ORIGINAL PAPER Effects of sucrose and plant growth regulators on acetylcholinesterase inhibitory activity of alkaloids accumulated in shoot cultures of Amaryllidaceae Anna El Tahchy • Simon Bordage • Agata Ptak • Franc ¸ois Dupire • Elvina Barre • Catherine Guillou • Max Henry • Yves Chapleur • Dominique Laurain-Mattar Received: 10 November 2010 / Accepted: 29 January 2011 / Published online: 16 February 2011 Ó Springer Science+Business Media B.V. 2011 Abstract The influence of sucrose (30, 60, 90 and 120 g/L), activated charcoal (5 and 10 g/L), and various levels of several plant growth regulators (6-benzyladenine, naph- thalene-1-acetic acid, 2,4-dichlorophenoxyacetic acid, and picloram) on organogenesis (bulb and root development) and the accumulation of alkaloid and galanthamine in shoot cultures of three Amaryllidaceae species (Narcissus pseudonarcissus, Galanthus elwesii, and Leucojum aes- tivum) was investigated in a full-factorial experiment. Alkaloid extracts were analyzed by gas chromatography– mass spectrometry, leading to the quantification of galan- thamine and to the identification of other alkaloids. The different extracts were then subjected to an Ellman test to evaluate the inhibitory activity of acetylcholinesterase. The highest contents of galanthamine [0.02–0.1% dry weight (DW) depending on the plant species] were always accompanied with a high level of acetylcholinesterase inhibition ( [ 30%). However, some samples containing low amounts of galanthamine (0.005% DW) showed high inhibitory activities (40–80%). These findings demonstrate the presence of Amaryllidaceae alkaloids that have not yet been identified as having anti-acetylcholinesterase activity. Keywords Narcissus pseudonarcissus Á Galanthus elwesii Á Leucojum aestivum Á Amaryllidaceae alkaloids Á Acetylcholinesterase inhibition Á Nutrient medium variation Abbreviations AChE Acetylcholinesterase BA 6-benzyladenine 2,4-D 2,4-dichlorophenoxyacetic acid DW Dry weight Gal Galanthamine NAA Naphthalene-1-acetic acid PGRs Plant growth regulators Picloram 4-Amino-3,5,6-trichloropicolinic acid Rt Retention time Introduction Cholinesterase inhibitors are approved drugs for the palli- ative treatment of Alzheimer’s disease, a progressive neurodegenerative disorder that is one of the most common causes of mental deterioration in the elderly population (Henchman et al. 2002). Plants of the Amaryllidaceae family produce pharmacologically active alkaloids that have interesting pharmacological properties, such as ace- tylcholinesterase (AChE) inhibitory activity, cytotoxicity, antitumoral activity, among others (Bastida et al. 2006). Galanthamine (Gal) (Fig. 1a) and lycorine (Fig. 1b) are two of the best known alkaloids that have been found in plants of different Amaryllidaceae genera, including A. El Tahchy Á S. Bordage Á M. Henry Á Y. Chapleur Á D. Laurain-Mattar (&) Groupe S.U.C.R.E.S., UMR 7565 CNRS-Nancy-Universite ´, BP 70239, 54506 Nancy-Vandoeuvre, France e-mail: dominique.laurain-mattar@pharma.uhp-nancy.fr A. Ptak Department of Plant Breeding and Seed Science, Agricultural University, 31-140 Krakow, Poland F. Dupire Service Commun de Spectrome ´trie de Masse, UHP-Institut Jean Barriol, 54506 Nancy-Vandoeuvre, France E. Barre Á C. Guillou Centre de Recherche de Gif, Institut de Chimie des Substances Naturelles-CNRS, Bt 27, Avenue de la Terrasse, 91198 Gif-sur-Yvette, France 123 Plant Cell Tiss Organ Cult (2011) 106:381–390 DOI 10.1007/s11240-011-9933-7