following physical properties were evaluated: appearance (texture, presence or absence of entrapped air, contraction holes), liquefaction time, uniformity of weight and in-vitro release profile of the crude marijuana resin from the suppositories. The torpedo shaped suppositories were smooth in texture with absence of entrapped air and contraction holes. The suppositories had uniform greenish brown colour and low weight variation. The liquefaction time was also low. The 300 mg Cannabis crude in 4 % Tween 85 showed highest melting time (11.67 ± 057 min) while the incorporation of Tween 85 improved the release profile (0.0452-0.0650 %) in different batches. It is possible to formulate marijuana suppositories with satisfactory physical properties; however, release profile of marijuana from the suppository bases was generally low even though the addition of Tween 85 greatly enhanced drug release. Key words: Crude Cannabis resin, sustained release, liquefaction time, weight uniformity, release profile.

Multidimensional analysis of cannabis volatile constituents: Identification of 5,5-dimethyl-1-vinylbicyclo[2.1.1]hexane as a volatile marker of hashish, the resin of Cannabis sativa L.

The volatile constituents of drug samples derived from Cannabis sativa L. were investigated by means of headspace solid phase microextraction (HS-SPME) and gas chromatography techniques (GC–MS, GC×GC–MS). Samples of cannabis herb and hashish showed clear differences in their volatile chemical profiles, mostly resulting from photo-oxidation processes occurring during the transformation of fresh cannabis herb into hashish. Most unexpectedly, we could demonstrate hashish samples as containing remarkable amounts of a rare and unusual monoterpene – 5,5-dimethyl-1-vinylbicyclo[2.1.1]hexane – among the volatile compounds detected in their headspaces. We gave evidence for the formation of this compound from the light induced rearrangement of β-myrcene during the manufacture of hashish. In view of its high abundance among volatile constituents of cannabis resin and its scarce occurrence in other natural volatile extracts, we propose to rename this specific monoterpene hashishene.

O39: Death by self-mutilation after cannabis consumption in a patient with cortical dysplasia

Aims The use of cannabis has been linked to an increased risk for acute psychosis, especially delirious episodes or schizophrenicspectrum disorders. Self-mutilation and dissociative disorders are rare (Bergua A et al. Ophthalmologica 2002;216:269–71). On the contrary, according to some authors, cannabis resin consumption would reduce these disorders (Passie T et al. Drug Test Anal. 2012;4:649-59). We report on original case of death by automutilation in a patient with cortical dysplasia following consumption of a highly concentrated resin cannabis. Case report Mr. B., 36 years-old, presents on the public roads a major state of agitation with self-mutilation and bilateral enucleation. Despite the intervention of emergency services, he impales on the fence and dies in an array of cataclysmic hemorrhage. According to statements of two witnesses who have spent the day with the victim, he would have presented a state of major agitation after spending few minutes in his apartment. They also indicate that the victim was a regular cannabis user and had presented unlabeled psychiatric disorders few years earlier. Four fragments of a brown substance resembling cannabis resin were seized at the home of the victim. Methods The autopsy was performed 12 hours later. Toxicological samples (cardiac blood, peripheral blood, pleural fluid, gastric content, vitreous humor and hair) were taken. Blood screening for major drugs and narcotics was done by LC-MS/MS. Dosages of amphetamines, opiates and cocaine were done by GC-MS, quantification of cannabinoids by GC-MS/MS. Identification and dosage of hallucinogens, new synthetic drugs, and synthetic cannabinoids were performed by LC-MS and LC-MS/MS. The composition of the four brown fragments seized at the home of the victim was determined by GC-MS and GC-FID. A pathological expertise was also performed. Results Toxicological analysis of the biological fluids and hair revealed the presence of different markers of cannabis consumption: THC 11-OH-THC THC-COOH Cannabinol Cannabidiol Cardiac blood (ng/mL) 25 3.2 13 0.6 0.3 Femoral blood (ng/mL) 13.5 4.1 14.7 1.3 0.7 Gastric content (ng/mL) 868 0.8 26 – 22 Hair (1,5 cm) (pg/mg) 137 1 ns* 9 94 These results indicate a recent cannabis consumption and the high gastric concentrations are in favor of an orally absorption. Hair analysis confirms regular consumption. A postmortem redistribution between the blood compartments can be observed. The analysis of fragments (F1 to F4) confirmed that it was cannabis resin with following composition (%): THC Cannabinol Cannabidiol F1- F3 31.1 5.5 1 F2 34.8 6 1.1 F4 35.4 6 1 The 4 pieces of cannabis resin highlights similar qualitative and quantitative composition with very high levels of active compounds. The pathological expertise reveals brain cortical dysplasia and lung lesions alveolitis type with pigmented macrophages related to regular toxic inhalation. Discussion and conclusion cortical dysplasia induces seizures and hallucinations. In this particular case, the absorption of a large dose of an highly concentrated cannabis extract may have caused hallucinations and dissociative disorders.

O3: Cannabinoids in young children hair: Preliminary data from some cases

Aims The differentiation between an active cannabis consumption and a passive drug exposure (i.e. through side stream marijuana smoke, or transfer from contaminated hands), remains an unsolved problem in hair analysis [Uhl M., Sachs H. Forensic Science International 2004;145:143–147] . The presence in hair of Δ 9 -tetrahydrocannabinolic acid A [THCA-A; a non-psychoactive precursor of Δ 9 -tetrahydrocannabinol (THC) and the main cannabinoid in a fresh plant] was recently proposed as a specific marker of external contamination [Roth N et al. Journal of Mass Spectrometry 2013;48:227–233] . However, the evidence of active consumption of cannabis (inhalated and/or oral) currently remains based on the detection of 11-nor-9-carboxy-Δ 9 -tetrahydrocannabinol acid (THC-COOH) in hair. Nevertheless, the missing detection of this metabolite does not exclude an active consumption, as the low pg/mg range of THCCOOH concentrations in hair of cannabis users can unable to detect it despite using extremely sensitive analytical methods. In this context, the aim of this presentation is to highlight this problem in young children hair. Cases stories Twelve hair samples collected from 9 young children (4F/5M – 2 to 14 months old) were sent to the laboratory for cannabinoid determination in order to explore their cannabis exposure (their parents being regular cannabis users) after (or not) an acute cannabis intoxication (cannabis resin intake): overall eighteen hair segments were analysed. Methods A recently published liquid chromatography – tandem mass spectrometry (LC – MS/MS) method was used for the simultaneous quantitative determination in hair of THC-COOH, THC, Cannabinol (CBN) and Cannabidiol (CBD) [Dulaurent S et al. Forensic Science International 2014;236:151–156.] . Briefly, after a decontamination step, the sample preparation consisted of an alkaline hydrolysis of hair samples followed by a liquid-liquid extraction of compounds in acidic conditions. The chromatographic detection was performed by means of MRM and MS 3 transitions using an API 5500 QTRAP mass spectrometer (AB Sciex, Courtaboeuf, France), with intra- and inter-assay accuracies and relative standard deviation below 9% and 15%, respectively, for the four compounds. LOQ (= LOD) was 0.2 pg/mg for THC-COOH and 50 pg/mg for THC, CBD and CBN. Results and conclusion Some selected results in hair segments (which do not correspond to a documented acute accidental intoxication period), are shown in the following table (concentration in ng/mg): Case Age (M) Sexe Hair segment THC CBN CBD THC-COOH 1 2 M global – 2,5 cm nd nd 0.41 nd 2 11 F 0 to 2 cm 0.13 0.17 0.13 nd 3 13 F 0 to 3 cm 1.17 0.38 2.17 nd 4 11 M 1 to 3 cm 1.21 0.38 2.16 0.00068 Interpretation of these results could be as follows: (case 1) low exposure in relation to low environmental exposure or in utero exposure, (case 2) low and (case 3) high exposure in relation to environmental exposure, and (case 4) high exposure in relation to environmental exposure with significant absorption by his organism. Nevertheless, in order to improve the interpretation of such results in this particular population and due to the tricky context, (i) many pitfalls have to be overcome, in particular ontogenic issues (i.e. CYP2C9 and UGT expression in infants), and (ii) additional clinical and analytical data are needed.

Sampling of illicit drugs for quantitative analysis – Part III: Sampling plans and sample preparations

The findings in this paper are based on the results of our drug homogeneity studies [1] and particle size investigations [2]. Using that information, a general sampling plan (depicted in the form of a flow-chart) was devised that could be applied to the quantitative instrumental analysis of the most common illicit drugs: namely heroin, cocaine, amphetamine, cannabis resin, MDMA tablets and herbal cannabis in 'bud' form (type I). Other more heterogeneous forms of cannabis (type II) were found to require alternative, more traditional sampling methods.A table was constructed which shows the sampling uncertainty expected when a particular number of random increments are taken and combined to form a single primary sample. It also includes a recommended increment size; which is 1g for powdered drugs and cannabis resin, 1 tablet for MDMA and 1 bud for herbal cannabis in bud form (type I).By referring to that table, individual laboratories can ensure that the sampling uncertainty for a particular drug seizure can be minimised, such that it lies in the same region as their analytical uncertainty for that drug.The table shows that assuming a laboratory wishes to quantitatively analyse a seizure of powdered drug or cannabis resin with a ‘typical’ heterogeneity, a primary sample of 15×1g increments is generally appropriate. The appropriate primary sample for MDMA tablets is 20 tablets, while for herbal cannabis (in bud form) 50 buds were found to be appropriate.Our study also showed that, for a suitably homogenised primary sample of the most common powdered drugs, an analytical sample size of between 20 and 35mg was appropriate and for herbal cannabis the appropriate amount was 200mg.The need to ensure that the results from duplicate or multiple incremental sampling were compared, to demonstrate whether or not a particular seized material has a ‘typical’ heterogeneity and that the sampling procedure applied has resulted in a ‘correct sample’, was highlighted and the setting up of suitable control charts (R or S charts), for quality control purposes, was strongly recommended and examples given.Furthermore, although this particular study relates to the sampling of illicit drugs, it should be remembered that it is based on general sampling theory and therefore the same approach could be applied to other disciplines where ‘correct sampling’ of powders and solids is important.

The retail value of the illicit drug market in Italy: a consumption-based approach

This study estimates the retail value of the illicit drug market in Italy from a consumption-based approach. The illicit drugs considered in this analysis are heroin, cocaine, cannabis (herbal and resin), amphetamines and ecstasy. Results show that the value of the illicit drug market in Italy is much less than previously estimated and quantified at € 3.3 bn. Heroin and cocaine retain the biggest markets in terms of revenues, while cannabis is the most-consumed illicit drug. Synthetic illicit drugs account for roughly 10% of the illicit drug market. Conclusions offer some suggestions as to how uncertainties about estimates of the illicit drug market can be reduced.

Extraction of High Quality DNA from Seized Moroccan Cannabis Resin (Hashish)

The extraction and purification of nucleic acids is the first step in most molecular biology analysis techniques. The objective of this work is to obtain highly purified nucleic acids derived from Cannabis sativa resin seizure in order to conduct a DNA typing method for the individualization of cannabis resin samples. To obtain highly purified nucleic acids from cannabis resin (Hashish) free from contaminants that cause inhibition of PCR reaction, we have tested two protocols: the CTAB protocol of Wagner and a CTAB protocol described by Somma (2004) adapted for difficult matrix. We obtained high quality genomic DNA from 8 cannabis resin seizures using the adapted protocol. DNA extracted by the Wagner CTAB protocol failed to give polymerase chain reaction (PCR) amplification of tetrahydrocannabinolic acid (THCA) synthase coding gene. However, the extracted DNA by the second protocol permits amplification of THCA synthase coding gene using different sets of primers as assessed by PCR. We describe here for the first time the possibility of DNA extraction from (Hashish) resin derived from Cannabis sativa. This allows the use of DNA molecular tests under special forensic circumstances.

Sampling of illicit drugs for quantitative analysis—Part II. Study of particle size and its influence on mass reduction

The basic goal in sampling for the quantitative analysis of illicit drugs is to maintain the average concentration of the drug in the material from its original seized state (the primary sample) all the way through to the analytical sample, where the effect of particle size is most critical.The size of the largest particles of different authentic illicit drug materials, in their original state and after homogenisation, using manual or mechanical procedures, was measured using a microscope with a camera attachment. The comminution methods employed included pestle and mortar (manual) and various ball and knife mills (mechanical). The drugs investigated were amphetamine, heroin, cocaine and herbal cannabis.It was shown that comminution of illicit drug materials using these techniques reduces the nominal particle size from approximately 600μm down to between 200 and 300μm.It was demonstrated that the choice of 1g increments for the primary samples of powdered drugs and cannabis resin, which were used in the heterogeneity part of our study (Part I) was correct for the routine quantitative analysis of illicit seized drugs. For herbal cannabis we found that the appropriate increment size was larger.Based on the results of this study we can generally state that:An analytical sample weight of between 20 and 35mg of an illicit powdered drug, with an assumed purity of 5% or higher, would be considered appropriate and would generate an RSDsampling in the same region as the RSDanalysis for a typical quantitative method of analysis for the most common, powdered, illicit drugs. For herbal cannabis, with an assumed purity of 1% THC (tetrahydrocannabinol) or higher, an analytical sample weight of approximately 200mg would be appropriate.In Part III we will pull together our homogeneity studies and particle size investigations and use them to devise sampling plans and sample preparations suitable for the quantitative instrumental analysis of the most common illicit drugs.

Variability of cannabis potency in the Venice area (Italy): A survey over the period 2010–2012

Cannabis is the most widely used illicit substance globally, with an estimated annual prevalence in 2010 of 2.6-5.0% of the adult population. Concerns have been expressed about increases in the potency of cannabis products. A high tetrahydrocannabinol (THC) content can increase anxiety, depression, and psychotic symptoms, and can increase the risk of dependence and adverse effects on the respiratory and cardiovascular systems in regular users. The aim of this study was to report statistical data about the potency of cannabis products seized in the north-east of Italy, in a geographical area centred in Venice and extending for more than 10,000 km(2) with a population of more than two million, by investigating the variability observed in THC levels of about 4000 samples of cannabis products analyzed over the period 2010-2012. Overall median THC content showed an increasing trend over the study period from about 6.0% to 8.1% (6.2-8.9% for cannabis resin, 5.1-7.6% for herbal cannabis). The variation in the THC content of individual samples was very large, ranging from 0.3% to 31% for cannabis resin and from 0.1 to 19% for herbal cannabis. Median CBN:THC ratios showed a slightly decreasing trend over the study period, from 0.09 (2010) to 0.03 (2012), suggesting an increasing freshness of submitted materials. Median CBD:THC ratios also showed a decreasing trend over the study from about 0.52 (2010) to 0.18 (2012), likely due to the increase in submissions of materials from indoor and domestic cultivation with improved breeding methods.

Sampling of illicit drugs for quantitative analysis. Part I: Heterogeneity study of illicit drugs in Europe

Sampling of illicit drugs for qualitative and quantitative analysis would normally be considered as routine and comparable tasks in forensic drugs laboratories and previously similar statistical sampling approaches have been applied. However, we believe that two different sampling approaches, based on two different theoretical and statistical backgrounds are more appropriate. Furthermore the application of the qualitative sampling approach can be impractical for quantitative sampling as it could generate many analytical samples from a single seizure. In some countries the purity of the illicit drug in a seizure may affect the criminal sentence and therefore, reliable results for quantitative analysis are crucial.It was decided to investigate a new approach, which although incorporating some statistics also took account of our background knowledge about the composition of the drugs we were analysing. The ultimate goal was to produce recommendations for a practical sampling plan for quantitative analysis. It was found that the two key factors which had a significant effect on obtaining a representative analytical sample from a bulk seizure were the heterogeneity of the drug powder and the particle sizes of its components. This article concentrates on drug heterogeneity. Particle size effects will be addressed in part II of this study.A sampling plan was devised for a range of drug seizure types and asked ENFSI member laboratories to use it when analysing real drug seizures to provide heterogeneity data for the most common illicit drugs (heroin, cocaine, amphetamine, MDMA and cannabis (herbal and resin)). It was found that for routine quantitative drugs analysis, the sampling problems caused by heterogeneity can be solved by using an incremental sampling protocol. Furthermore, the number of increments that need to be taken for a particular drug is dependent on the relative standard deviation (RSD) required by an individual laboratory and the analytical method that they employ. A 1g increment size was found to be suitable for powdered drugs and cannabis resin. However, 1g increments were not suitable for herbal cannabis, because of particle size issues. Sampling of herbal cannabis will be addressed in Part II of this study. Recommendations for a sampling plan, based on the heterogeneity and particle size of specific drugs seizures in casework will be discussed in Part III of this study.

Effect of Cannabis sativa on oxidative stress and organ damage after systemic endotoxin administration in mice

The effect of Cannabis sativa extract on oxidative stress and organ tissue damage during systemic inflammation was studied. For this purpose, Swiss mice were challenged with a single intraperitoneal dose of lipopolysaccharide (LPS; 200 μg/kg) to mimic aspects of mild systemic infection. Cannabis resin extract (5, 10, or 20 mg/kg) (expressed as Δ9-tetrahydrocannabinol) was given via subcutaneous route for 2 days prior to and at the time of endotoxin administration. Mice were euthanized 4 h after LPS injection. Malondialdehyde (MDA), reduced glutathione (GSH), and nitric oxide (nitrite/nitrate) in the brain, liver, kidney, lung, and heart as well as brain glucose were measured. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) were measured in liver homogenates. Histopathological examination of different organs was performed, and immunohistochemical techniques were used to evaluate expression levels of inducible nitric oxide synthase (iNOS) and caspase-3 in the brain and liver. The administration of only cannabis (20 mg/kg) decreased MDA, increased GSH, and decreased glucose level in the brain. No significant effects were observed for cannabis alone on MDA, GSH, or nitric oxide in other organs or on liver enzymes. The administration of LPS increased MDA and nitric oxide, while GSH decreased in different organs. Brain glucose increased by endotoxin. AST, ALT, and ALP were markedly increased in the liver tissue. In LPS-treated mice, cannabis (20 mg/kg) decreased MDA. GSH increased in the brain, kidney, and lung, nitric oxide decreased in the brain and lung while brain glucose decreased after the highest dose of cannabis. Cannabis failed to alter the level of liver enzymes. Histological damage in the brain, kidney, heart, lung, and liver due to endotoxin is increased by cannabis. Increased immunoreactivity of caspase-3 in the cytoplasm of the hepatocytes was observed after LPS and cannabis cotreatment compared with the LPS only group. Caspase-3 immunoreactivity markedly increased in degenerating neurons of the cortex following cannabis and LPS cotreatment. iNOS inmmunoreactivity increased after LPS and more intense iNOS expression was detected in hepatocytes after cannabis and LPS cotreatment. iNOS expression increased after cannabis and LPS treatment especially in the cerebral cortex. Thus, the administration of cannabis decreased tissue oxidative stress but increased organ damage after endotoxin injection in mice.

Cloud point extraction of Δ9-tetrahydrocannabinol from cannabis resin

A cloud point extraction coupled with high performance liquid chromatography (HPLC/UV) method was developed for the determination of Δ(9)-tetrahydrocannabinol (THC) in micellar phase. The nonionic surfactant "Dowfax 20B102" was used to extract and pre-concentrate THC from cannabis resin, prior to its determination with a HPLC-UV system (diode array detector) with isocratic elution. The parameters and variables affecting the extraction were investigated. Under optimum conditions (1 wt.% Dowfax 20B102, 1 wt.% Na2SO4, T = 318 K, t = 30 min), this method yielded a quite satisfactory recovery rate (~81 %). The limit of detection was 0.04 μg mL(-1), and the relative standard deviation was less than 2 %. Compared with conventional solid-liquid extraction, this new method avoids the use of volatile organic solvents, therefore is environmentally safer.

Endocannabinoids in nervous system health and disease: the big picture in a nutshell

The psychoactive component of the cannabis resin and flowers, delta9-tetrahydrocannabinol (THC), was first isolated in 1964, and at least 70 other structurally related 'phytocannabinoid' compounds have since been identified. The serendipitous identification of a G-protein-coupled cannabinoid receptor at which THC is active in the brain heralded an explosion in cannabinoid research. Elements of the endocannabinoid system (ECS) comprise the cannabinoid receptors, a family of nascent lipid ligands, the 'endocannabinoids' and the machinery for their biosynthesis and metabolism. The function of the ECS is thus defined by modulation of these receptors, in particular, by two of the best-described ligands, 2-arachidonoyl glycerol and anandamide (arachidonylethanolamide). Research on the ECS has recently aroused enormous interest not only for the physiological functions, but also for the promising therapeutic potentials of drugs interfering with the activity of cannabinoid receptors. Many of the former relate to stress-recovery systems and to the maintenance of homeostatic balance. Among other functions, the ECS is involved in neuroprotection, modulation of nociception, regulation of motor activity, neurogenesis, synaptic plasticity and the control of certain phases of memory processing. In addition, the ECS acts to modulate the immune and inflammatory responses and to maintain a positive energy balance. This theme issue aims to provide the reader with an overview of ECS pharmacology, followed by discussions on the pivotal role of this system in the modulation of neurogenesis in the developing and adult organism, memory processes and synaptic plasticity, as well as in pathological pain and brain ageing. The volume will conclude with discussions that address the proposed therapeutic applications of targeting the ECS for the treatment of neurodegeneration, pain and mental illness.