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Calystegines in root cultures of Atropa belladonna respond to sucrose, not to elicitation.

Rothe G, Garske U, Drager B.

Institute of Pharmaceutical Biology, Martin-Luther-University Halle-Wittenberg, Hoher Weg 8, D-06120, Saale, Halle, Germany

Calystegines are norpseudotropine alkaloids accumulating in root cultures of Atropa belladonna, together with tropine derivatives, e.g. hyoscyamine. Both alkaloid groups are derived from the tropane alkaloid pathway. For the investigation of the regulation and individual steps of tropane biosynthesis, methods for the induction of the pathway were tested. Elicitation by chitosan, or defence responses to ABA and methyl jasmonate did not enhance calystegine accumulation, but led to a more or less pronounced decrease. By blocking one arm of the diverged tropane pathway, calystegine accumulation can be increased, but total tropane alkaloid formation does not increase considerably. By elevation of sucrose supply, both, total alkaloids and calystegines in particular were increased approximately threefold. The mechanism of the induction of the biosynthesis by sucrose is not known and needs further experiments.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11297802&dopt=Abstract hyoscyamine Levbid SL



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Decreased scopolamine yield in field-grown Duboisia plants regenerated from hairy roots.

Roig Celma C, Palazon J, Cusido RM, Pinol MT, Keil M.

Departament de Productes Naturals, Biologia Vegetal i Edafologia, Facultat de Farmacia, Universitat de Barcelona, Barcelona, Spain.

Hairy root cultures were obtained from hybrid clones of Duboisia myoporoides x D. leichhardtii following transformation by Agrobacterium rhizogenes strain A4. Shoots spontaneously regenerating from the hairy root cultures were rooted and transferred to soil. The plants displayed typical morphological alterations known as hairy root syndrome to varying degrees. PCR analysis confirmed that all transformed plants contained the rolA, rolB and rolC genes, irrespective of the degree of morphological alterations. A field test of the transformed regenerated plants revealed that those plants displaying the strongest hairy root syndrome symptoms had the highest content of the tropane alkaloid scopolamine. However, the overall scopolamine and hyoscyamine yield of all transformed plants was clearly reduced compared to untransformed control plants. These results demonstrate that the A. rhizogenes-transformed plants tested in this study do not provide a viable alternative to agricultural farming of hybrid clones of D. myoporoides x D. leichhardtii obtained by conventional breeding.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11345697&dopt=Abstract hyoscyamine Levbid SL



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Occurrence of cadaverine in hairy roots of Brugmansia candida.

Carrizo CN, Pitta-Alvarez SI, Kogan MJ, Giulietti AM, Tomaro ML.

Catedra de Microbiologia Industrial y Biotecnologia, Facultad de Farmacia y Bioquimica, Universidad de Buenos Aires, Junin 956, (1113) Buenos Aires, Argentina.

The polyamine, cadaverine, was detected in transformed root cultures of Brugmansia candida (syn. Datura candida), a Solanaceae which produces the tropane alkaloids scopolamine and hyoscyamine. To the best of our knowledge, this is the first time that the existence of this uncommon polyamine has been detected in a Datura species. Cadaverine, however, could not be found in the whole plant. The occurrence of cadaverine in hairy roots could be a consequence of either the transformation or a response to stress. Also, cadaverine could be participating in other secondary pathways rather than to the tropane alkaloids. The common polyamines, putrescine, spermidine and spermine were also observed.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11397445&dopt=Abstract hyoscyamine Levbid SL



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Hairy roots of Brugmansia candida that grow without agitation: biotechnological implications.

Pitta-Alvarez SI, Giulietti AM.

Catedra de Microbiologia Industrial y Biotecnologia, Facultad de Farmacia y Bioquimica, Universidad de Buenos Aires, Junin 956, 6to. piso, Buenos Aires 1113, Argentina.

Hairy roots of Brugmansia candida that grew without agitation were obtained. Kinetics of growth and production of the tropane alkaloids scopolamine and hyoscyamine, with and without agitation, were studied. The exponential growth rate was higher in the roots that were exposed to shaking (0.13 d(-1)) than in the nonagitated ones (0.09 d(-1)). The specific production and the levels per flask of both alkaloids were enhanced without shaking. The use of these roots in large-scale productions could be economically advantageous. It remains to be seen if the data obtained in shake flasks can be extrapolated to large-scale bioreactors.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11485426&dopt=Abstract hyoscyamine Levbid SL



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Interactions between allosteric modulators and 4-DAMP and other antagonists at muscarinic receptors: potential significance of the distance between the N and carboxyl C atoms in the molecules of antagonists.

Lysikova M, Havlas Z, Tucek S.

Institute of Physiology, Academy of Sciences of the Czech Republic, Prague.

Allosteric enhancement of the affinity of muscarinic receptors for their ligands offers a new way to influence cholinergic neurotransmission. The structure of the allosteric binding domain(s) and the features of agonists, antagonists and modulators which determine the occurrence of either positive or negative cooperativity require clarification. We tested interactions between allosteric modulators alcuronium, strychnine and brucine and eight antagonists at muscarinic receptors expressed in CHO cells. In experiments with unlabeled antagonists, all three modulators enhanced the affinity for 4-diphenylacetoxy-N-dimethylpiperidinium (4-DAMP) at the M2 receptors, and strychnine did so also at the M4 receptors. Positive interactions were also observed between alcuronium and L-hyoscyamine (M2) and scopolamine (M2), between strychnine and butylscopolamine (M4), L-hyoscyamine (M2 and M4) and scopolamine (M4), and between brucine and scopolamine (M2). Positive effects of alcuronium, strychnine and brucine on the affinity of the M2 receptors for 4-DAMP have been confirmed by direct measurements of the binding of [3H]-4-DAMP. A comparison of molecular models of several antagonists which are esters revealed that antagonists in which the distance between the N and the carboxyl C atoms corresponds to five chemical bonds are more likely to display positive cooperativity with alcuronium at the M2 receptors than the antagonists in which the N-carboxyl C distance corresponds to four chemical bonds.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11495349&dopt=Abstract hyoscyamine Levbid SL