In vitro propagation methods of the succulent plant Euphorbia resinifera and quantification of its major compounds

Being native from Morocco and found mostly on the slopes of the Atlas, Euphorbia resinifera, a succulent Euphorbiaceae, was used in antiquity as a medicinal plant by applying its dried latex (Euphorbium) against snake bites and during poisoning. More recently, this plant was shown to contain the strongest “spice” found in nature so far, the diterpenoid resiniferatoxin (RTX) [1]. This complex molecule is 1000 more potent on the Scoville scale than pure capsaicin and RTX, which strongly activate TRPV1 receptors in neurons, is now being tested in Phase II clinical trials to manage pain in terminally ill cancer patients [2]. The NIH is also trying to develop new analgesics based on RTX. Additionally, some triterpenoids present in the latex have been shown to be potent cytotoxics [3 – 4], therefore highlighting the pharmaceutical interest in this plant species. The endemic location of Euphorbia resinifera and its slow growth are a bottleneck for the large-scale production of active ingredients based on this plant species. Here we described for the first time the in-vitro propagation methods for the production of undifferentiated (callus and cell suspension cultures) as well as the micropropagation of Euphorbia resinifera. Moreover, quantification of major metabolites Euphol, Euphorbol and RTX, in the ex vivo and in vitro plants as well as in the cell suspensions was performed by HPLC-DAD and the identity of the compounds was confirmed by LC-MS. The impact of plant tissue differentiation is discussed in relation to the production of complex diterpenoids and triterpenoids in Eurphorbia resinifera plants and latex-free cell suspensions.