Adatok az Atropa belladonna L. gentranszformalt kulturainak jellemzesehez.

摘要

Atropa belladonna L. (deadly nightshade) contains several agents with pharmacological importance; it is an official drug in the VIII th Hungarian Pharmacopoeia. The plant contains tropane- and pyrrolidine-base alkaloids; its main alkaloid is the hyoscyamine which has parasympatholytic effect. Nowadays gene technological methods are widely used to increase the production of active ingredient beside the conventional tissue culturing procedures.;During our experiments we investigated the growth of the genetically modified and non-transformed cultures of in vitro and in vivo A. belladonna, and its tropane alkaloid composition, specifically its hyoscyamine, scopolamine and 6OH-hyoscyamine content (LC-MS-MS, HPLC). Genetical transformations mediated by Agrobacterium rhizogenes (R1601) in cultures was demonstrated by using polymerase chain reaction (PCR) and opine detection. However several methods can be found in the bibliography for determination of alkaloid content using HPLC assessment, we elaborated a new, simple validated method that is suitable for the analysis of numerous and complex samples and that make possible a quick, accurate, qualitative and quantitative determination of alkaloid content [1.,2.].;Alkaloid contents (0.52%, 0.40%, 0.35%) of the investigated hairy root clones (;Both three hairy root clones spontaneously organized using liquid B5 culture medium. The in vitro transgenic plants were cultivated in vivo after micropropragation. Phenotypes of ;In order to know more about the metabolism of tropane-base alkaloids, and to understand more accurately the changes during abiotic stress effects, the measurable endogeous HCHO-content of the samples has been determined (OPLC, HPLC) [4.]. Our biological experiments that have been performed in BioArena-system demonstrated that the investigated components of A. belladonna most likely affects through HCHO.;1Hank H., Szo&huml;ke E., Toth K., Laszlo I., Kursinszki L. (2004): Investigation of Tropane Alkaloids in Genetically Transformed Atropa belladonna L. Cultures. Chromatographia 60. 55-59. 2Kursinszki L., Hank H., Laszlo I., Szo&huml;ke E. (2005): Simultaneous analysis of hyoscyamine, scopolamine, 6beta-hydroxyhyoscyamine and apoatropine in Solanaceous hairy roots by reversed phase high-performance liquid chromatography. Journal of chromatography A 1091, 32-39. 3Hank H., Laszlo I., Balvanyos I., Kursinszki L., Kovacs Gy., Toth K., Szo&huml;ke E. (2003): Effect of Magnesium on the Growth and Alkaloid Production of Hairy Root Cultures. Acta Horticult.597, 271-274. 4Hank, H., Tyihak, E., Katay, Gy., Kursinszki, L., Szo&huml;ke, E. (2005): Tropane alkaloids and endogenous formaldehyde levels in vivo and in vitro in genetically modified Atropa belladonna L. European Journal of Pharmaceutical Sciences 25S1. 110-111.