Enantiomeric differentiation of atropine/hyoscyamine by (13) C NMR spectroscopy and its application to Datura stramonium extract.
BACKGROUND
The two enantiomers of hyoscyamine, an alkaloid found in many plant species, have distinct pharmacological and biological properties. Methods for the discrimination of both enantiomers are almost exclusively based on chiral HPLC/UV. Determination of the enantiomeric ratio (e.r.) of hyoscyamine is a challenging problem since this compound tends to racaemise, forming atropine during acid-base extraction.


OBJECTIVE
To develop a protocol for the calculation of enantiomeric ratio of hyoscyamine in a plant extract using a (13) C NMR method.


METHODS
Samples were prepared by extraction of dried Datura stramonium seeds. Observation of C12 and C15 NMR signals of hyoscyamine in the presence of one equivalent of TFA and sub-stoichiometric amount of Yb(hfc)(3) allowed the calculation of the e.r. of S-(-) and R-(+)-hyoscyamine.The method was optimised with various mixtures of (+) and (-)-hyoscyamine ranging from 50:50 (racaemic mixture, i.e. atropine) to 98.5:1.5. The e.r. measured by NMR on the signals of aromatic C12 and C15 were in agreement with the gravimetrically prepared samples. The method was then applied to an extract of Datura stramonium and S-(-)-hyoscyamine was the unique enantiomer.


CONCLUSIONS
The study showed that the e.r. determination of atropine/hyoscyamine was achieved with a routine NMR spectrometer, using CLSR/TFA on pure compounds as well as on the crude extract of Datura stramonium.
Antoine Lanfranchi
Don
D
Université de Corse-CNRS, UMR 6134 SPE, Equipe Chimie et Biomasse, Route des Sanguinaires, 20000 Ajaccio, France.
Tomi
Félix
F
Casanova
Joseph
J
D000588:Amines / Q000737:chemistry; D001285:Atropine / Q000737:chemistry*; D013286:Datura stramonium / Q000737:chemistry*; D028581:Lanthanoid Series Elements / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D018727:Muscarinic Antagonists / Q000737:chemistry*; D010936:Plant Extracts / Q000032:analysis; D011522:Protons; D012639:Seeds / Q000737:chemistry; D013237:Stereoisomerism
D000588:Amines; D028581:Lanthanoid Series Elements; D018727:Muscarinic Antagonists; D010936:Plant Extracts; D011522:Protons; D001285:Atropine