Simultaneous analysis of hyoscyamine, scopolamine, 6beta-hydroxyhyoscyamine and apoatropine in Solanaceous hairy roots by reversed-phase high-performance liquid chromatography.
A new high-performance liquid chromatographic method is described for tropane alkaloid analysis in genetically transformed root cultures of Datura innoxia Mill. and Atropa belladonna L. Sample preparation, tropane alkaloid extraction with chloroform-methanol-concentrated ammonia 15:5:1 (v/v/v), was followed by solid-phase extraction on Supelclean LC-18 cartridges. Optimized conditions and careful pH control resulted in high recovery and reproducibility. Simultaneous determination of apoatropine, 6beta-hydroxyhyoscyamine, hyoscyamine and scopolamine was performed by HPLC on C18 (2) reversed-phase column. The application of Luna new-generation silica-based stationary phase resulted in excellent peak shapes using an ion-pair reagent and triethanolamine free mobile phase and allowed to exploit the full power of pH-dependent selectivity. Simplicity and improved selectivity make this method a preferred alternative of published ion-pair chromatographic methods. Validation studies proved that the global method has good repeatability and satisfactory recovery. Absolute limits of detection were 0.6, 0.6, and 0.8 ng for hyoscyamine, 6beta-hydroxyhyoscyamine, and scopolamine respectively.
Kursinszki
László
L
Department of Pharmacognosy, Semmelweis University, Ulloi út 26, H-1085 Budapest, Hungary. kurslasz@drog.sote.hu
Hank
Hajnalka
H
László
Imre
I
Szoke
Eva
E
D001285:Atropine / Q000032:analysis*; D001286:Atropine Derivatives / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D010936:Plant Extracts; D018517:Plant Roots / Q000737:chemistry*; D012044:Regression Analysis; D015203:Reproducibility of Results; D012601:Scopolamine / Q000032:analysis*; D019657:Solanaceae / Q000737:chemistry*; D012991:Solanaceous Alkaloids / Q000032:analysis*
D001286:Atropine Derivatives; D010936:Plant Extracts; D012991:Solanaceous Alkaloids; C003922:anisodamine; D001285:Atropine; D012601:Scopolamine