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      Identification of Cannabis sativa L. (hemp) Retailers by Means of Multivariate Analysis of Cannabinoids

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          Abstract

          In this work, the concentration of nine cannabinoids, six neutral cannabinoids (THC, CBD, CBC, CBG, CBN and CBDV) and three acidic cannabinoids (THCA CBGA and CBDA), was used to identify the Italian retailers of Cannabis sativa L. (hemp), reinforcing the idea that the practice of categorizing hemp samples only using THC and CBD is inadequate. A high-performance liquid chromatography/high-resolution mass spectrometry (HPLC-MS/MS) method was developed for screening and simultaneously analyzing the nine cannabinoids in 161 hemp samples sold by four retailers located in different Italian cities. The hemp samples dataset was analyzed by univariate and multivariate analysis with the aim to identify the hemp retailers without any other information on the hemp samples like Cannabis strains, seeds, soil and cultivation characteristics, geographical origin, product storage, etc. The univariate analysis highlighted that the hemp samples could not be differentiated by using any of the nine cannabinoids analyzed. To evaluate the real efficiency of the discrimination among the four hemp retailers a partial least squares discriminant analysis (PLS-DA) was applied. The PLS-DA results showed a very good discrimination between the four hemp retailers with an explained variance of 100% and low classification errors in both calibration (5%) and cross validation (6%). A total of 92% of the hemp samples were correctly classified by the cannabinoid variables in both fitting and cross validation. This work contributed to show that an analytical method coupled with multivariate analysis can be used as a powerful tool for forensic purposes.

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          Classification tools in chemistry. Part 1: linear models. PLS-DA

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            Cannabis sativa: A comprehensive ethnopharmacological review of a medicinal plant with a long history

            Cannabis sativa L. (C. sativa) is an annual dioecious plant, which shares its origins with the inception of the first agricultural human societies in Asia. Over the course of time different parts of the plant have been utilized for therapeutic and recreational purposes, for instance, extraction of healing oils from seed, or the use of inflorescences for their psychoactive effects. The key psychoactive constituent in C. sativa is called Δ-9-tetrahydrocannabinol (D9-THC). The endocannabinoid system seems to be phylogenetically ancient, as it was present in the most primitive vertebrates with a neuronal network. N-arachidonoylethanolamine (AEA) and 2-arachidonoyl glycerol (2-AG) are the main endocannabinoids ligands present in the animal kingdom, and the main endocannabinoid receptors are cannabinoid type-1 (CB1) receptor and cannabinoid type-2 (CB2) receptor.
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              Identification of olivetolic acid cyclase from Cannabis sativa reveals a unique catalytic route to plant polyketides.

              Δ(9)-Tetrahydrocannabinol (THC) and other cannabinoids are responsible for the psychoactive and medicinal properties of Cannabis sativa L. (marijuana). The first intermediate in the cannabinoid biosynthetic pathway is proposed to be olivetolic acid (OA), an alkylresorcinolic acid that forms the polyketide nucleus of the cannabinoids. OA has been postulated to be synthesized by a type III polyketide synthase (PKS) enzyme, but so far type III PKSs from cannabis have been shown to produce catalytic byproducts instead of OA. We analyzed the transcriptome of glandular trichomes from female cannabis flowers, which are the primary site of cannabinoid biosynthesis, and searched for polyketide cyclase-like enzymes that could assist in OA cyclization. Here, we show that a type III PKS (tetraketide synthase) from cannabis trichomes requires the presence of a polyketide cyclase enzyme, olivetolic acid cyclase (OAC), which catalyzes a C2-C7 intramolecular aldol condensation with carboxylate retention to form OA. OAC is a dimeric α+β barrel (DABB) protein that is structurally similar to polyketide cyclases from Streptomyces species. OAC transcript is present at high levels in glandular trichomes, an expression profile that parallels other cannabinoid pathway enzymes. Our identification of OAC both clarifies the cannabinoid pathway and demonstrates unexpected evolutionary parallels between polyketide biosynthesis in plants and bacteria. In addition, the widespread occurrence of DABB proteins in plants suggests that polyketide cyclases may play an overlooked role in generating plant chemical diversity.
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                Author and article information

                Contributors
                Role: Academic Editor
                Journal
                Molecules
                Molecules
                molecules
                Molecules
                MDPI
                1420-3049
                07 October 2019
                October 2019
                : 24
                : 19
                : 3602
                Affiliations
                Faculty of Bioscience and Technology for Food, Agriculture and Environment, University of Teramo, 64100 Teramo, Italy; spalmieri@ 123456unite.it (S.P.); aricci@ 123456unite.it (A.R.); ffanti@ 123456unite.it (F.F.); cottaviani@ 123456unite.it (C.O.); closterzo@ 123456unite.it (C.L.S.)
                Author notes
                [* ]Correspondence: mmascini@ 123456unite.it (M.M.); msergi@ 123456unite.it (M.S.); Tel.: +39-861-266949 (M.S.)
                Author information
                https://orcid.org/0000-0003-2508-5680
                https://orcid.org/0000-0002-7604-1584
                Article
                molecules-24-03602
                10.3390/molecules24193602
                6804059
                31591294
                1582cb17-601e-4ebb-80e3-735b1fcf1156
                © 2019 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 31 August 2019
                : 04 October 2019
                Categories
                Article

                cannabis sativa l.,hplc-ms/ms analysis,cannabinoids,multivariate analysis,partial least squares discriminant analysis (pls-da)

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