title: Multi-development-HPTLC method for quantitation of hyoscyamine, scopolamine and their biosynthetic precursors in selected solanaceae plants grown in natural conditions and as in vitro cultures.
abstract: BACKGROUND
  Hyoscyamine and scopolamine, anti-cholinergic agents widely used in medicine, are typically obtained from plants grown under natural conditions. Since field cultivation entails certain difficulties (changeable weather, pests, etc.), attempts have been made to develop a plant in vitro culture system as an alternative source for the production of these compounds. During experiments to locate the limiting steps in the biotechnological procedure, it is important to monitor not only the levels of the final products but also the changes in the concentration of their precursors.
  
  
  OBJECTIVE
  To develop a HPTLC method for the separation and quantitation of the main tropane alkaloids hyoscyamine and scopolamine, their respective direct precursors littorine and anisodamine, and cuscohygrine, a product of a parallel biosynthetic pathway that shares a common precursor (N-methyl-∆(1) -pyrrolium cation) with tropane alkaloids.
  
  
  METHODS
  Using alkaloid extracts from Atropa baetica hairy roots, different TLC chromatographic systems and developing procedures were investigated.
  
  
  RESULTS
  Full separation of all compounds was obtained on HPTLC Si60 F254 plates preconditioned with mobile phase vapours (chloroform:methanol:acetone:25% ammonia ratios of 75:15:10:1.8, v/v/v/v). The chromatograms were developed twice (at distances of 4.0 and 3.0 cm) in a Camag twin trough chamber and visualised with Dragendorff's reagent. Densitometric detection (λ = 190 and 520 nm) was used for quantitative analyses of the different plant samples.
  
  
  CONCLUSIONS
  This method can be recommended for quantitation of hyoscyamine, scopolamine, anisodamine, littorine and cuscohygrine in different plant material (field grown vs. in vitro cultures).
authors.affiliation: Department of Pharmacognosy, Medical University of Gdansk, al. gen. J. Hallera 107, 80-416, Gdansk, Poland.
authors.forename: Zbigniew
authors.initials: Z
authors.lastname: Jaremicz
authors.forename: Maria
authors.initials: M
authors.lastname: Luczkiewicz
authors.forename: Mariusz
authors.initials: M
authors.lastname: Kisiel
authors.forename: Rafael
authors.initials: R
authors.lastname: Zárate
authors.forename: Nabil
authors.initials: N
authors.lastname: El Jaber-Vazdekis
authors.forename: Piotr
authors.initials: P
authors.lastname: Migas
mesh_terms: D000096:Acetone / Q000031:analogs & derivatives / Q000032:analysis; D032282:Atropa / Q000737:chemistry / Q000378:metabolism; D001286:Atropine Derivatives / Q000032:analysis* / Q000378:metabolism; D002855:Chromatography, Thin Layer / Q000379:methods*; D064692:Hyoscyamine / Q000032:analysis*; D018517:Plant Roots / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D011759:Pyrrolidines / Q000032:analysis; D015203:Reproducibility of Results; D012601:Scopolamine / Q000032:analysis*; D019657:Solanaceae / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D012991:Solanaceous Alkaloids / Q000032:analysis* / Q000378:metabolism; D046509:Tissue Culture Techniques
chemical_list: D001286:Atropine Derivatives; D011759:Pyrrolidines; D012991:Solanaceous Alkaloids; C003922:anisodamine; D000096:Acetone; C063158:littorine; C098327:cuscohygrine; D012601:Scopolamine; D064692:Hyoscyamine
keywords: HPTLC; Solanaceae; anisodamine; cuscohygrine; hyoscyamine; in vitro cultures; littorine; scopolamine