Monitoring Medical Cannabis

Giuseppe Cannazza and Cinzia Citti from the University of Modena and Reggio Emilia discuss the growing interest in the use of medicinal cannabis in Europe and the analytical challenges involved. - Interview by Alasdair Matheson

Q. Cannabis for medicinal purposes has gained a lot of attention in the US recently. What is the situation for medicinal cannabis in Europe?

Giusepppe Cannazza: In Europe there is a growing interest in medicinal cannabis, and cannabis is used to treat a series of pathologies. Nonetheless, cannabis is a controlled drug under specific regulations and is illegal even for therapeutic purposes in many European countries. At present, the only European nations implementing state-controlled medicinal cannabis production are Italy, the Netherlands, and the Czech Republic. 

The Netherlands has awarded the sole license to produce medicinal cannabis to the company Bedrocan BV. Although the UK is not among the countries producing medicinal cannabis, GW Pharmaceuticals has the license to grow cannabis and manufacture Sativex, which is not distributed as whole-plant, herbal cannabis flowers, but as an extract with a known amount of tetrahydrocannabinol (THC) and cannabidiol (CBD) in a 1:1 ratio. 

In the Czech Republic, the logistics company Elkoplast is licensed to produce medicinal cannabis using Bedrocan strains. In Italy, the army recently completed its first crop of cannabis intended for distribution to patients with a doctor’s prescription for medicinal cannabis. Therefore, the use of medicinal cannabis is currently evolving in Europe and the production is expected to increase going forward.

Q. What treatments and in what forms is it being used for medicinal purposes in Europe?

GC: Certified medicinal cannabis is currently used for the treatment of a number of pathologies, including multiple sclerosis, epilepsy, neuropathic pain, arthritis, depression, anxiety disorders, sleep disorders, psychosis, glaucoma, and Tourette’s syndrome, or for the relief of symptoms such as nausea and vomiting as a result of chemotherapy, and appetite stimulation in HIV and AIDS. There are numerous studies in the scientific literature testifying the efficacy of cannabis derivatives for therapeutic use (1,2,3,4). The most popular forms of delivery are vaporization, herbal tea, and oral spray. 

Q. Are there problems associated with sample preparation?

Cinzia Citti: The issue with sample preparation, if we intend the preparation of cannabis extracts to be delivered to patients, is that there is no standardized protocol for the extraction. It is clear that, in this context, the pharmacist who has to prepare the extracts can use different methods. This creates a great variability in the chemical composition of the galenical preparation in terms of the substances contained and their concentration, and, as a result, in the therapeutic effect.

Q. Your group recently developed a method to identify the principal cannabinoids: cannabidiolic acid (CBDA), cannabidiol (CBD), cannabinol (CBN), tetrahydrocannabinol (THC), and tetrehydrocannabinolic acid (THCA) (5). How did this project arise and what were the main challenges you had to overcome?

GC: Our project was born after the publication of a recent Italian law decree that has imposed the quantitative determination of the main cannabinoids in each cannabis‑based galenical preparation. The main issue in this scenario is the lack of a standardized analytical method for this specific purpose in galenical preparations. All published methods are suitable for cannabis plant material and not for pharmaceutical preparations. The matrix makes the difference in the analysis. Moreover, the methods reported in the literature are relatively long and too complicated to be reproduced by a pharmacist.

Q. What is novel about your approach compared to previous methods?

CC: Our method is highly selective and specific towards the main cannabinoids under investigation; it is rapid, and requires less than 10 min for the analysis; it is very sensitive, because it can reveal even traces of cannabinoids (0.1 μg/mL); and lastly, it is very simple to be applied in a pharmacy not only for the analysis but also for the sample preparation, which involves only a simple dilution of a minimum amount (50 μL) of extract.

Q. What were your main findings?

CC: The main goal achieved in our study is that it can be considered the first analytical method for the qualitative and quantitative determination of the main cannabinoids in galenical preparations. The method was tested with two types of detectors: ultraviolet (UV) and mass spectrometry (MS). The Italian legislation imposes the use of mass spectrometry as the only specific detector, which is very expensive for a pharmacist and requires qualified personnel for the analysis. 

We demonstrated that UV and mass spectrometry can be equally employed because they provided the same quantitative results (5). Therefore, it is more convenient for a pharmacist to buy and use a liquid chromatograph interfaced with a UV detector. These are not the only results we obtained: in the paper that we published (5), we proposed an alternative extraction method for medicinal cannabis. I will be more specific: cannabis is not only cannabinoids, but also a lot of other substances, including terpenes, which are volatile compounds thought to exert a synergic action with cannabinoids. Our extraction procedure proposes the use of a condenser, which is an apparatus that brings the vapours produced by the heat of cannabis inflorescence back to the extraction mixture. This prevents the loss of the terpene volatile component of cannabis, which is extremely important for the overall pharmaceutical effect.

Q. Do you have any advice for analysts embarking on analyzing cannabis?

GC: We suggest paying specific attention to the storage of analytical standards. In particular to the acid forms, such as CBDA, THCA, and CBGA because they are extremely sensitive to heat. In fact, they can easily decarboxylate (losing carbon dioxide), leading to the formation of their neutral derivatives, CBD, THC, and cannabigerol (CBG). Moreover, we observed the presence of an additional compound in the CBD/methanol solution by liquid chromatography (LC)–MS with a molecular weight different from that of CBD, which is likely to be derived from a reaction of the latter with methanol.

Another important point is in relation to the coelution of CBDA with CBGA and CBG with CBD, which is hardly detectable by UV detection alone. In this case, it is important either to use a mass spectrometer to allow their separation based on their m/z or to have a highly efficient column able to distinguish between the two pairs of cannabinoids. Therefore, we can say that UV is always appropriate, considering that it is the cheapest and easiest method of detection. However, I think it is necessary to check for any interfering (coeluting) compound by MS before validating a quantitative method.

Q. Does your group intend to explore cannabis analysis further?

CC: We have already started to explore cannabis from other points of view. In particular, we are exploring the chemical variability of cannabis extracts when different solvents and times of extraction are involved applying a metabolomics approach. We also published a review in the Journal of Pharmaceutical and Biomedical Analysis on the analysis of cannabis in both plant material and biological matrices (6). Our research is now focused on the investigation of the comprehensive chemical composition of cannabis extracts.

We believe that the field of cannabis with regards to its analytical, chemical, and pharmaceutical aspects needs to be explored further. Very little is known about the biological activity of cannabinoid acids and other minor cannabinoids. As the use of this interesting plant is spreading in a plethora of applications, such as food, textile, medicinal, and recreational, it is necessary to embark on an in-depth study of all its facets.

References

1. B.S. Koppel et al., Neurology82, 1556–1563 (2014). 
2. P.F. Whiting et al., JAMA 313, 2456–2473 (2015). 
3. O. Devinsky et al., Lancet Neurol. 15, 270–278 (2016). 
4. L.M. Borgelt et al., Pharmacotherapy: The Journal of Human Pharmacology and Drug Therapy 33, 195–209 (2013). 
5. C. Citti, G. Ciccarella, D. Braghiroli, C. Parenti, M.A. Vandelli, and G. Cannazza, Journal of Pharmaceutical and Biomedical Analysis 128, 201–209 (2016).
6. C. Citti, D. Braghiroli, M.A. Vandelli, and G. Cannazza, Journal of Pharmaceutical and Biomedical Analysis, DOI: 10.1016/j.jpba.2017.06.003.

Giuseppe Cannazza is a researcher in medicinal chemistry at the Department of Life Science of the University of Modena and Reggio Emilia (Italy). His expertise in pharmaceutical chemistry and analytical sciences is testified by a long experience in Italy and abroad (University of Graz, Austria, and National Institute of Aging, NIH, Baltimore, USA). His scientific interests include the development of analytical methods for the evaluation of endogenous metabolites in biological fluids and plant extracts, their stereochemical configuration, and stereo- and chemical stability. He is the author of more than 50 papers in peer‑reviewed scientific journals.

Cinzia Citti is a postdoctoral research sassistant at the University of Modena and Reggio Emilia (Italy) with previous experience at the University of Salento and the School of Chemistry at the University of Bristol, UK. After a few years dedicated to synthetic chemistry, she turned to analytical and pharmaceutical chemistry collaborating with Giuseppe Cannazza. Her interest span metabolomics for the study of drugs effects on the central nervous system, chemistry of cannabis, and stereochemistry of drug pharmacokinetics.