Difference between revisions of "Journal:Fast detection of 10 cannabinoids by RP-HPLC-UV method in Cannabis sativa L."
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==Abstract== | ==Abstract== | ||
[[wikipedia:Cannabis | [[wikipedia:Cannabis|Cannabis]] has regained much attention as a result of updated legislation authorizing many different uses, and it can be classified on the basis of the content of [[wikipedia:Tetrahydrocannabinol|Δ9-tetrahydrocannabinol]] (Δ9-THC), a psychotropic substance for which there are legal limitations in many countries. For this purpose, accurate qualitative and quantitative determination is essential. The relationship between THC and [[wikipedia:Cannabidiol|cannabidiol]] (CBD) is also significant, as the latter substance is endowed with many specific and non-psychoactive proprieties. For these reasons, it becomes increasingly important and urgent to utilize fast, easy, validated, and harmonized procedures for determination of [[wikipedia:Cannabinoid|cannabinoids]]. The procedure described herein allows rapid determination of 10 cannabinoids from the [[wikipedia:Inflorescence|inflorescences]] of ''Cannabis sativa'' L. by extraction with organic solvents. Separation and subsequent detection are by [[wikipedia:Reversed-phase chromatography|reversed-phase]] [[high-performance liquid chromatography]] with ultraviolet detector (RP-HPLC-UV). Quantification is performed by an external standard method through the construction of calibration curves using pure standard [[Chromatography|chromatographic]] reference compounds. The main cannabinoids dosed (g/100 g) in actual samples were cannabidiolic acid (CBDA), CBD, and Δ9-THC (Sample L11 CBDA 0.88 ± 0.04, CBD 0.48 ± 0.02, Δ9-THC 0.06 ± 0.00; Sample L5 CBDA 0.93 ± 0.06, CBD 0.45 ± 0.03, Δ9-THC 0.06 ± 0.00). The present validated RP-HPLC-UV method allows determination of the main cannabinoids in ''Cannabis sativa'' L. inflorescences and appropriate legal classification of it as either [[wikipedia:Hemp|hemp]] or a [[wikipedia:Cannabis (drug)|drug-type]]. | ||
'''Keywords''': cannabinoids, ''Cannabis sativa'' L., HPLC, validation | '''Keywords''': cannabinoids, ''Cannabis sativa'' L., HPLC, validation | ||
==Introduction== | |||
''[[wikipedia:Cannabis|Cannabis]]'' is classified into the family of Cannabaceae and initially encompassed three main species: ''[[wikipedia:Cannabis sativa|Cannabis sativa]]'', ''[[wikipedia:Cannabis indica|Cannabis indica]]'', and ''[[wikipedia:Cannabis ruderalis|Cannabis ruderalis]]''.<ref name="MontserratHemp14">{{cite journal |title=Hemp (''Cannabis sativa'' L.) seed oil: Analytical and phytochemical characterization of the unsaponifiable fraction |journal=Journal of Agricultural and Food Chemistry |author=Montserrat-de la Paz, S.; Marín-Aguilar, F.; García-Giménez, M.D. et al. |volume=62 |issue=5 |pages=1105–10 |year=2014 |doi=10.1021/jf404278q |pmid=24422510}}</ref> Nowadays, ''Cannabis'' has only one species due to continuous crossbreeding of the three species to generate hybrids. In fact, all plants are categorized as belonging to ''Cannabis sativa'' and classified into chemotypes based on the concentration of the main [[wikipedia:Cannabinoid|cannabinoids]]. Depending on the ratio of [[wikipedia:Tetrahydrocannabinolic acid||tetrahydrocannabinolic acid]] (THCA) to cannabidiolic acid (CBDA) (the THCA/CBDA ratio), some chemotypes have been distinguished. In particular, chemotype I or “drug plants” have a TCHA/CBDA ratio >1.0, chemotype II plants exhibit an intermediate ratio, chemotype III or “fiber plants” have a THCA/CBDA ratio <1.0, chemotype IV plants contain cannabigerolic acid (CBGA) as the main cannabinoid, and chemotype V plants contain almost no cannabinoids.<ref name="AppendinoCannab11">{{cite journal |title=Cannabinoids: Occurrence and medicinal chemistry |journal=Current Medicinal Chemistry |author=Appendino, G.; Chianese, G.; Taglialatela-Scafati, O. |volume=18 |issue=7 |pages=1085–99 |year=2011 |doi=10.2174/092986711794940888 |pmid=21254969}}</ref><ref name="AndreCannabis16">{{cite journal |title=''Cannabis sativa'': The Plant of the Thousand and One Molecules |journal=Frontiers in Plant Science |author=Andre, C.M.; Hausman, J.F.; Guerriero, G. |volume=7 |pages=19 |year=2016 |doi=10.3389/fpls.2016.00019 |pmid=26870049 |pmc=PMC4740396}}</ref><ref name="Aizpurua-OlaizolaEvo16">{{cite journal |title=Evolution of the Cannabinoid and Terpene Content during the Growth of Cannabis sativa Plants from Different Chemotypes |journal=Journal of Natural Products |author=Aizpurua-Olaizola, O.; Soydaner, U;. Öztürk, E. et al. |volume=79 |issue=2 |pages=324–31 |year=2016 |doi=10.1021/acs.jnatprod.5b00949 |pmid=26836472}}</ref><ref name="BrighentiDevelop17">{{cite journal |title=Development of a new extraction technique and HPLC method for the analysis of non-psychoactive cannabinoids in fibre-type Cannabis sativa L. (hemp) |journal=Journal of Pharmaceutical and Biomedical Analysis |author=Brighenti, V.; Pellati, F.; Steinbach, M. et al. |volume=143 |pages=228–36 |year=2017 |doi=10.1016/j.jpba.2017.05.049 |pmid=28609672}}</ref> | |||
In Italy, the interest in ''Cannabis sativa'' L. has increased recently, mainly due to December 2016 legislation (Legge 2 Dicembre 2016, n. 242).<ref name="ItalyLegge16">{{cite web |url=https://www.gazzettaufficiale.it/eli/id/2016/12/30/16G00258/sg |title=Legge 2 Dicembre 2016, n. 242 - Disposizioni per la promozione della coltivazione e della filiera agroindustriale della canapa |work=Gazzetta Ufficiale della Repubblica Italiana |date=30 December 2016 |accessdate=01 June 2019}}</ref> As a result, requests have been made to develop cost-effective and easy-to-use quantitative and qualitative methods for analysis of cannabinoids. | |||
The Italian regulatory framework has classified two types of ''Cannabis sativa'' L. depending on the content of [[wikipedia:Tetrahydrocannabinol|Δ9-tetrahydrocannabinol]] (Δ9-THC). In particular, fiber-type plants of ''Cannabis sativa'' L., also called “hemp,” are characterized by a low content of Δ9-THC (<0.2% w/w). If the content of Δ9-THC is >0.6% w/w, it is considered as drug-type, also called “therapeutic” or “marijuana.” | |||
Industrial hemp is used in several sectors, such as in the pharmaceutical, cosmetic, food, and textile industries, as well as in energy production and building. In general, fiber-type plants are less used in the pharmaceutical field, where drug-type plants are more often employed.<ref name="BrighentiDevelop17" /> However, there is also an increased interest in hemp varieties containing non-psychoactive compounds. In fact, the European Union has approved 69 varieties of ''Cannabis sativa'' L. for commercial use.<ref name="EUPlant19">{{cite web |url=http://ec.europa.eu/food/plant/plant_propagation_material/plant_variety_catalogues_databases/search//public/index.cfm |title=EU Plant variety database |publisher=European Commission |date=2019 |accessdate=01 June 2019}}</ref> | |||
Revision as of 21:15, 2 December 2019
| Full article title | Fast detection of 10 cannabinoids by RP-HPLC-UV method in Cannabis sativa L. |
|---|---|
| Journal | Molecules |
| Author(s) | Mandrioli, Mara; Tura, Matilde; Scotti, Stefano; Toschi, Tullia Gallina |
| Author affiliation(s) | University of Bologna, Shimadzu Italia |
| Primary contact | Email: tullia dot gallinatoschi at unibo dot it |
| Editors | Locatelli, Marcello |
| Year published | 2019 |
| Volume and issue | 24(11) |
| Page(s) | 2113 |
| DOI | 10.3390/molecules24112113 |
| ISSN | 1420-3049 |
| Distribution license | Creative Commons Attribution 4.0 International |
| Website | https://www.mdpi.com/1420-3049/24/11/2113/htm |
| Download | https://www.mdpi.com/1420-3049/24/11/2113/pdf (PDF) |
Abstract
Cannabis has regained much attention as a result of updated legislation authorizing many different uses, and it can be classified on the basis of the content of Δ9-tetrahydrocannabinol (Δ9-THC), a psychotropic substance for which there are legal limitations in many countries. For this purpose, accurate qualitative and quantitative determination is essential. The relationship between THC and cannabidiol (CBD) is also significant, as the latter substance is endowed with many specific and non-psychoactive proprieties. For these reasons, it becomes increasingly important and urgent to utilize fast, easy, validated, and harmonized procedures for determination of cannabinoids. The procedure described herein allows rapid determination of 10 cannabinoids from the inflorescences of Cannabis sativa L. by extraction with organic solvents. Separation and subsequent detection are by reversed-phase high-performance liquid chromatography with ultraviolet detector (RP-HPLC-UV). Quantification is performed by an external standard method through the construction of calibration curves using pure standard chromatographic reference compounds. The main cannabinoids dosed (g/100 g) in actual samples were cannabidiolic acid (CBDA), CBD, and Δ9-THC (Sample L11 CBDA 0.88 ± 0.04, CBD 0.48 ± 0.02, Δ9-THC 0.06 ± 0.00; Sample L5 CBDA 0.93 ± 0.06, CBD 0.45 ± 0.03, Δ9-THC 0.06 ± 0.00). The present validated RP-HPLC-UV method allows determination of the main cannabinoids in Cannabis sativa L. inflorescences and appropriate legal classification of it as either hemp or a drug-type.
Keywords: cannabinoids, Cannabis sativa L., HPLC, validation
Introduction
Cannabis is classified into the family of Cannabaceae and initially encompassed three main species: Cannabis sativa, Cannabis indica, and Cannabis ruderalis.[1] Nowadays, Cannabis has only one species due to continuous crossbreeding of the three species to generate hybrids. In fact, all plants are categorized as belonging to Cannabis sativa and classified into chemotypes based on the concentration of the main cannabinoids. Depending on the ratio of |tetrahydrocannabinolic acid (THCA) to cannabidiolic acid (CBDA) (the THCA/CBDA ratio), some chemotypes have been distinguished. In particular, chemotype I or “drug plants” have a TCHA/CBDA ratio >1.0, chemotype II plants exhibit an intermediate ratio, chemotype III or “fiber plants” have a THCA/CBDA ratio <1.0, chemotype IV plants contain cannabigerolic acid (CBGA) as the main cannabinoid, and chemotype V plants contain almost no cannabinoids.[2][3][4][5]
In Italy, the interest in Cannabis sativa L. has increased recently, mainly due to December 2016 legislation (Legge 2 Dicembre 2016, n. 242).[6] As a result, requests have been made to develop cost-effective and easy-to-use quantitative and qualitative methods for analysis of cannabinoids.
The Italian regulatory framework has classified two types of Cannabis sativa L. depending on the content of Δ9-tetrahydrocannabinol (Δ9-THC). In particular, fiber-type plants of Cannabis sativa L., also called “hemp,” are characterized by a low content of Δ9-THC (<0.2% w/w). If the content of Δ9-THC is >0.6% w/w, it is considered as drug-type, also called “therapeutic” or “marijuana.”
Industrial hemp is used in several sectors, such as in the pharmaceutical, cosmetic, food, and textile industries, as well as in energy production and building. In general, fiber-type plants are less used in the pharmaceutical field, where drug-type plants are more often employed.[5] However, there is also an increased interest in hemp varieties containing non-psychoactive compounds. In fact, the European Union has approved 69 varieties of Cannabis sativa L. for commercial use.[7]
Supplementary material
The following are available online here (.zip):
File S1: Standard operating procedure (SOP) of the method presented in this article, Table S1: Calibration curves relating to the standard solution of 10 cannabinoids determined by RP-HPLC-UV method, Figure S1: Calibration curves relating to the standard solution of 10 cannabinoids determined by RP-HPLC-UV method, File S2: Preliminary tests carried out for development of the analytical procedure by RP-HPLC-UV
Acknowledgements
The authors gratefully acknowledge Enecta Srl for providing samples. The experimentation was conducted in the context of a PhD project entitled Harmonized procedures of analysis of medical, herbal, food and industrial cannabis: Development and validation of cannabinoids’ quality control methods, of extraction and preparation of derivatives from the plant raw material, according to the product destination and funded by Enecta Srl.
Author contributions
Conceptualization, T.G.T., M.M. and M.T.; Methodology, M.M.; Software, M.M. and S.S.; Validation, M.M.; Formal analysis, M.M.; Investigation, T.G.T. and M.M.; Resources, T.G.T. and M.M.; Data curation, M.M., T.G.T. and M.T.; Writing—original draft preparation, M.M., M.T. and T.G.T.; writing—review and editing, T.G.T. and S.S.; Visualization, T.G.T.; Supervision, T.G.T.; project administration, T.G.T.; funding acquisition, T.G.T.
Funding
This research received no external funding; this trial received financial support from Enecta Srl.
Conflicts of interest
The authors declare no conflict of interest.
References
- ↑ Montserrat-de la Paz, S.; Marín-Aguilar, F.; García-Giménez, M.D. et al. (2014). "Hemp (Cannabis sativa L.) seed oil: Analytical and phytochemical characterization of the unsaponifiable fraction". Journal of Agricultural and Food Chemistry 62 (5): 1105–10. doi:10.1021/jf404278q. PMID 24422510.
- ↑ Appendino, G.; Chianese, G.; Taglialatela-Scafati, O. (2011). "Cannabinoids: Occurrence and medicinal chemistry". Current Medicinal Chemistry 18 (7): 1085–99. doi:10.2174/092986711794940888. PMID 21254969.
- ↑ Andre, C.M.; Hausman, J.F.; Guerriero, G. (2016). "Cannabis sativa: The Plant of the Thousand and One Molecules". Frontiers in Plant Science 7: 19. doi:10.3389/fpls.2016.00019. PMC PMC4740396. PMID 26870049. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4740396.
- ↑ Aizpurua-Olaizola, O.; Soydaner, U;. Öztürk, E. et al. (2016). "Evolution of the Cannabinoid and Terpene Content during the Growth of Cannabis sativa Plants from Different Chemotypes". Journal of Natural Products 79 (2): 324–31. doi:10.1021/acs.jnatprod.5b00949. PMID 26836472.
- ↑ 5.0 5.1 Brighenti, V.; Pellati, F.; Steinbach, M. et al. (2017). "Development of a new extraction technique and HPLC method for the analysis of non-psychoactive cannabinoids in fibre-type Cannabis sativa L. (hemp)". Journal of Pharmaceutical and Biomedical Analysis 143: 228–36. doi:10.1016/j.jpba.2017.05.049. PMID 28609672.
- ↑ "Legge 2 Dicembre 2016, n. 242 - Disposizioni per la promozione della coltivazione e della filiera agroindustriale della canapa". Gazzetta Ufficiale della Repubblica Italiana. 30 December 2016. https://www.gazzettaufficiale.it/eli/id/2016/12/30/16G00258/sg. Retrieved 01 June 2019.
- ↑ "EU Plant variety database". European Commission. 2019. http://ec.europa.eu/food/plant/plant_propagation_material/plant_variety_catalogues_databases/search//public/index.cfm. Retrieved 01 June 2019.
Notes
This presentation is faithful to the original, with only a few minor changes to presentation. Some grammar and punctuation was cleaned up to improve readability. In some cases important information was missing from the references, and that information was added.
