Enhanced production of scopolamine and atropine alkaloids in in vitro cell suspension cultures of Atropa belladonna L. under 2-iP, 2,4-D, and ornithine

The belladonna plant, Atropa belladonna L. (Solanaceae), is the most important commercial source of tropane alkaloids such as atropine and scopolamine used for medicinal and pharmaceutical purposes. The chemical biosynthesis and extraction of tropane alkaloids using wild-collected plants has been demonstrated to be difficult, prolong and commercially unachievable. Therefore, this study investigated the combinational impact of different (1) leaf sterilization treatments [mercuric chloride (HgCl2) (0.1-0.3%), sodium hypochlorite (NaOCl) (20-40%), thiophanate-methyl (fungicide) (10-30%)] on viability, clean explants, and contamination %, (2) 2-isopentenyladenine (2-iP) (2-3 mg/L) + 2,4-dichlorophenoxyacetic acid (2,4-D) (2-6 mg/L) on callus induction % and fresh weight (FW), and (3) 2-iP + 2,4-D + 1-3 mM ornithine on dry weight (DW), atropine and scopolamine content in in vitro cell suspension cultures of A. belladonna. The 0.1% HgCl2 + 30% thiophanate-methyl + 40% NaOCl sterilization treatment was the most effective (100% disinfection, 70% viability). Maximized callus induction (100%) and FW were recorded in MS medium containing 3 mg/L 2-iP + 2 mg/L 2,4-D (4 weeks) and 2 mg/L 2-iP + 5 mg/L 2,4-D (2 months), respectively. DW and scopolamine (4.33 mg/g), and atropine (4.14 mg/g) content were higher in liquid medium with 3 mg/L 2-iP + 3 mg/L 2,4-D + 1 or 3 mM ornithine, respectively (10 weeks). In vitro production of atropine and scopolamine in cell suspension cultures of A. belladonna in shake flasks can be increased and accelerated using elicitors such as plant growth regulators (i.e., 2-iP, 2,4-D) and amino acids precursors (i.e., ornithine) involved in alkaloids biosynthetic pathway.