Chemical Quantification of CBD and THC and Bacteriological Analysis of Artisan Cannabis Sativa Extracts: TheImportance of Regulation

doi:10.21203/rs.3.rs-3759988/v1

Objectives

Cannabis sativa is a plant species that has been used for millennia, especially in medicinal form, due to its great therapeutic potential. Tetrahydrocannabinol (THC) and cannabidiol (CBD) are the main pharmacologically active substances found in the plant. Efficient methods of extraction, purification and, sometimes, the separation of these cannabinoids are necessary to obtain extracts and compounds that can be used in the search for safe and effective therapeutic responses.

Methods

In this work, it was possible to analyze the composition of cannabinoids in 41 extracts of Cannabis sp. (32 produced by associations and 9 produced by the patients themselves), determining the amount of THC and CBD in the samples, using the High Performance Liquid Chromatography technique (RP-HPLC), as well as the presence of bacteria, using the Gram method.

Results

The median concentrations of THC and CBD measured in the samples were 51% and 58%, lower than the expected concentration, respectively. Furthermore, the presence of a Gram positive microorganism was observed in only one sample, suggestive of Staphylococcus Aureus.

Conclusion

Although associations have a noble social function, quality control in the production of medicines is important, with proper titration of cannabinoids and microbiological analysis of samples, which can be implemented through regulation by the Public Power.

Cannabidiol

Cannabis sativa

cannabinoids

HPLC

THC

Archaeological findings demonstrate that Cannabis sativa has existed for millennia. Records of the plant's pollen were found in northwest China and date back to 19.6 million years ago. The oldest fossil, found in Japan, dates from approximately 10,000 years before Christ (BC) (McPartland et al. 2019). Cannabis sativa spread across Eurasia and, thanks to the domestication process, reached different regions of the world. Its use has always been associated with food, religion and health (Pisanti and Bifulco 2018).

In Brazil, Cannabis sativa has been used medicinally for various purposes, such as chronic pain and neurological diseases. According to studies, the control of seizures in patients with refractory epilepsy is the most prevalent and effective therapeutic use of this plant. The pharmacological effects depend on the dose and on the interaction between plant components (cannabinoids, terpenes and flavonoids), which is called the entourage effect, or entourage effect. For refractory epilepsy, the use of oil extracts with cannabidiol (CBD) levels greater than tetrahydrocannabinol (THC), two of the main phytocannabinoids found in this plant, is the most indicated (Bitencourt et al. 2021; Costa et al. 2022).

CBD and THC are two of the many chemicals that have been isolated from Cannabis sp. and they are the main molecules responsible for the therapeutic properties, due to the interaction with the endocannabinoid receptors CB1, distributed in the Central Nervous System, and CB2, present in greater quantity in the Peripheral Nervous System (Russo 2011; Hill et al. 2011; Salentijn et al. 2015). Endocannabinoid receptors recognize the cannabinoids found in the plant, interact with them and thus provide therapeutic benefits, with anticonvulsant, analgesic, antiemetic and other properties (Zuardi et al. 2010).

The most common route of administration of the oil extract of Cannabis sp. is the sublingual route (MacCallum and Russo 2018). Such oils are produced by pharmaceutical companies, patient associations and by patients themselves. Currently, there are no regulations in Brazil for the due standardization of these extracts. In this way, it is important to investigate the quality of the process of cultivation and production of the medicine, aiming to verify the use of appropriate techniques, the titration of the substances present in the plant and the guarantee of a product free of contamination, thus meeting the needs of patients. Given the present context, the objective of this work was to evaluate the microbiological safety and quantify the THC and CBD content from Cannabis sativa extracts produced by hand by patients and associations from various regions of Brazil.

Materials and reagents

The samples were obtained by LabNeC (Laboratory of Behavioral Neuroscience) at the University of Southern Santa Catarina (UNISUL) and the analyzes were carried out in partnership with MULTILAB (Multi-user Laboratory of the Graduate Program in Health Sciences) at the University of Extremo South Santa Catarina (UNESC).

Certified reference materials were used as a standard for the THC and CBD chemical quantification method. The standards used were obtained at concentrations of 1 mg mL-1 (Sigma-Aldrich®). The equipment used was RP-HPLC, and the column used was Ascentis® C18 (25 cm x 2.1 mm 5 µm; Supelco®, USA). The temperature in the CTO-20A unit was 30°C and the composition of the mobile phase used was: (i) HPLC grade methanol; (ii) Milli-Q ultrapurified water. For bacteriological analysis, Mueller Hinton and Blood Agar plates were prepared. Positive samples on Blood and Mueller Agar plating were analyzed by gram stain.

Sampling and sample preparation

Medicinal extracts of Cannabis sp. (n = 41) were donated voluntarily by patient associations or patients with their own cultivation from February 2020 to December 2020. All extracts were produced in Brazil through the ethanolic extraction of flowers and/or leaves and stems of Cannabis sp., with subsequent evaporation of the solvent. The sample is presented in the form of a dark paste of high viscosity, which may contain insoluble particles, assuming colors that vary from light yellow to intense green. Due to the imprecision regarding the way of preparation and plant material of origin of each sample, it was not possible to define the cause, but associating the reports provided by the sample donors, factors such as the presence of leaves and time of exposure to the solvent can collaborate so that the extract end takes on different colors.

Aliquots of 100 mg for samples in oily vehicle and 20 mg for samples of resins (IFAVs) were transferred to polypropylene tubes and 10 mL of the solvent methanol:n-hexane (9:1 v/v) containing internal standard at the concentration of 0.005 mg mL-1. The samples were homogenized at 2000 rpm for 1 min and submitted to an ultrasound bath for 30 min. Samples in oily vehicle were refrigerated at -20°C for 30 min and centrifuged at 4000 rpm for 20 min. The resin samples diluted and homogenized by US and the supernatants of the centrifuged samples were filtered through a 0.22 µm PTVF membrane and transferred to RP-HPLC vials and adopted the appropriate dilution according to the chromatographic profile. For example, for extracts around 50 mg mL-1, the final dilution factor was 200 times.

For an evaluation and determination of cannabinoids, RP-HPLC is perfect for not requiring heating for its operation. The analyzes are carried out at temperatures close to the ambient temperature, without changing the molecular structure of the substances (Peschel and Politi 2015).

Chromatographic conditions

The analysis carried out in this work was adapted from De Backer et al (2009). The Shimadzu Prominence HPLC system equipment was used, UV-VIS SPD-20ª20A detector, analytical column Ascentis® C18 (250 cm mm x 2.1 mm 5 µm; Supelco®, USA). Milli-Q ultrapurified water mobile phase (A); HPLC grade methanol (B); Milli-Q ultrapurified water. The initial setting was 68% methanol (v/v), which was increased linearly until reaching 90.5% methanol over 35 minutes, then increased to 95% in 1 minute.

After three minutes, the column was adjusted to the initial condition and re-equilibrated under that condition for sixteen minutes. The total running time was 56 minutes. The flow rate was adjusted to 0.3 mL/min, the injection volume used was 30 µL. All experiments and analyzes were carried out at a temperature of 30°C.

Statistical analyzes

Variables are presented as mean ± standard deviation (SD), number and frequency. Statistical analysis was performed using the SPSS 21.0 statistical program. Data were assessed by one-way ANOVA, followed by Dunnet and / or Tukey post hoc (parametric data) and Kruskal-Wallis test, followed by Dunns post-tests (non-parametric data).

Cannabis sp. oily extracts were analyzed, 32 from Brazilian medical Cannabis associations and nine extracts produced by the patients themselves. Table 1 presents the specification regarding the expected dosage of the products (%, mg/ml), as informed by the association/patient, and the actual CBD and THC levels present in the samples, quantified using the RP-HPLC method. It is possible to observe that there is a greater number of extracts rich in THC, 24 samples, 7 samples of extracts rich in THC/CBD and 10 samples of extracts rich in CBD.

Table 1 CBD and THC concentration per analyzed sample
#	Origin	Feature	THC Expected (mg/ml)	THC Dosed (mg/ml)	Expected CBD (mg/ml)	Dosed CBD (mg/ml)
1	Association	Rich in THC	10	1.2	-	0.61
2	Association	Rich in THC	10	17.7	-	0.46
3	Association	Rich in THC	10	86.5	-	0.55
4	Association	Rich in THC	10	4	-	1.47
5	Association	Rich in THC	10	4.6	-	2.14
6	Association	Rich in THC	20	7.4	-	0.62
7	Association	Rich in THC	20	10.8	-	0.36
8	Association	Rich in THC	25	15.7	-	0.83
9	Association	Rich in THC	40	34.2	-	0.81
10	Association	Rich in THC	50	198.8	-	0.39
11	Association	Rich in THC	50	14.3	-	1.84
12	Association	Rich in THC	50	126.8	-	0.46
13	Association	Rich in THC	70	116.7	-	3.71
14	Association	Rich in THC	100	53.3	-	9.98
15	Association	Rich in THC	100	103.8	-	1.95
16	Association	Rich in THC	100	140.2	-	0.33
17	Association	Rich in CBD	-	0.46	10	8.1
18	Association	Rich in CBD	-	8.65	10	3.9
19	Association	Rich in CBD	-	0.74	20	19.5
20	Association	Rich in CBD	-	1.08	40	21.5
21	Association	Rich in CBD	-	1.57	50	36.8
22	Association	Rich in CBD	-	1.38	50	66.2
23	Association	Rich in CBD	-	19.88	100	4.6
24	Association	Rich in CBD	-	17.8	100	NM*
25	Association	Rich in CBD	-	3.42	100	3.3
26	Association	Rich in THC/CBD	5	8.49	5	2.15
27	Association	Rich in THC/CBD	10	4.99	10	3.68
28	Association	Rich in THC/CBD	10	12.67	10	4.99
29	Association	Rich in THC/CBD	20	8.34	20	6.62
30	Association	Rich in THC/CBD	25	35.07	25	10.14
31	Association	Rich in THC/CBD	50	83.9	50	20.83
32	Association	Rich in THC/CBD	50	23.71	50	17.01
33	Patient	Rich in THC	10	3.01	-	0.31
34	Patient	Rich in THC	10	1.43	-	0.46
35	Patient	Rich in THC	10	0.87	-	0.21
36	Patient	Rich in THC	10	6.02	-	0.38
37	Patient	Rich in THC	10	1.68	-	0.21
38	Patient	Rich in THC	15	0.41	-	0
39	Patient	Rich in THC	25	0.84	-	0.21
40	Patient	Rich in THC	30	0.35	-	0.2
41	Patient	Rich in CBD	-	1.77	10	16.87

*Not Measurable

Source: elaborated by the author (2023)

When analyzing the extracts separately, according to the cannabinoid present, the proportion of measured and expected THC and CBD is shown in Fig. 1.

The THC-rich extracts had a median THC concentration of 51% lower than expected values. CBD-rich extracts, on the other hand, had a median of 58% lower than expected CBD concentration.

Table 2 shows the absolute and relative difference of THC and CBD in the extracts. Only two extracts rich in THC showed a dosed value greater than the expected value, the others showed an absolute and relative difference of THC below the expected value. Only one extract rich in CBD showed a dosed value greater than the expected value, the others showed an absolute and relative difference of CBD below the expected value. In the extracts rich in THC/CBD, four extracts presented a measured THC value greater than the expected value, the others presented an absolute and relative difference of THC below the expected value.

Table 2

Absolute and relative difference of THC and CBD between analyzed samples
#	Origin	Feature	Absolute THC Difference (mg/ml)	Relative THC Difference (%)	Absolute CBD Difference (mg/ml)	Relative CBD Difference (%)
1	Association	Rich in THC	-4.67	-46.70	-	-
2	Association	Rich in THC	-6.75	-67.50	-	-
3	Association	Rich in THC	-1.28	-12.80	-	-
4	Association	Rich in THC	2.68	26.80	-	-
5	Association	Rich in THC	1.67	16.70	-	-
6	Association	Rich in THC	-9.62	-48.10	-	-
7	Association	Rich in THC	-11.83	-59.15	-	-
8	Association	Rich in THC	-10.98	-43.92	-	-
9	Association	Rich in THC	-23.19	-57.98	-	-
10	Association	Rich in THC	-47.78	-95.56	-	-
11	Association	Rich in THC	-1.55	-3.10	-	-
12	Association	Rich in THC	-43.56	-87.12	-	-
13	Association	Rich in THC	-2.65	-3.79	-	-
14	Association	Rich in THC	30.87	30.87	-	-
15	Association	Rich in THC	-38.8	-38.80	-	-
16	Association	Rich in THC	-85.66	-85.66	-	-
17	Patient	Rich in THC	-6.99	-69.90	-	-
18	Patient	Rich in THC	-8.57	-85.70	-	-
19	Patient	Rich in THC	-9.13	-91.30	-	-
20	Patient	Rich in THC	-3.98	-39.80	-	-
21	Patient	Rich in THC	-8.32	-83.20	-	-
22	Patient	Rich in THC	-14.59	-97.27	-	-
23	Patient	Rich in THC	-24.16	-96.64	-	-
24	Patient	Rich in THC	-29.65	-98.83	-	-
25	Association	Rich in CBD	-	-	-3.00	-30.00
26	Association	Rich in CBD	-	-	-9.66	-96.60
27	Association	Rich in CBD	-	-	-9.90	-49.50
28	Association	Rich in CBD	-	-	-18.45	-46.13
29	Association	Rich in CBD	-	-	-49.78	-99.56
30	Association	Rich in CBD	-	-	-24.58	-49.16
31	Association	Rich in CBD	-	-	-99.30	-99.30
32	Association	Rich in CBD	-	-	-7.03	-7.03
33	Association	Rich in CBD	-	-	-61.88	-61.88
34	Patient	Rich in CBD	-	-	6.87	68.7
35	Association	Rich in THC/CBD	3.49	69.8	-2.85	-57
36	Association	Rich in THC/CBD	-5.01	-50.1	-6.32	-63.2
37	Association	Rich in THC/CBD	2.67	26.7	-5.01	-50.1
38	Association	Rich in THC/CBD	-11.66	-58.3	-13.38	-66.9
39	Association	Rich in THC/CBD	10.07	40.28	-14.86	-59.44
40	Association	Rich in THC/CBD	33.9	67.8	-29.17	-58.34
41	Association	Rich in THC/CBD	-26.29	-52.58	-32.99	-65.98
Source: elaborated by the author (2023)
Source: elaborated by the author (2023)
Source: elaborated by the author (2023)
Source: elaborated by the author (2023)

In Figs. 2 and 3 it is possible to observe the absolute and relative difference of THC and CBD in the samples, respectively. Furthermore, when performing the Mann-Whitney test and analyzing the distribution of the relative difference in THC in relation to the way the extract was produced, via association or patient, there was a statistically significant difference (p < 0.005), with smaller relative differences in THC in the extracts from associations.

Regarding the presence of microorganisms, the presence of bacterial growth was observed in Muller Hinton Agar (yellow) and in Blood Agar (red), after 24 hours of incubation of sample 7, rich in THC. The analysis of the Gram staining result of sample 07 showed the presence of a Gram-positive bacterium, with colony characteristics compatible with Staphylococcus aureus.

In this work, it was possible to analyze the composition of cannabinoids in 41 extracts of Cannabis sp. (32 produced by associations and 9 produced by the patients themselves), determining the amount of THC and CBD in the samples, which presented a median of 51% and 58% of lower concentration in relation to the expected values, respectively. Furthermore, the presence of a Gram-positive microorganism was observed in only one sample, suggestive of Staphylococcus Aureus.

An Argentine study published recently, in 2022, analyzed the composition of artisanal extracts of Cannabis sativa and revealed that the concentration of cannabinoids found was lower than expected, especially CBD and, even, in some extracts, the presence of no cannabinoid (Manzo et al. 2022). Two other studies, one from Brazil and one from the United States, compared the amount of cannabinoids in artisanal extracts and imported extracts and observed a lower concentration of cannabinoids, especially CBD in artisanal samples. Another relevant point recorded was the predominance of THC in artisanal extracts, which is possibly related to the consumption profile, the lack of regulation and the difficult access to plants rich in CBD (Carvalho et al. 2020; Miller et al. 2022).

Along the same lines, a third study, published in 2022, evaluated the variability of blood concentrations of cannabinoids in patients who used artisanal and imported extracts. The concentration of CBD in the bloodstream of participants using artisanal oil was lower than that observed in those using imported medicine (Cohen et al. 2022). However, according to researchers, artisanal extracts are effective and safe, especially in the treatment of refractory epilepsy, which ensures the importance of access to artisanal extract as a treatment option (Tzadok et al. 2022; Sulak et al. 2017).

In the present study, higher concentrations of THC (lower relative difference) were also observed in the extracts from the associations when compared with the extracts produced by the patients themselves. This fact is possibly associated with the quality of the raw material and the extraction technique used, which elucidates the need for proper regulation and, therefore, instruction on the production methods of Cannabis sp extract. for patients, thus improving the quality of medicines (Figueiredo 2019; Azevedo 2020).

Furthermore, when relating the presence of bacteria in the extracts by the Gram method, the presence in only one sample indicated a low rate of contamination, which is the opposite of the results found by the Oswaldo Cruz Foundation, which evaluated some artisanal extracts and concluded that 40% of the extracts were contaminated with some bacteria or fungus (such as: Candida albicans, Bacillus mycoides, Aspergillus brasiliensis, Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli). It is likely that investigating only the presence of bacteria using the Gram method has contributed to the findings presented in this article, given the wide variety of microorganisms investigated in the comparative study (Carmo 2019).

Currently, in Brazil, there are several drugs based on CBD and THC available in drugstores, such as: Cannabidiol Prati Donaduzzi 200mg/ml and Mevatyl (THC 27mg/ml and CBD 25mg/ml), which are products that have a high cost, around BRL 2,000 to BRL 3,000 reais, which makes it difficult for patients to access treatment (National Health Surveillance Agency 2017). Therefore, the presence of associations and artisanal production of oils by users is crucial for the medicine to be accessible to the population. There are three associations that can legally produce the medicine based on Cannabis sp. in Brazil, namely: ABRACE, Brazilian Cannabis Support Association Esperança, which has its headquarters in João Pessoa (PB); APEPI, Support and Research for Cannabis Patients, in the city of Rio de Janeiro (RJ); and Cultive, in the city of São Paulo (SP) (National Health Surveillance Agency 2022; Figueiredo 2019; Legal Consultant 2021). In addition to the production and supply of the drug, associations represent the largest support platform for patients and their families, playing a key role in articulating demands for legal access to Cannabis, providing contact between patients, doctors and lawyers. Therefore, they provide an essential service to all those involved and interested in the medicinal potential of the plant (Figueiredo 2019; Azevedo 2020).

It is worth mentioning that the medicinal extracts of Cannabis sp. are produced, especially, with the flowers of pistillate specimens (female) due to the higher concentration of cannabinoids (Lewis-Bakker et al. 2019). Cannabis extraction is a very old process, used to remove substances from the plant (cannabinoids, terpenes and flavonoids) that can be used for various treatments. The methods used for extraction can vary and it is important to avoid the use of heat due to the sensitivity of cannabinoids to high temperatures. There are numerous methods used for Cannabis extraction, such as: Rosin method; Butane extraction; extraction by CO2 and, the most used artisanally, extraction by means of isopropyl alcohol, in which the flowers, previously dried, are submerged in the solvent, in a saturation process, and the extract is filtered and goes on to the distillation process. Next, an oily vehicle (for example, corn oil and medium-chain triglyceride) is added, with the appropriate titration of cannabinoids (Ferreira and Filev 2020; Liang et al. 2010).

Therefore, it is evident that it is important to promote the quality of the process of cultivation and production of medicines derived from Cannabis sp., including the standardization of techniques, titration of substances present in the plant and the guarantee of a product free of contamination, with the proper regulation implemented, thus meeting the needs of patients and promoting greater care for their health (Carvalho et al. 2020; Legal Consultant 2021).

As for the limitations of this study, it is worth mentioning the low number of samples of artisanal extracts analyzed, the non-quantification of other cannabinoids, terpenes and flavonoids and the absence of analysis of the mycological profile and of other bacteria in the samples. We have no conflicts of interest for the preparation of this work.

The analysis of substances present in products based on extracts of Cannabis sp. it is essential for the treatment of several clinical conditions to be effective and safe. Therefore, it is indicated that the producers of such drugs carry out studies capable of quantifying the cannabinoids, terpenes and flavonoids present in the extract extracted from the plant, thus aiming at a reliable and effective quality control.

In Brazil, there is a lack of scientific research related to the dosage of products derived from Cannabis sp. and its extracts. For all this to be possible, it is necessary that the public agents responsible for regulation facilitate access to plants for scientific research, thus increasing the quality control of medicines produced by medical Cannabis associations and by the patients themselves.

Ethics approval

Not applicable.

Conflict of interest

The authors declare no competing interests.

Funding

This work was carried out with the support of the Coordination for the Improvement of Higher Education Personnel - Brazil (CAPES) - Financing Code 001.

Author contribution

PAC and LRNJ conception, design of the study, acquisition of data, analysis and interpretation of data, drafting and revising, final approval; LMS writing; RS analyzes HPLC; EP microbiological analysis; SSR writing review; KSK mathematical calculations; PAA writing review; FKR writing review; RAMA laboratory support; RMB writing review, final approval.

Acknowledgements

This work was carried out with the support of the Coordination for the Improvement of Higher Education Personnel-Brazil (CAPES).

Azevedo CF (2020) Legal access to medical cannabis in Brazil: a fundamental right. Dissertation, Federal University of Santa Catarina, Florianopolis.
Benjamin De Backer. Innovative development and validation of an HPLC/DAD method for the qualitative and quantitative determination of major cannabinoids in cannabis plant material. Journal of Chromatography B, 877, 4114-4124, 2009. Com adaptações.
Bitencourt RM, Takahashi RN, Carlini EA (2021) From an Alternative Medicine to a New Treatment for Refractory Epilepsies: Can Cannabidiol Follow the Same Path to Treat Neuropsychiatric Disorders?. Front Psychiatry 12:638032. https://doi.org/10.3389/fpsyt.2021.638032
Carmo JS (2019) Evaluation of the microbiological quality of flowers and medicinal extracts of Cannabis sativa. Dissertation, Oswaldo Cruz Foundation, Rio de Janeiro.
Carvalho VM, Aguiar AFL, Baratto LC, Souza FLC, Rocha ED (2020) Quantification of cannabinoids in medicinal cannabis extracts by high performance liquid chromatography. Quim Nova 43(1):90–7. https://doi.org/10.21577/0100-4042.20170457
Cohen NT, Bahar B, Conry JA, Schreiber JM (2022)Variability in Serum Concentrations and Clinical Response in Artisanal Versus Pharmaceutical Cannabidiol Treatment of Pediatric Pharmacoresistant Epilepsy. J Pediatr Pharmacol Ther 27(6):558–63. https://doi.org/10.5863/1551-6776-27.6.558 _
Costa PA, Junior LRN, Silva LM, Bitencourt RM (2022) Full-spectrum or purified CBD: which is the best treatment for epilepsy?. Neuroscience Magazine 30:1-4. https://doi.org/10.34024/rnc.2022.v30.12864
Ferreira R, Filev R (2020) Artisanal Extraction Methods for Cannabis. Introduction to Cannabis Associativism 76–85.
Figueiredo E (2019) The Production of the Legal Truth about Cannabis in Brazil. Representative body. https://www2.camara.leg.br. Accessed 2022.
Hill AJ, Williams CM, Whalley BJ, Stephens GJ (2012) Phytocannabinoids as novel therapeutic agents in CNS disorders. Pharmacol Therapeut 133(1):79–97. https://doi.org/10.1016/j.pharmthera.2011.09.002 _
Legal Consultant – ConJur (2021) Judge grants collective HC for association to plant marijuana for medicinal purposes. https://www.conjur.com.br/2021-fev-13/associacao-autorizada-plantar-maconha-fins-mediciais. Accessed 2022.
Lewis-Bakker MM, Yang Y, Vyawahare R, Kotra LP (2019) Extractions of Medical Cannabis Cultivars and the Role of Decarboxylation in Optimal Receptor Responses. Cannabis Cannabinoid Res 4(3):183–94. https://doi.org/10.1089/can.2018.0067
Liang YZ, Xie PS, Chan K (2010) Chromatographic Fingerprinting and Metabolomics for Quality Control of TCM. Comb Chem High T Scr 13(10):943–53. https://doi.org/10.2174/138620710793360310.
MacCallum CA, Russo EB (2018) Practical considerations in medical cannabis administration and dosing. Eur J Intern Med 49:12–9. https://doi.org/10.1016/j.ejim.2018.01.004
Manzo PG, Martin S, Uema S, Charles G, Bruni FM, Montoya SN, Bertotto ME, Eynard M, Armando P, Fierro CB (2022) Characterization of the problem of the therapeutic use of Cannabis Oil in Córdoba, Argentina. Rev Fac Cien Med Univ Nac Cordoba 79(2):123-31. https://doi.org/10.31053/1853.0605.v79.n2.30922 _
McPartland JM, Hegman W, Long T (2019) Cannabis in Asia: its center of origin and early cultivation, based on a synthesis of subfossil pollen and archaeobotanical studies. Veget Hist Archaeobot 28:691-702. https://doi.org/10.1007/s00334-019-00731-8
Miller OS, Junior EJE, Jones KJ, Gidal BE (2022) Analysis of cannabidiol (CBD) and THC in nonprescription consumer products: Implications for patients and practitioners. Epilepsy Behav 127:108514. https://doi.org/10.1016/j.yebeh.2021.108514
National Health Surveillance Agency (2017) RDC Resolution nº 156. Official Diary of the Union. https://www.in.gov.br/materia/-/asset_publisher/Kujrw0TZC2Mb/content/id/20198336/do1-2017-05-08-resolucao-rdc-n-156-de- 5-May-2017-20198229. Accessed 2022.
National Health Surveillance Agency (2022) RDC Resolution nº 660. Official Diary of the Union. https://www.in.gov.br/en/web/dou/-/resolucao-rdc-n-660-de-30-de-marco-de-2022-389908959. Accessed 2022.
Peschel W, Politi M (2015) H NMR HPLC/DAD for Cannabis sativa L. chemotype distinction, extract profiling and specification. Talanta 140:150–65. https://doi.org/10.1016/j.talanta.2015.02.040
Pisanti S, Bifulco M (2018) Medical Cannabis: A plurimillennial history of an evergreen. J Cell Physiol 234(6):8342-51. https://doi.org/10.1002/jcp.27725 _
Russo EB (2011) Taming THC: potential cannabis synergy and phytocannabinoid-terpenoid entourage effects. Brit J Pharmacol 163(7):1344-64. https://doi.org/10.1111/j.1476-5381.2011.01238.x_
Salentijn EMJ, Zhang Q, Amaducci S, Yang M, Trindade LM (2015) New developments in fiber hemp (Cannabis sativa L.) breeding. Ind Crop Prod 68:32–41. https://doi.org/10.1016/j.indcrop.2014.08.011
Sulak D, Saneto R, Goldstein B (2017) The current status of artisanal cannabis for the treatment of epilepsy in the United States. Epilepsy Behav 70(b):328-33. https://doi.org/10.1016/j.yebeh.2016.12.032
Tzadok M, Hamed N, Heimer G, Zohar-Dayan E, Rabinowicz S, Zeev BB (2022) The Long-Term Effectiveness and Safety of Cannabidiol-Enriched Oil in Children With Drug-Resistant Epilepsy. Pediatr Neurol 136:15-9. https://doi.org/10.1016/j.pediatrneurol.2022.06.016
Zuardi AW, Crippa JAS, Hallak JEC (2010) Cannabis sativa: The plant that can produce unwanted effects and also treat them. Braz J Psychiatry 32:51–2. https://doi.org/10.1590/S1516-44462010000500001

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Chemical Quantification of CBD and THC and Bacteriological Analysis of Artisan Cannabis Sativa Extracts: TheImportance of Regulation

Status:

Version 1

Abstract

Objectives

Methods

Results

Conclusion

Figures

INTRODUCTION

MATERIALS AND METHODS

Materials and reagents

Sampling and sample preparation

Chromatographic conditions

Statistical analyzes

RESULTS

DISCUSSION

CONCLUSION

Declarations