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quantification of metabolites in the indole alkaloid pathways of catharanthus roseus: implications for metabolic engineeringin this article, we present a review of the current state of metabolic engineering in catharanthus roseus. a significant amount of research has contributed to characterization of several individual steps in the biosynthetic pathway of medicinally valuable alkaloids. however, knowledge of the regulation of these pathways is still sparse. using hairy root cultures, we studied the responses of alkaloid metabolism to environmental stimulation such as light and elicitation. through precursor feeding ...199810191413
transient studies of light-adapted cultures of hairy roots of catharanthus roseus: growth and indole alkaloid accumulation.cultures of c. roseus transgenic ("hairy") root clones lbe-6-1 and lbe-4-2 were adapted with periodic daily illumination to investigate the effect of light on growth and nutrient utilization, and the accumulation of the indole alkaloids. light-adapted roots appeared green and had radially thickened morphology compared with dark-grown controls. their growth rates were higher than dark-grown controls, with 45% lower doubling times: lbe-6-1, 3.6 days; lbe-4-2, 2.8 days. relative to dark-grown contr ...199810099477
determination of inorganic phosphate by coupling thymidine phosphorylase and reversed-phase high-performance liquid chromatography: application to tonoplast pyrophosphatase activity.we developed an hplc method for the measurement of inorganic phosphate using thymidine phosphorylase (ec 2.4.2.4). this enzyme catalyzes the phosphorolysis of thymidine to 2-deoxyribose 1-phosphate and thymine. thymine release was measured at 265 nm after separation by reverse-phase hplc. the assay was sensitive enough to detect as little as 10 pmol of pi. the response to the phosphate concentration was linear from 1 to 100 microm. the value of this method was demonstrated in an analysis of the ...19979417785
terpenoid biosynthesis from 1-deoxy-d-xylulose in higher plants by intramolecular skeletal rearrangement.the incorporation of [1-13c]- and [2,3,4,5-13c4]1-deoxy-d-xylulose into beta-carotene, lutein, phytol, and sitosterol in a cell culture of catharanthus roseus was analyzed by nmr spectroscopy. the labeling patterns of the isoprene precursors, isopentenyl pyrophosphate and dimethylallyl pyrophosphate, were obtained from the terpenes by a retrobiosynthetic approach. 13c enrichment and 13c13c coupling patterns showed conclusively that 1-deoxy-d-xylulose and not mevalonate is the predominant isopren ...19979380681
transient studies of nutrient uptake, growth, and indole alkaloid accumulation in heterotrophic cultures of hairy roots of catharanthus roseus.the kinetics of growth, the uptake of macronutrients, and the accumulation of indole alkaloids were investigated in long-term, heterotrophically cultured transgenic ("hairy") roots of catharanthus roseus.tabersonine, ajmalicine, and serpentine were monitored over a 70-day period. the doubling time [dry-weight (dw) basis] of c. roseus hairy roots in b5/2 nutrients supplemented with 3% sucrose was 3.6 days. nh(4)(+), no(3),(-) and p(i) were depleted sequentially from culture medium by hairy roots, ...199718636519
studies on biosynthesis of brassinosteroids.biosynthesis of steroidal plant hormones, brassinosteroids, was studied using the cell culture system of catharanthus roseus. feeding labeled compounds of possible intermediates to the cultured cells, followed by analyzing the metabolites by gas chromatography-mass spectrometry disclosed the pathways from a plant sterol, campesterol, to brassinolide. there are two pathways, named the early c6-oxidation pathway and late c6-oxidation pathway, both of which would be operating in a wide variety of p ...19979178548
[characterization of a protein related to bet v 1, the major birch pollen antigen, from cells of the madagascar periwinkle. universal presence of proteins of this type among higher plants].in this work we present the characterisation of a gene that codes for a protein of 17 kda, in in vitro cultures of a plant with ornamental and pharmaceutical properties, the madagascan periwinkle, (catharanthus roseus [l] g. don). this protein is very close to the principal allergen of birch and also to allergens isolated from or demonstrated in some foods such as celery, parsley, apple tree, peas, asparagus and potato, but it has no allergenic characteristics.19979453736
three differentially expressed s-adenosylmethionine synthetases from catharanthus roseus: molecular and functional characterization.we describe the molecular and functional characterization of three closely related s-adenosyl-l-methionine synthetase (sams) isoenzymes from catharanthus roseus (madagascar periwinkle). the genes are differentially expressed in cell cultures during growth of the culture and after application of various stresses (elicitor, nutritional down-shift, increased nacl). seedlings revealed organ-specific expression and differential gene regulation after salt stress. a relationship analysis indicated that ...19979037140
comparison of the activities of camv 35s and fmv 34s promoter derivatives in catharanthus roseus cells transiently and stably transformed by particle bombardment.activities of several camv 35s and fmv 34s promoter derivatives fused to the gusa reporter gene were compared in suspension-cultured catharanthus roseus cells that were transiently and stably transformed using particle bombardment. our data demonstrate that the 35s and a deletion derivative of the 34s promoter combined with particle bombardment form useful tools for genetic engineering of c. roseus cells. our results disagree on several points with activities of 35s and 34s promoter derivatives ...19979106518
cell-cycle-regulated transcription of a- and b-type plant cyclin genes in synchronous cultures.synchronously dividing cell cultures of catharanthus roseus were used to isolate cdnas for two mitotic cyclins, named cys and cym. the deduced protein sequence of cys is similar to that of a-type cyclins, and cym belongs to the group of b-type cyclins. in a fashion similar to the pattern of expression seen for a-type and b-type cyclins in mammalian cells, cys is expressed before cym in c. roseus cells during the cell cycle. cys mrna accumulated at the onset of s phase and disappeared early in th ...19979193070
an important developmental role for oligosaccharides during early embryogenesis of cyprinid fish.derivatives of chitin oligosaccharides have been shown to play a role in plant organogenesis at nanomolar concentrations. here we present data which indicate that chitin oligosaccharides are important for embryogenesis in vertebrates. we characterize chitin oligosaccharides synthesized in vitro by zebrafish and carp embryos in the late gastrulation stage by incorporation of radiolabeled n-acetyl-d-[u14c]glucosamine and by hplc in combination with enzymatic conversion using the bradyrhizobium nod ...19979223299
molecular cloning and characterization of desacetoxyvindoline-4-hydroxylase, a 2-oxoglutarate dependent-dioxygenase involved in the biosynthesis of vindoline in catharanthus roseus (l.) g. don.a 2-oxoglutarate-dependent dioxygenase (ec 1.14.11.11) which catalyzes the 4-hydroxylation of desacetoxyvindoline was purified to homogeneity. three oligopeptides isolated from a tryptic digest of the purified protein were microsequenced and one oligopeptide showed significant homology to hyoscyamine 6 beta-hydroxylase from hyoscyamus niger. a 36-mer degenerate oligonucleotide based on this peptide sequence was used to screen a catharanthus roseus cdna library and three clones, cd4h-1 to -3, wer ...19979290645
theophylline metabolism in higher plants.metabolism of [8-(14)c]theophylline was investigated in leaf segments from camellia sinensis (tea), camellia irrawadiensis, ilex paraguariensis (maté) and avena sativa, root segments of vigna mungo seedlings and cell suspension cultures of catharanthus roseus. there was extensive uptake and metabolism of [8-(14)c]theophylline by leaves of tea and camellia irrawadiensis and, to a lesser extent, maté. these purine alkaloid-containing species converted [8-(14)c]theophylline into 3-methylxanthine, x ...19979305805
a promoter region that controls basal and elicitor-inducible expression levels of the nadph:cytochrome p450 reductase gene (cpr) from catharanthus roseus binds nuclear factor gt-1.nadph:cytochrome p450 reductase (cpr) is essential for the activation of cytochrome p450 enzymes, which are involved in a wide variety of metabolic pathways in plants, including those related to defence responses. in the subtropical plant catharanthus roseus several cytochrome p450 enzymes operate in the biosynthesis of defence-related terpenoid indole alkaloids (tias). in agreement with the importance of cpr in defence, cpr mrna levels in c. roseus were found to be enhanced by fungal elicitor p ...19979435792
in vitro evaluation of medicinal plant extracts against pestalotiopsis mangiferae.a serious leaf-spot disease of mangifera indica was noted during the last 10 years in satpura plateau of india. on the basis of characteristic symptoms and cultural characters, the pathogen was identified as pestalotiopsis mangiferae which is hitherto not reported from satpura plateau of india. screening of 17-medicinal plants against the test pathogen revealed 14 antimycotic whereas 3-plants, viz., argemone mexicana, caesalpinia bonducella, and casia fistula acclerated the growth of the pathoge ...19969676046
novel type of receptor-like protein kinase from a higher plant (catharanthus roseus). cdna, gene, intramolecular autophosphorylation, and identification of a threonine important for auto- and substrate phosphorylation.we characterize crrlk1, a novel type of receptor-like kinase (rlk), from the plant catharanthus roseus (madagascar periwinkle). the protein (90.2 kda) deduced from the complete genomic and cdna sequences is a rlk by predicting a n-terminal signal peptide, a large extracytoplasmic domain, a membrane-spanning hydrophobic region followed by a transfer-stop signal, and a c-terminal cytoplasmic protein kinase with all 11 conserved subdomains. it is a novel rlk type because the predicted extracytoplas ...19968900145
analysis of several iridoid and indole precursors of terpenoid indole alkaloids with a single hplc run.an isocratic hplc system is described which allows the separation of the iridoid and indole precursors of terpenoid indole alkaloids, which are present in a single crude extract. the system consists of a column of lichrospher 60 rp select b 5 microm, 250 x 4 mm (merck) with an eluent of 1% formic acid-acetonitrile-trichloroacetic acid (100:10:0.25, v:v:w) at a flow of 1.2 ml/min. in the suspension cultures of catharanthus roseus secologanin and tryptophan were detected. in the cultures of tabern ...199617252445
characterization of a cdna encoding a proline-rich 14 kda protein in developing cortical cells of the roots of bean (phaseolus vulgaris) seedlings.a cdna clone, corresponding to mrnas preferentially expressed in the roots of bean (phaseolus vulgaris l.) seedlings, was isolated. this clone contains a 381 bp open reading frame encoding a polypeptide of 13.5 kda, designated pvr5 (phaseolus vulgaris root 5). the amino acid sequence of this clone is rich in proline (13.5%) and leucine (12.7%) and shares significant amino acid sequence homology with root-specific and proline-rich proteins from monocots (maize and rice), and proline-rich proteins ...19968639755
enzymology of indole alkaloid biosynthesis in catharanthus roseus.indole alkaloids in catharanthus roseus have been in focus because of their medicinal value. these alkaloids consist of an indole moiety provided by tryptamine and a terpenoid portion provided by the secologanin. the most important catharanthus alkaloids are vinblastine (vlb), vincristine (vcr) and ajmalicine. vlb and vcr are clinically useful anticancer agents whereas ajmalicine is used for the treatment of circulatory diseases. vcr and vlb are the most expensive because of their low abundance ...19968936815
uv-b- and oxidative stress-induced increase in nicotinamide and trigonelline and inhibition of defensive metabolism induction by poly(adp-ribose)polymerase inhibitor in plant tissue.nicotinamide and trigonelline contents increased in catharanthus roseus tissue culture after exposure to 2,2'-azobis(2-amidinopropane)dihydrochloride (aaph) or vanadylsulfate and in pisum sativum leaves after exposure to uv-b radiation. vanadylsulfate increased phenylalanine ammonia-lyase (pal) activity and the content of reduced and oxidized gluthathione in c. roseus tissue culture. the increases in pal activity caused by 2 mm aaph or 0.2mm vanadylsulfate were prevented by 0.1 mm 3-aminobenzami ...19968603735
the role of glucose in ajmalicine production by catharanthus roseus cell cultures.the role of glucose in ajmalicine production by catharanthus roseus was investigated in the second stage of a two-stage batch process. activities of tryptophan decar-boxylate (tdc) and anthranilate synthase (as), two enzymes in the pathway leading to ajmalicine, were higher after induction with 40 g/l glucose than after induction with 60 or 80 g/l glucose. experiments with different media containing mixtures of glucose and the nonpermeating osmotic agent xylose, and using an already induced cult ...199518623431
a cytochrome p-450 monooxygenase catalyzes the first step in the conversion of tabersonine to vindoline in catharanthus roseus.hydroxylation at the c-16 position of the indole alkaloid tabersonine has been suggested as the first step toward vindoline biosynthesis in catharanthus roseus. tabersonine 16-hydroxylase (16-oh) activity was detected in total protein extracts from young leaves of c. roseus using a novel coupled assay system. enzyme activity was dependent on nadph and molecular oxygen and was inhibited by co, clotrimazole, miconazole, and cytochrome c. 16-oh was localized to the endoplasmic reticulum by linear s ...199512228585
plant defense metabolism is increased by the free radical-generating compound aaph.effects of the free radical-generating substance 2,2'-azobis(2-amidinopropane) dihydrochloride (aaph) on defense systems in plant tissue cultures were investigated. exposure of catharanthus roseus, c. tricophyllus, and pisum sativum cultures to aaph caused altered levels of reduced and oxidized glutathione. an increased total glutathione content in c. roseus was prevented by the glutathione biosynthesis inhibitor buthionine-sulfoximine. the specific phenylalanine ammonia-lyase activity in a c. r ...19957557546
a cdna encoding a plant homologue to animal hmg box proteins involved in structure-specific recognition of dna (ssrp family).we report a cdna encoding a 71-kda protein with a single high-mobility group (hmg) box and two nuclear localization signals from the higher plant catharanthus roseus (madagascar periwinkle). the protein had 40% amino acid identity with animal dna-binding proteins of the ssrp (structure-specific recognition protein) family that recognize bent, unwound dna structures. genomic southern analysis suggested the presence of two genes.19957665097
two-stage batch process for the production of ajmalicine by catharanthus roseus: the link between growth and production stage.the link between the growth stage and the production stage in a two-stage batch process was investigated using (filtered) inocula from different periods of the stationary phase of the growth cycle. in the production stage, ajmalicine production by catharanthus roseus in a 3-l stirred tank reactor was induced with a high glucose concentration (80 g/l). ajmalicine production in cultures started with cells from the late stationary phase was five times higher than in cultures started with cells from ...199518623366
lack of control in inorganic phosphate uptake by catharanthus roseus (l.) g. don cells (cytoplasmic inorganic phosphate homeostasis depends on the tonoplast inorganic phosphate transport system?).inorganic phosphate (pi) uptake by catharanthus roseus (l.) g. don cells was studied in relation to its apparent uncontrolled uptake using 31p-nuclear magnetic resonance spectroscopy. kinetics of pi uptake by the cells indicated that apparent km and vm were about 7 [mu]m and 20 [mu]mol g-1 fresh weight h-1, respectively. pi uptake in murashige-skoog medium under different pi concentrations and different initial cell densities followed basically the same kinetics. when supplied with abundant pi, ...199512228474
relation between dissolved oxygen concentration and ajmalicine production rate in high-density cultures of catharanthus roseus.the relation between dissolved oxygen (do) and the ajmalicine production rate of catharanthus roseus was investigated in 15-l tank reactors at constant stirrer speed and gas flow rate. below a do concentration of 29% of air saturation the ajmalicine production rate was less than 0.06 micromol/g/d. above a do of 43% the ajmalicine production rate was constant at 0.21 micromol/g/d. between a do of 29% and 43% there was a strong relation between the ajmalicine production rate and the do concentrati ...199518623236
strictosidine synthase from catharanthus roseus: purification and characterization of multiple forms.multiple (six) forms of strictosidine synthase from catharanthus roseus cell suspension cultures were purified and characterized. a purification protocol is presented composed of hydrophobic-interaction, gel-permeation and ion-exchange chromatography and chromatofocusing. four of six isoforms were purified to apparent homogeneity, whereas two others were nearly homogeneous. all strictosidine synthase isoforms were found to be glycoproteins. the isoforms were also found in leaves and roots of the ...19957887913
cytochrome p-450 in plant/insect interactions: geraniol 10-hydroxylase and the biosynthesis of iridoid monoterpenoids.the interactions between plant secondary metabolites (particularly monoterpenes) and insects are discussed. such metabolites are likely to have influenced the evolution of cyt p450-linked detoxification systems in animals, through animal/plant coevolution. the biosynthesis of many classes of plant secondary metabolites involves cyt p450 enzymes. of these, one of the best characterised is the geraniol/nerol 10-hydroxylase which catalyses a key step in the biosynthesis of the iridoid class of plan ...19958820862
tryptophan decarboxylase, tryptamine, and reproduction of the whitefly.tryptophan decarboxylase (tdc) from catharanthus roseus (periwinkle) converts tryptophan to the indole-alkaloid tryptamine. when the tdc gene was expressed in transgenic tobacco, the 55-kd tdc enzyme and tryptamine accumulated. bemisia tabaci (sweetpotato whitefly) reproduction on transgenic plants decreased up to 97% relative to controls. production of tryptamine, its derivatives, or other products resulting from tdc activity may discourage whitefly reproduction and provide a single-gene-based ...199512228625
overexpression of a tryptophan decarboxylase cdna in catharanthus roseus crown gall calluses results in increased tryptamine levels but not in increased terpenoid indole alkaloid production.the enzyme tryptophan decarboxylase (tdc) (ec 4.1.1.28) catalyses a key step in the biosynthesis of terpenoid indole alkaloids in c. roseus by converting tryptophan into tryptamine. hardly any tdc mrna could be detected in hormone-independent callus and cell suspension cultures transformed by the oncogenic t-dna of agrobacterium tumefaciens. supply of tryptamine may therefore represent a limiting factor in the biosynthesis of alkaloids by such cultures. to investigate this possibility, chimaeric ...19958589734
cdnas for s-adenosyl-l-methionine decarboxylase from catharanthus roseus, heterologous expression, identification of the proenzyme-processing site, evidence for the presence of both subunits in the active enzyme, and a conserved region in the 5' mrna leader.s-adenosyl-l-methionine decarboxylases (adometdc) are pyruvoyl-dependent enzymes producing the aminopropyl group for spermidine biosynthesis, and this reaction is the rate-limiting step in polyamine biosynthesis. we characterized cdnas from catharanthus roseus (madagascar periwinkle) and investigated the enzyme after heterologous expression. the largest cdna (1842 bp) had an 5' leader of 469 bp and encoded a protein of 357 residues and 30-35% identity with mammalian adometdc. the proenzyme expre ...19957883014
cinnamate 4-hydroxylase from catharanthus roseus, and a strategy for the functional expression of plant cytochrome p450 proteins as translational fusions with p450 reductase in escherichia coli.a pcr-based approach was used to isolate cdnas for cinnamate 4-hydroxylase (c4h) from catharanthus roseus cell cultures. the protein shared 75.9% identity with c4h from other plants, and the transcription was induced under various stress conditions. the cloned protein was used to investigate the functional expression of plant p450/p450-reductase fusions in e. coli. fusions containing a modified n-terminal membrane anchor were located in the membrane and possessed c4h activity without solubilizat ...19957589568
vitamin-b12-independent methionine synthase from a higher plant (catharanthus roseus). molecular characterization, regulation, heterologous expression, and enzyme properties.methionine synthases catalyze the formation of methionine by the transfer of a methyl group from 5-methyltetrahydrofolate to homocysteine. this reaction is the last step in l-methionine biosynthesis, and it also serves to regenerate the methyl group of s-adenosylmethionine, a cofactor required for biological methylation reactions. we describe the cloning, expression and characterization of a methionine synthase from the higher plant catharanthus roseus. cdnas were identified that encoded a prote ...19957601135
creation of a metabolic sink for tryptophan alters the phenylpropanoid pathway and the susceptibility of potato to phytophthora infestans.the creation of artificial metabolic sinks in plants by genetic engineering of key branch points may have serious consequences for the metabolic pathways being modified. the introduction into potato of a gene encoding tryptophan decarboxylase (tdc) isolated from catharanthus roseus drastically altered the balance of key substrate and product pools involved in the shikimate and phenylpropanoid pathways. transgenic potato tubers expressing the tdc gene accumulated tryptamine, the immediate decarbo ...199512242360
sensitivity of bedding plants to southern root-knot nematode, meloidogyne incognita race 3.thirty-two cultivars of 10 commonly-grown bedding plants, representing eight families, were evaluated for their response to infection by the root-knot nematode, meloidogyne incognita race 3, under greenhouse conditions. four ageratum cultivars, two marigold, and two salvia cultivars were rated resistant after exposure for 8 weeks. four begonia, four celosia, one dianthus, one verbena, one vinca, and three pansy cultivars were susceptible. three salvia, one begonia, one gerber, one verbena, and t ...199419279964
introduction of oxygenated functional groups into 3-carene and 2-pinene by cultured cells.the biotransformation of the monoterpene hydrocarbons 3-carene and 2-pinene by cell suspension cultures of nicotiana tabacum and catharanthus roseus was investigated. the cultures have the ability to regio- and enantioselectively introduce the oxygenated functional groups into the c = c double bond and the allylic positions of the substrates.19947765622
isolation and characterization of a rice cdna similar to the s-phase-specific cyc07 gene.we isolated a rice cdna clone (t151) which encodes an open reading frame of 262 amino acids. this clone is similar to the s-phase-specific cyc07 gene of catharanthus roseus. expression of this gene is much higher in callus than in seedlings and regulated by external stresses such as high osmotic pressure, salinity, low temperature and submergence.19947510136
suspension culture of catharanthus roseus crown gall cell induced by agrobacterium c58.in comparison with calli from leaf or stem, catharanthus roseus crown gall cell cultured on ms basic medium was superior in growth, total indole alkaloids and ajmalicine contents. the effects of illumination, cultural temperature, sucrose level of the medium and exogenous l-trp on the growth, total indole alkaloids and ajmalicine contents of c. roseus crown gall cell cultures were studied. the results will provide a theoretical basis for the attempt of using suspension cultures of c. roseus crow ...19947893941
nucleotide sequence of the tryptophan decarboxylase gene of catharanthus roseus and expression of tdc-gusa gene fusions in nicotiana tabacum.the enzyme tryptophan decarboxylase (tdc; ec 4.1.1.28) converts tryptophan into tryptamine. in catharanthus roseus and other plants capable of producing terpenoid indole alkaloids (tias) tdc links primary metabolism to the secondary metabolic pathway involved in the biosynthesis of these compounds. the accumulation of tdc mrna in c. roseus cells is developmentally regulated and transcriptionally influenced by elicitors (induction) and auxins (repression). here we report that tdc is encoded by a ...19948159173
mechanisms of the proliferation and differentiation of plant cells in cell culture systems.plant cell functions have been investigated in various cell culture systems. in this review, we summarize results obtained from investigations of gene expression during the cell cycle in synchronized cultures of catharanthus roseus during somatic embryogenesis in suspension cultures of daucus carota, during organogenesis in tissue cultures of arabidopsis thaliana and during the transdifferentiation of isolated mesophyll cells to tracheary elements in single-cell cultures of zinnia elegans.19947981037
elicitor-mediated induction of isochorismate synthase and accumulation of 2,3-dihydroxy benzoic acid in catharanthus roseus cell suspension and shoot cultures.after elicitation, cell suspension cultures of catharanthus roseus accumulate phenolic compounds. the major phenolic compound produced was isolated and identified as 2,3-dihydroxybenzoic acid (dhba). the accumulation of this compound is a rapid response to the addition of elicitor; within 6 h after the addition of elicitor, dhba concentration reached 6.3 mg/l cell suspension. dhba was not detected in non-elicited cells. the formation of dhba in elicited cells was correlated with the induction of ...199424192892
the negligible role of carbon dioxide and ethylene in ajmalicine production by catharanthus roseus cell suspensions.removal of gaseous metabolites in an aerated fermenter affects ajmalicine production by catharanthus roseus negatively. therefore, the role of co2 and ethylene in ajmalicine production by c. roseus was investigated in 3 l fermenters (working volume 1.8 l) with recirculation of a large part of the exhaust air. removal of co2, ethylene or both from the recirculation stream did not have an effect on ajmalicine production. inhibition of ethylene biosynthesis in shake flasks with co(2+), ni(2+) or am ...199424192885
relationship between cell morphology and indole alkaloid production in suspension cultures of catharanthus roseus.the relationship between the morphology and indole alkaloid production of catharanthus roseus cells was investigated. eleven cell lines were randomly selected from protoplast-derived clones. in each line, most of the cells maintained only one of the two shapes, either spherical or cylindrical. the cell aspect ratio (cell length/width) for most isolates was stable for more than two years of subculture. cell division patterns of spherical and cylindrical cell isolates were different and patterns o ...199424194221
effects of mercury (ii) species on cell suspension cultures of catharanthus roseus. 19947833617
application of high-performance liquid chromatography to the determination of vinblastine in catharanthus roseus.a method for the determination of vinblastine in catharanthus roseus by hplc is described. a crude alkaloid extract, obtained by extraction of leaves with toluene, 2% citric or tartaric acid and benzene, was separated by tlc. the vinblastine fraction was cut out and vinblastine was eluted from the sorbent. the amount of vinblastine was determined by hplc with peak-height measurement. the standard deviation is 0.2 microgram/ml. the detection limit is 0.05 ng of vinblastine in a sample applied to ...19947866534
effects of oxygen and nutrients limitation on ajmalicine production and related enzyme activities in high density cultures of catharanthus roseus.oxygen and nutrient limitation was investigated in order to identify the origin of a lower specific ajmalicine production in catharanthus roseus cultures at high cell densities in an induction medium. the effect of oxygen limitation was explored by comparing two identically aerated and agitated high cell density bioreactor cultures with dissolved oxygen (do) concentration of 15% and 85% of air saturation, with respect to alkaloid formation and related enzymes activities. oxygen had an evident ef ...199418618780
phosphatidate kinase, a novel enzyme in phospholipid metabolism (characterization of the enzyme from suspension-cultured catharanthus roseus cells).phosphatidate kinase (adenosine 5[prime]-triphosphate:phosphatidic acid phosphotransferase), a novel enzyme of phospholipid metabolism, was detected recently in the plasma membranes of suspension-cultured catharanthus roseus cells and purified (j.b. wissing, h. behrbohm [1993] plant physiol 102: 1243-1249). in the present work the properties of phosphatidate kinase are described. the enzyme showed a ph optimum of 6.1 and an isoelectric point of 4.8, and was rather stable in the presence of its s ...199412232252
large-scale cultivation of catharanthus roseus cells: production of ajmalicine in a 20-l airlift bioreactor.bioreactor systems have been developed for the production of ajmalicine, an alkaloid used in the treatment of hypertension. cell cultures of catharanthus roseus produced higher levels of ajmalicine (323 micrograms g-1 dry weight) in a production medium enriched with tryptophan. the cell cultures were grown in medium prepared in tap water and market sugar with a view to minimise the costs of production. large-scale cultivation of cell suspension was performed in a 20-l airlift bioreactor under co ...19947765026
hexose uptake by catharanthus roseus cell suspensions is inhibited by a high medium salt content.the uptake of glucose and fructose from the medium by catharanthus roseus cell suspensions was strongly inhibited by high medium salt concentration, such as found in ls (linsmaier and skoog 1965) medium. after inoculation into standard ls nutrient medium with less than 5 mm hexose no uptake occurred, while in low salt medium hexose was completely depleted. at a hexose concentration of 50 mm the uptake rate was higher in low salt medium than in standard medium. the lower rate of uptake at high sa ...199424194027
long-term effect of a pythium elicitor treatment on the growth and alkaloid production of catharanthus roseus cell suspensions.the treatment of a catharanthus roseus cell suspension culture with a low concentration of pythium elicitor stimulated the alkaloid production. when these pretreated cells were resuspended in a medium that did not contain the fungal extract, the positive effects of the treatment on alkaloid synthesis and excretion were lost and, moreover, the standard level of production was not recovered. a second treatment of these cells with pythium elicitor at day 5 of the second culture cycle greatly impair ...199417236032
high frequency plant regeneration from anther-derived cell suspension cultures via somatic embryogenesis in catharanthus roseus.a system for high frequency plant regeneration from cell suspension cultures in catharanthus roseus is described. calli were obtained from anthers cultured on murashige and skoog's medium supplemented with 1 mgl(-1) α-naphthaleneacetic acid and 0.1 mgl(-1) kinetin. after the second subculture on solid medium, embryogenic callus was identified and transferred to liquid medium to initiate suspension cultures. cells dispersed finely in the medium were subcultured at 14-day intervals. upon plating o ...199424193829
meristem-specific gene expression directed by the promoter of the s-phase-specific gene, cyc07, in transgenic arabidopsis.a genomic clone for the cyc07 gene, which is expressed specifically at the s phase during the cell cycle in synchronous cultures of periwinkle (catharanthus roseus) cells, was isolated. determination of the nucleotide sequence of the clone revealed that the cyc07 gene consists of seven exons separated by six introns. genomic southern analysis indicated that the cyc07 gene is present as a single copy per haploid genome in periwinkle. expression of related genes was detected in a wide range of oth ...19948204824
cdna for s-adenosyl-l-homocysteine hydrolase from catharanthus roseus. 19948165255
cdna for a 14-kilodalton polypeptide from madagascar periwinkle (catharanthus roseus). eight conserved cysteines in six related proteins from different plants suggest common functional elements. 19948165254
2,4-d and alkaloid accumulation in periwinkle cell suspensions.omission of 2,4-d from culture medium during one subculture of catharanthus roseus cells, strain c20, resulted in an increased alkaloid accumulation, without effect on growth. alkaloid accumulation, rather than growth, seemed to be more sensitive to 2,4-d. 2,4-d inhibited alkaloid accumulation essentially during growth phase, but its inhibitory effect during this period was partially reversible. as this reversibility was underlined only during the stationary phase, this suggested that this actio ...19947849107
purification and partial characterization of microsomal nadh-cytochrome b5 reductase from higher plant catharanthus roseus.a simple three step procedure was used to purify microsomal nadh-cytochrome b5 (ferricyanide) reductase to homogeneity from the higher plant c. roseus. the microsomal bound reductase was solubilized using zwitterionic detergent-chaps. the solubilized reductase was subjected to affinity chromatography on octylamino sepharose 4b, blue 2-sepharose cl-6b and nad(+)-agarose. the homogeneous enzyme has an apparent molecular weight of 33,000 as estimated by sds-page. the purified enzyme catalyzes the r ...19938267585
putative sites of cytokinin action during their enhancing effect on indole alkaloid accumulation in periwinkle cell suspensions.the effects of cytokinins on the different branches of the indole alkaloid pathway were investigated in catharanthus roseus cell cultures. addition of zeatin to a 2,4-dichlorophenoxyacetic acid-containing medium decreased tryptamine levels and increased the bioconversion of secologanin to ajmalicine. zeatin also enhanced the geraniol-10 hydroxylase activities and modified the indole alkaloid pattern. the results are discussed in the light of previous works showing that cytokinins have a positive ...199324201970
effect of terpenoid precursor feeding and elicitation on formation of indole alkaloids in cell suspension cultures of catharanthus roseus.the effects of terpenoid precursor feeding and elicitation by a biotic elicitor on alkaloid production of catharanthus roseus suspension cultures were studied. after addition of secologanin, loganin or loganic acid an increase in the accumulation of ajmalicine and strictosidine and a decrease of tryptamine level was observed in non-elicited cells. elicitation increased tryptamine accumulation in non-fed cells but it did not further increase alkaloid accumulation in precursor-fed cells. a decreas ...199324201968
induced responses in three alkaloid-containing plant species.in this paper we test three plant species for the inducibility of their alkaloid production. the plants were heavily damaged by cutting off 50% of their leaf surface using a pair of scissors. the cut-off leaf tips were used as controls for possible diurnal fluctuations. after 3, 6, 12, 24 and 48 h, respectively, the leaf bases of the damaged plants were harvested and the alkaloid concentration was measured. in senecio jacobaea the pyrrolizidine alkaloid (pa) concentration in damaged plants decre ...199328314020
subcellular localization of tryptophan decarboxylase, strictosidine synthase and strictosidine glucosidase in suspension cultured cells of catharanthus roseus and tabernaemontana divaricata.the subcellular localization of tryptophan decarboxylase, strictosidine synthase and strictosidine glucosidase in suspension cultured cells of catharanthus roseus (l.) g. don and tabernaemontana divaricata (l.) r. br. ex roem. et schult, was investigated. it was found that tryptophan decarboxylase is an extra-vacuolar enzyme, whereas strictosidine synthase is active inside the vacuole. strong indications were obtained for the localization of strictosidine glucosidase on the outside of the tonopl ...199324201788
phosphatidate kinase, a novel enzyme in phospholipid metabolism (purification, subcellular localization, and occurrence in the plant kingdom).microsomal membranes from suspension-cultured catharanthus roseus cells possess an enzymic activity that catalyzes the atp-dependent phosphorylation of phosphatidic acid (pa) to form diacylglycerol pyrophosphate (h. behrbohm, j.b. wissing [1993] febs lett 315: 95-99). this enzyme activity, pa kinase, was purified and characterized. plasma membranes, obtained from c. roseus microsomes by aqueous two-phase partitioning, were extracted, and pa kinase was purified 3200-fold by applying different chr ...199312231900
isolation of cytochrome p-450 cdna clones from the higher plant catharanthus roseus by a pcr strategy.cytochrome p-450 monooxygenases are membrane-bound enzymes involved in a wide range of biosynthetic pathways in plants. an efficient pcr strategy for isolating cytochrome p-450 cdna clones from plant cdna libraries is described. a set of degenerate primers for pcr amplification was designed to recognize nucleotide sequences specifying the highly conserved haembinding region of cytochrome p-450 proteins. using this primer set and a non-specific primer, complementary to either the poly(a) tail of ...19938507838
a structured model describing carbon and phosphate limited growth of catharanthus roseus plant cell suspensions in batch and chemostat culture.the growth of plant cell suspension cultures of catharanthus roseus in batch fermentors was studied at different initial phosphate levels of the medium. on the basis of the observations and existing knowledge with respect to phosphate metabolism in cultured c. roseus cells, a structured mathematical model was developed for the description of the kinetics of growth and intracellular accumulation of glucose and phosphate, as a function of glucose and phosphate supply. it was shown that the model o ...199318609621
induction of ajmalicine formation and related enzyme activities in catharanthus roseus cells: effect of inoculum density.in catharanthus roseus cell cultures the time courses of four enzyme activities, tryptophan decarboxylase (tdc), strictosidine synthase (sss), geraniol-10-hydroxylase (g10h) and anthranilate synthase (as), and alkaloid accumulation were compared under two different culture conditions (low-inoculum density and high-inoculum density on induction medium) and a control on growth medium. in growth medium a transient increase in tdc activity was first observed after which g10h reached its maximum acti ...19937763550
purification, characterization, and kinetic analysis of a 2-oxoglutarate-dependent dioxygenase involved in vindoline biosynthesis from catharanthus roseus.a 2-oxoglutarate-dependent dioxygenase (ec 1.14.11.11) which catalyzes the hydroxylation at position 4 of the indole alkaloid, desacetoxyvindoline has been purified to near homogeneity from catharanthus roseus. the purification procedure combined conventional chromatographic methods and cosubstrate affinity chromatography on alpha-ketoglutarate-sepharose. the specific activity of the 4-hydroxylase was enriched over 2000-fold compared to the crude homogenate with a recovery of 1.6%. the molecular ...19938449913
effects of near-ultraviolet light on alkaloid production in catharanthus roseus plants.artificial near-ultraviolet light with a peak at 370 nm and the light of natural radiation with wavelengths between 290 and 380 nm stimulated the synthesis of dimeric indole alkaloids in intact plants of catharanthus roseus. the artificial light also specifically stimulated an in vitro fmn-mediated, non-enzymatic coupling of vindoline and catharanthine to synthesize an iminium intermediate, the in vivo precursor of dimeric alkaloid synthesis. these results suggest that near-ultraviolet light is ...199317230336
scaleup of ajmalicine production by plant cell cultures of catharanthus roseus.the effect of scaleup on he production of ajmalicine by a catharanthus roseus cell suspension culture in a selected induction medium were studied. in preliminary experiments it was observed that the culture turned brown and the production was inhibited upon transfer from a shake flask to a stirred bioreactor with forced aeration. two factors were recognized as the potential origin of the differences between shake flask and bioreactor cultures: gas composition and mechanical shear forces. these f ...199318609545
isolation and characterization of a cdna clone from catharanthus roseus encoding nadph:cytochrome p-450 reductase, an enzyme essential for reactions catalysed by cytochrome p-450 mono-oxygenases in plants.the membrane-bound flavoprotein nadph:cytochrome p-450 (cytochrome c) reductase, that functions in electron transfer to cytochrome p-450 monooxygenases, was purified from a cell suspension culture of the higher plant catharanthus roseus. anti-serum raised against the purified protein was found to inhibit nadph:cytochrome c reductase activity as well as the activities of the cytochrome p-450 enzymes geraniol 10-hydroxylase and trans-cinnamate 4-hydroxylase, which are involved in alkaloid biosynth ...19938220474
isolation and characterization of a cdna clone for plant nuclear antigen 21d7 associated with cell division.a cdna clone was isolated from a carrot (daucus carota l.) cdna expression library using monoclonal antibody 21d7, which recognizes a nuclear antigen associated with cell division in plant cells. to show that the isolated cdna encodes the 21d7 antigen, a polyclonal antiserum was raised against a recombinant fusion protein specified by the cdna. both the polyclonal antiserum and the monoclonal antibody 21d7 recognized the same plant protein on immunoblots, in immunoprecipitation experiments, and ...19938310059
a chimaeric tryptophan decarboxylase gene as a novel selectable marker in plant cells.a novel selection system for plant genetic transformation was developed based on the enzyme tryptophan decarboxylase (tdc; ec 4.1.1.28) from catharanthus roseus. this enzyme converts the toxic tryptophan analogue 4-methyl tryptophan (4-mt) into the non-toxic compound 4-methyl tryptamine. expression of tdc in transgenic plants that have no endogenous tdc-activity allows selection on 4-mt. a vector was constructed containing a tdc cdna clone under control of the constitutively expressed cauliflowe ...19938358036
exploration of the frontiers of tradomedical practices: basis for development of alternative medical healthcare services in developing countries.the study is a brief exploration of the functions and roles of the traditional healers in the total health care delivery system as a basis for tapping the salient features of this age old art: for the purpose of refining, and establishing it as an alternative medical health-care service. the investigation is considered relevant particularly in the developing countries where, in addition to the dearth of orthodox medical services, institutions and personnel, it is relatively cheaper, socio-cultur ...19938410912
purification and characterization of anthranilate synthase from catharanthus roseus.anthranilate synthase (ec 4.1.3.27) has been purified from cell cultures of catharanthus roseus by poly(ethylene glycol) precipitation/fractionation and subsequent separation by anion exchange on q-sepharose, orange a dye chromatography, mono q anion-exchange chromatography and superose 6 gel filtration. by analogy to anthranilate synthases from other sources it does look like the enzyme is a tetramer composed of two large and two small subunits, with molecular mass 67 and 25.5 +/- 0.5 kda, resp ...19938444181
hsp90 homologue from madagascar periwinkle (catharanthus roseus): cdna sequence, regulation of protein expression and location in the endoplasmic reticulum.we describe cdnas for a hsp90 homologue from catharanthus roseus and studies on the regulation of expression. the largest cdna (2670 bp) coded for a protein of 817 amino acids with a calculated size of 93,491 da and a pi of 4.61. it contained a eucaryotic secretory signal, the endoplasmic reticulum (er) targeting and retention signal (lys-asp-glu-leu), and the hsp90 protein family signature with one conservative exchange (asn-lys-asp-ile-phe-leu instead of asn-lys-glu-ile-phe-leu). rna blots rev ...19938106014
production of indole alkaloids by selected hairy root lines of catharanthus roseus.hairy root cultures of catharanthus roseus were established by infection of seedlings with agrobacterium rhizogenes 15834. approximately 150 transformants from four different. c. roseus cultivars were screened for desirable traits in growth and indole alkaloid production. five hairy root clones grew well in liquid culture with doubling times similar to those reported for cell suspensions. fast growing clones had similar morphologies, characterized by thin, straight, and regular branches with thi ...199318609590
kinetics and stoichiometry of growth of plant cell cultures of catharanthus roseus and nicotiana tabacum in batch and continuous fermentors.plant cell suspension cultures of catharanthus roseus and nicotiana tabacum were grown in stirred tank bioreactors operated in batch and continuous mode. the stoichiometry of growth of both species in steady-state glucose limited chemostats was studied at a range of different dilution rates. a linear relation was applied to describe specific glucose uptake, oxygen consumption, and carbon dioxide production as a function of the growth rate. specific respiration deviated greatly from the linear re ...199218601193
visualization of vacuoplasts in isolated vacuole preparations from mesophyll protoplasts of periwinkle [catharanthus roseus (l.) g. don].a procedure was developed for the rapid detection of vacuoplasts in vacuole preparations isolated from mesophyll protoplasts of catharanthus roseus (l.) g. don (periwinkle). the procedure relies on the staining of surface carbohydrates on the plasma membrane surrounding vacuoplasts with fluorescein-labeled lectins. when isolated under conditions of constant osmotic strength, approximately 15-20% of the vacuoles isolated showed surface labeling with fitc-agglutinin from abrus precatorius. isolati ...199224203146
transient induction of tryptophan decarboxylase (tdc) and strictosidine synthase (ss) genes in cell suspension cultures of catharanthus roseus.when cell suspension cultures of catharanthus roseus are treated with autoclaved elicitor from the fungus pythium aphanidermatum, they respond with the rapid transient induction of tryptophan decarboxylase (tdc) and strictosidine synthase (ss) enzyme activities, followed by the accumulation of indole alkaloids (eilert et al., 1987). in this report, we demonstrate that expression of tdc and ss enzyme activities is preceeded by the transient appearance of mrnas for both enzymes, suggesting transcr ...199224213491
purification of an indole alkaloid biosynthetic enzyme, strictosidine synthase, from a recombinant strain of escherichia coli.the gene for the indole alkaloid biosynthetic enzyme, strictosidine synthase, of catharanthus roseus has been cloned into an inducible escherichia coli expression vector using an expression cassette polymerase chain reaction technique. induction of the gene resulted in overexpression of the enzyme which accumulated mainly as insoluble inclusion bodies. denaturation and refolding of the insoluble protein resulted in the ability to purify up to 6 mg of active enzyme from a single liter of cell cul ...19921330135
a gene family homologous to the s-phase specific gene in higher plants is essential for cell proliferation in saccharomyces cerevisiae.previously we reported the isolation and characterization of the gene, cyc07, which was specifically expressed in the s phase during the cell cycle in synchronous cell division cultures of the higher plant, catharanthus roseus. we found that the yeast saccharomyces cerevisiae contains two closely related genes which show a high degree of similarity (about 64% at the amino acid level) to cyc07 of c. roseus. site-directed disruption mutations demonstrated that the two yeast genes, homologous to cy ...19921451783
the pharmacology of extinction.it is impossible to predict what compounds of pharmacological interest may be present in an unexamined species. the extinction of such species may result, therefore, in the loss of therapeutically significant compounds. the fact that science will never know what has been lost does not lessen the significance of the loss. a number of species are discussed to exemplify the potential loss. ginkgo biloba is an ancient plant, apparently saved from a natural extinction by human intervention. from this ...19921453701
sensitive detection and identification of mycoplasma-like organisms in plants by polymerase chain reactions.dna amplification by polymerase chain reactions (pcr) was employed to detect host plant infection by several mycoplasmalike organisms (mlos), including the aster yellows (ay), dwarf aster yellows (day), and periwinkle little leaf (0-1) mlos. for pcr, two pairs of oligonucleotide primers, designated ay18pm and ay19pm, respectively, were synthesized on the basis of partial sequences of cloned ay mlo dna fragments ay18 and ay19. reaction mixtures containing primer pair ay18pm yielded a dna product ...19921510676
auxin rapidly down-regulates transcription of the tryptophan decarboxylase gene from catharanthus roseus.the enzyme tryptophan decarboxylase (tdc) (ec 4.1.1.28) converts tryptophan into tryptamine, and thereby channels primary metabolites into indole alkaloid biosynthesis. the production of these secondary metabolites in suspension cells of catharanthus roseus depends on medium composition. of the possible variables, we investigated the effect of hormones on the expression of the tdc gene in cell cultures. omission of naa from the growth medium resulted in accumulation of tdc mrna. the addition of ...19921600147
coordinated regulation of two indole alkaloid biosynthetic genes from catharanthus roseus by auxin and elicitors.catharanthus roseus (periwinkle) produces a wide range of terpenoid indole alkaloids, including several pharmaceutically important compounds, from the intermediate strictosidine. the complete mrna sequence for the enzyme strictosidine synthase (sss) was determined. comparison of the primary structure of the encoded protein with the amino-terminal sequence of purified sss indicated the presence of a signal peptide of 31 amino acids in the putative primary translation product. sss is encoded by a ...19921600148
molecular analysis and heterologous expression of an inducible cytochrome p-450 protein from periwinkle (catharanthus roseus l.).we screened cdna libraries from periwinkle (catharanthus roseus) cell cultures induced for indole alkaloid synthesis and selected clones for induced cytochrome p-450 (p-450) proteins by differential hybridization, size of the hybridizing mrna, and presence of amino acid motifs conserved in many p-450 families. four cdnas satisfying these criteria were analyzed in detail. they were grouped in two classes (pcros1, pcros2) that represented two closely related genes of a new p-450 family designated ...199216653087
nucleotide sequence of a cdna encoding 3-hydroxy-3-methylglutaryl coenzyme a reductase from catharanthus roseus. 199216653173
phytochrome is involved in the light-regulation of vindoline biosynthesis in catharanthus.the enzyme acetylcoenzyme a:deacetylvindoline 4-o-acetyl-transferase (dat) catalyzes the final step in the biosynthesis of the monoterpenoid indole alkaloid, vindoline. previous studies have shown that the appearance of dat activity in etiolated seedlings of catharanthus roseus is induced by exposure of seedlings to light and that enzyme activity is restricted principally to the cotyledons. evidence is now presented that phytochrome is involved in the light-mediated induction of dat activity in ...199216653011
auxins induce tryptophan decarboxylase activity in radicles of catharanthus seedlings.germinating seedlings of catharanthus roseus produce monoterpenoid indole alkaloids as a result of a transient increase of tryptophan decarboxylase (tdc) activity. the influence of auxins on this transient rise of tdc activity was studied. external application of indolebutyric acid or 2,4-dichlorophenoxyacetic acid at a concentration of 20 to 40 mum enhanced and prolonged the rise in tdc activity in developing seedlings. auxin treatment also influenced the morphology of the seedlings; it induced ...199216653009
biotransformation of tabersonine in cell suspension cultures of catharanthus roseus.to investigate the reactions involved in the biosynthesis of vindoline from tabersonine, the bioconversion products formed when the latter compound was fed to cell suspension cultures of catharanthus roseus were isolated and characterized. two biotransformation products of tabersonine were isolated and shown to be lochnericine, which is formed by epoxidation of tabersonine at positions 14, 15, and lochnerinine, the 11-methoxylation product of lochnericine. the bioconversion ratio of the main bio ...19921368411
identification of an essential histidine residue at the active site of the tonoplast malate carrier in catharanthus roseus cells.the involvement of a histidyl residue in the binding or translocation step was investigated in the malate carrier at the tonoplast of catharanthus roseus cells. the transport rate was strongly stimulated when the ph of the incubation medium was decreased from ph 7.0 to 5.0. the histidine-specific reagent diethylpyrocarbonate (depc) efficiently inhibited the activity of the malate carrier. inhibition developed rapidly and was completed after 5 min at a concentration of 2 mm depc. the original sub ...19921433274
cytokinin-enhanced accumulation of indole alkaloids in catharanthus roseus cell cultures - the factors affecting the cytokinin response.cytokinins were found to stimulate the alkaloid synthesis induced by removing auxin from the medium of a cell line of catharanthus roseus. diluting the mineral salts of the culture medium decreased the alkaloid production but increased the "sensitivity" of the cells. addition of high levels of ca(2+), mg(2+) or sr(2+) to b5 media in which the mineral salts were diluted to 5-40%, increased the alkaloid production. the latter effect is related only partially to enhanced osmotic potential.199224201541
the effect of temperature on hairy root cultures of catharanthus roseus: growth, indole alkaloid accumulation and membrane lipid composition.cultivation of catharanthus roseus hairy root cultures at different temperatures was found to have an effect on growth rate and indole alkaloid content as well as lipid composition. when lowering the temperature, the roots responded by increasing the degree of unsaturation of cellular lipids, which was mainly due to an increased proportion of linolenic acid in the main lipid classes. the modifications in lipid composition were obviously necessary for the roots to retain the proper cell membrane ...199224201540
the effect of temperature on growth, indole alkaloid accumulation and lipid composition of catharanthus roseus cell suspension cultures.cell suspension cultures of catharanthus roseus were used to study the effect of temperature on plant cell lipids and indole alkaloid accumulation. lowering the cultivation temperature increased the total fatty acid content per cell dry weight relative to that at higher temperatures, mainly because of increased accumulation of unsaturated c18 acids. in addition, an increase in the relative proportion of phosphatidylcholine and phosphatidylethanolamine was observed. within individual lipids, the ...199224201539
cytoplasmic acidification induced by inorganic phosphate uptake in suspension cultured catharanthus roseus cells: measurement with fluorescent ph indicator and p-nuclear magnetic resonance.cytoplasmic acidification during inorganic phosphate (pi) absorption by catharanthus roseus cells were studied by means of a fluorescent ph indicator, 2',7'-bis-(2-carboxyethyl)-5 carboxyfluorescein (acetomethylester) (bcecf), and (31)p-nuclear magnetic resonance spectroscopy. cytoplasmic acidification measured by decrease in the fluorescence intensity started immediately after pi application. within a minute or so, a stable state was attained and no further acidification occurred, whereas pi ab ...199216668939
development of a helical-ribbon impeller bioreactor for high-density plant cell suspension culture.a double helical-ribbon impeller (hri) bioreactor with a 11-l working volume was developed to grow high-density catharanthus roseus cell suspensions. the rheological behavior of this suspension was found to be shear-thinning for concentrations higher than 12 to 15 g dw . l(-1). a granulated agar suspension of similar rheological properties was used as a model fluid for these suspensions. mixing studies revealed that surface baffling and bottom profiling of the bioreactor and impeller speeds of 6 ...199218600977
diacylglycerol kinase from suspension cultured plant cells : characterization and subcellular localization.diacylglycerol kinase (adenosine 5'-triphosphate:1,2-diacylglycerol 3-phosphotransferase, ec 2.7.1.107), purified from suspension cultured catharanthus roseus cells (j wissing, s heim, kg wagner [1989] plant physiol 90: 1546-1551), was further characterized and its subcellular location was investigated. the enzyme revealed a complex dependency on lipids and surfactants; its activity was stimulated by certain phospholipids, with phosphatidylinositol and phosphatidylglycerol as the most effective ...199216668739
first field isolation of wound tumor virus from a plant host: minimal sequence divergence from the type strain isolated from an insect vector.a new strain of wound tumor virus (wtv) has been isolated from a periwinkle plant (catharanthus roseus) that was among several used as bait plants in a blueberry field. the 12 segments of double-stranded rna of the viral genome were isolated directly from infected tissue and found to have mobilities through agarose gels that were identical to those of the type strain wtv. coupled complementary dna (cdna) and polymerase chain reactions (pcr) primed with oligonucleotides complementary to the termi ...19911962460
large-scale (20 l) culture of surface-immobilized catharanthus roseus cells.surface-immobilized c. roseus cell cultures were grown in a 20-l modified airlift bioreactor operated at 0.51 vvm (kla approximately 8 h-1) under various gassing regimes [air, 2% (v/v) and 5% co2]. extracellular ammonium, phosphate, and nitrate ions as well as carbohydrate uptake and ph value of the medium were monitored together with on-line dissolved oxygen concentration, conductivity of the medium, and carbon dioxide production rate (cpr) of the cultures. cultures supplemented with 2% co2 sho ...19911367999
stimulated indole alkaloid release from catharanthus roseus immobilized cultures. initial studies.vacuolar sequestration of valuable secondary metabolites remains the major limitation to the use of immobilization technology for large scale plant-cell-based bioprocesses, which otherwise may be a more efficient culture system than suspension for this biomass. in this initial study, the release of indole alkaloids produced by immobilized catharanthus roseus cells cultured in zenk's alkaloid production medium was evaluated. unstimulated alkaloid release in immobilized cultures reached levels of ...19911367690
effect of culture process on alkaloid production by catharanthus roseus cells. ii. immobilized cultures.two processes for the production of indole alkaloids 2 l surface-immobilized bioreactor cultures of catharanthus roseus cells using zenk's alkaloid production medium (apm) were evaluated. the 1-stage process consisted of inoculating apm containing bioreactors and incubating for 15 d. the 2-stage process involved inoculating growth medium-containing bioreactors, growing the immobilized cultures for a certain period of time and subsequently replacing this medium with apm. the production stage whic ...19911367689
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