Expression of a heterologous S-adenosylmethionine decarboxylase cDNA in plants demonstrates that changes in S-adenosyl-L-methionine decarboxylase activity determine levels of the higher polyamines spermidine and spermine.
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We posed the question of whether steady-state levels of the higher polyamines spermidine and spermine in plants can be influenced by overexpression of a heterologous cDNA involved in the later steps of the pathway, in the absence of any further manipulation of the two synthases that are also involved in their biosynthesis. Transgenic rice (Oryza sativa) plants engineered with the heterologous Datura stramonium S-adenosylmethionine decarboxylase (samdc) cDNA exhibited accumulation of the transgene steady-state mRNA. Transgene expression did not affect expression of the orthologous samdc gene. Significant increases in SAMDC activity translated to a direct increase in the level of spermidine, but not spermine, in leaves. Seeds recovered from a number of plants exhibited significant increases in spermidine and spermine levels. We demonstrate that overexpression of the D. stramonium samdc cDNA in transgenic rice is sufficient for accumulation of spermidine in leaves and spermidine and spermine in seeds. These findings suggest that increases in enzyme activity in one of the two components of the later parts of the pathway leading to the higher polyamines is sufficient to alter their levels mostly in seeds and, to some extent, in vegetative tissue such as leaves. Implications of our results on the design of rational approaches for the modulation of the polyamine pathway in plants are discussed in the general framework of metabolic pathway engineering. (+info)
Amino acid sequences of ferredoxins from Scopolia japonica and Lycium chinense: their similarities to that of Datura arborea.
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The complete amino acid sequences of [2Fe-2S] ferredoxins from Scopolia japonica and Lycium chinense have been determined by automated Edman degradation of the entire Cm-proteins and of the peptides obtained by enzymatic digestions. These two ferredoxins exhibited only 2-7 differences in the amino acid sequence when compared to the Datura-ferredoxins (D. stramonium, D. metel, and D. arborea), and especially only 2 or 3 differences compared to D. arborea. On the contrary, 8-19 differences were observed among the other solanaceous ferredoxins. This suggests that S. japonica and L. chinense are closely related taxonomically to Datura plants, especially to D. arborea. (+info)
Effect of pmt gene overexpression on tropane alkaloid production in transformed root cultures of Datura metel and Hyoscyamus muticus.
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In order to increase the production of the pharmaceuticals hyoscyamine and scopolamine in hairy root cultures, a binary vector system was developed to introduce the T-DNA of the Ri plasmid together with the tobacco pmt gene under the control of CaMV 35S promoter, into the genome of Datura metel and Hyoscyamus muticus. This gene codes for putrescine:SAM N-methyltransferase (PMT; EC. 2.1.1.53), which catalyses the first committed step in the tropane alkaloid pathway. Hairy root cultures overexpressing the pmt gene aged faster and accumulated higher amounts of tropane alkaloids than control hairy roots. Both hyoscyamine and scopolamine production were improved in hairy root cultures of D. metel, whereas in H. muticus only hyoscyamine contents were increased by pmt gene overexpression. These roots have a high capacity to synthesize hyoscyamine, but their ability to convert it into scopolamine is very limited. The results indicate that the same biosynthetic pathway in two related plant species can be differently regulated, and overexpression of a given gene does not necessarily lead to a similar accumulation pattern of secondary metabolites. (+info)
During October 11-November 20, 2002, the Cincinnati Drug and Poison Information Center (DPIC) received notification of and offered treatment advice for 14 adolescents in the Akron/Cleveland, Ohio, area who became ill after intentional exposure to toxic seeds that DPIC identified as Datura inoxia. All became ill shortly after eating the seeds or drinking tea brewed using the seeds. All patients recovered fully after treatment. This report summarizes these cases, discusses the characteristics of the various plants known commonly as "moonflowers," and underscores the need for awareness of the potential toxicity from recreational use of a plant. (+info)
Comparison of the nucleotide sequences of ferredoxin-cDNAs among some Datura plants.
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The nucleotide sequences of partial ferredoxin (Fd)-cDNAs (corresponding to the amino acid sequence of 22-87 in the total 97 amino acids of ferredoxin) were determined for Datura arborea, D. stramonium, D. metel, and related Datura plants. Non-synonymous substitutions were noted at 4 positions and a synonymous substitution was seen at position 82 (Gln [CAA] (arborea) vs. Gln [CAG] (stramonium and metel)). The nucleotide sequence of Fd-cDNA may provide more detailed information regarding the relative taxonomic positions of plants than the amino acid sequence. However, Datura plants in the same section (metel, fastuosa, and innoxia) and of different varieties (stramonium var. stramonium and stramonium var. tatula) showed identical Fd-cDNA nucleotide sequences. This result suggests that there are very close relationships among the plants in each group. (+info)
Mydriasis due to Datura inoxia.
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Unilateral, dilated unresponsive pupil may be secondary to a wide range of ocular and neurological disorders. "Gardener's pupil" is a pharmacological mydriasis caused by exposure to plants containing alkaloids. We report a case of mydriasis related to Datura inoxia, and detail a number of plants that have toxic effects. This report emphasises the importance of accurate history taking when evaluating fixed dilated pupil. We recommend that labelling of such poisonous plants should detail the local and systemic effects of accidental exposure, rather than merely the fact that they are poisonous. (+info)
In vitro antifungal activity of 2-(3,4-dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1-methylethyl pentanoate, a dihydropyrrole derivative.
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A novel compound 2-(3,4-dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1-methylethyl pentanoate was isolated from the plant Datura metel L. The in vitro activity of this dihydropyrrole derivative against Aspergillus and Candida species was evaluated by using standard methods approved by the National Committee for Clinical Laboratory Standards. The compound was found to be active against all the species tested, namely Candida albicans, Candida tropicalis, Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger. The MIC at which more than 90% of growth was inhibited (MIC(90)) by the compound ranged from 21.87 to 43.75 microg ml(-1) against various fungal species by microbroth dilution assay. Since the compound 2-(3,4-dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1-methylethyl pentanoate has antifungal activity it can be explored further to develop new antimycotic drugs. (+info)
Testing for atropine and scopolamine in hair by LC-MS-MS after Datura inoxia abuse.
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Datura inoxia belongs to the family of Solanaceae. This is a very common plant in New Caledonia that contains two main toxic alkaloids, l-atropine and l-scopolamine. In this study, we report the case of a 20-year-old male admitted to an Emergency Unit after consumption of 6 dried flowers in hot water for hallucinations, mydriasis, and agitation associated with tachycardia and increase of systolic blood pressure to 180. Full recovery was observed after one week. Three weeks later, a lock of about 80 hairs (200 mg) was collected from the subject in vertex posterior with scissors to be tested for both atropine and scopolamine. After decontamination with dichloromethane, a strand of hair was segmented into three parts, cut into small segments (< 1 mm), incubated overnight in 1 mL pH 8.4 phosphate buffer in the presence of 2.5 ng atropine-d(3), the internal standard, then extracted with 5 mL dichloromethane/isopropanol/n-heptane (50:17:33). The residue was reconstituted in 100 microL of methanol, from which 10 microL was injected into an XTerra MS C18 column (100 x 2.1 mm, 3.5 microm) eluted with a gradient of acetonitrile and formate buffer delivered at a flow rate of 0.2 mL/min. A Quattro Micro triple-quadrupole mass spectrometer (MS) was used for analyses. Ionization was achieved using electrospray in the positive ionization mode. For each compound, detection was related to two daughter ions (atropine: m/z 290.2 to 124.0 and 92.9; atropine-d(3): m/z 293.1 to 127.0 and 92.9; scopolamine: m/z 304.1 to 138.0 and 156.0). Although atropine was never detected (limit of detection = 2 pg/mg), scopolamine was identified in the three segments, in the range 14 to 48 pg/mg. The absence of atropine in hair is consistent with its very low dosage in the flower of Datura inoxia. Hair segmentation indicated that the subject was previously exposed on several occasions to the plant. Liquid chromatography-tandem MS appears to be a necessity for testing tropane alkaloids of the Datura group, given the low concentrations to be measured. (+info)