Secondary Metabolites of Medicinal Plants. Bharat Singh

cultures. But when the cells were pretreated with cyclosporine A before the transfer of these cells in cell culture medium, then these cells survived. The DNA degradation was also performed by cannabinoid treatment in suspension of cells. The cannabinoids can lead to necrosis of cells via dysfunction of mitochondria during suspension of cells of C. sativa (Morimoto et al. 1998; Morimoto et al. 2007). The 10-Oxo-Δ^6a(10a)-tetrahydrocannabinol, cannabiglendol, Δ⁷-isotetrahydrocannabinol, and cannabicitran have been isolated from Cannabis (Flores-Sanchez and Verpoorte 2008).

      Cannabis sativa pollen grains were investigated for the presence of flavonoid glycosides (kaempferol 3-O-sophoroside and quercetin 3-O-sophoroside) and cannabinoids (Δ⁹-tetrahydrocannabiorcol, cannabidivarin, cannabicitran, Δ⁹-tetrahydrocannabivarin, cannabicyclol, cannabidiol, cannabichromene, Δ⁹-tetrahydrocannabinol, cannabigerol, cannabinol, dihydrocannabinol, cannabielsoin, 6a,7,10a-trihydroxytetrahydrocannabinol, 9,10-epoxycannabitriol, cannabinol, cannabitriol, cannabigerovarinic acid A, cannabinodiol, 10-O-ethylcannabitriol, (−)-(9R,10R)-trans-10-O-ethylcannabitriol and 7,8-dehydro-10-O-ethylcannabitriol, cannabicyclol, cannabinerolic acid, (+)-cannabichromenic acid, cannabichromevarin, cannabigerovarin, tetrahydrocannabiorcol, cannabifuran, dehydrocannabifuran, cannabichromanone-C3, cannabichromanone-C5, cannabicyclolic acid, cannabicyclovarin, cannabigerolic acid A, (−)-cannabitetrol) from the methanolic extract (ElSohly et al. 1977, 1984; Grote and Spiteller 1978a,1978b; Turner et al. 1981; Hartsel et al. 1983; Ross and ElSohly 1995; Yamaguchi et al. 1995; ElSohly and Slade 2005; Ross et al. 2005; Hanuš et al. 2016). The cannabielsoin acid, cannabinolic acid, orientin, cannabisin A, Δ⁹-tetrahydrocannabinolic acid, cannabigerolic acid, 11-hydroxy-Δ⁹-tetrahydrocannabinolic acid A, (±)-6,7-cis/trans-epoxycannabigerolic acid, 8-keto-Δ⁹-tetrahydrocannabinolic acid A, cannabidivarinic acid, 8β,11-bis-hydroxy-Δ⁹-tetrahydrocannabinolic acid, cannabidiolic acid, anandamide, and (−)-7R-cannabicourmaronic acid have been reported from Cannabis extracts (Mackie et al. 1993; Mahlberg and Kim 2004; Ahmed et al. 2008, 2015; Galal et al. 2009; Fischedick et al. 2010a; Happyana et al. 2013; Giacoppo et al. 2014; Lewis et al. 2017).

2.19.2 Culture Conditions

      Callus cultures were established in C. sativa by using the explants of seedlings and flowers. The fast synthesis of cannabinoids was observed in light, while slow response was achieved in dark. In suspension cultures, the cannabinoids could not be synthesized due to lack of polyketide synthase activity (Raharjo et al. 2006; Wang et al. 2009). By biotransformation, the cannabidiol biotransforms to cannabielsoin under normal growth culture conditions. The cannabidiol was converted to bound cannabielsoin and Δ-9-tetrahydrocannabinol to cannabicoumaronon (Braemer and Paris 1987). The cannabielsoin synthesis biologically catalyzed by the tissue culture technique for the first time was by Hartsel et al. (1983).

      Besides the successful establishment of callus cultures of C. sativa by the various authors, the cell culture studies are facing several types of challenges, e.g. slow accumulation of secondary metabolites and improper supply of raw materials. By unifying the microbial and plant metabolic pathways for the production of isoquinoline alkaloids, these problems could be overcome (Sato and Kumagai 2013). By supplementing the culture medium with cadmium and nickel, the accumulation of isoquinoline alkaloids was enhanced by two to three times (Srivastava and Srivastava 2010). Both biotic and abiotic elicitors were used to increase the accumulation of isoquinoline alkaloids. For the enhancement of the production of isoquinoline alkaloids, the tetrahydrocannabinolic acid synthase gene signaling pathway was monitored (Flores-Sanchez et al. 2009). The production of isoquinoline alkaloids was optimized by developing the hairy root cultures. The hemp hypocotyls respond faster to the several strains of Agrobacterium rhizogenes for