increase in the indole alkaloid production and its excretion into the culture medium by calcium antagonists in catharanthus roseus hairy roots.treatment of catharanthus roseus hairy roots with antagonists, like verapamil and cdcl2, that block the ca2+ flux across the plasma membrane enhanced the total alkaloid content by 25% and their secretion 10 times. the specific ca2+ chelator, egta, stimulated 90% of the total alkaloid secretion. treatment with inhibitors of intracellular ca2- movement, like tmb-8 and trapsigargin, enhanced the total alkaloid content by 74% and their secretion into the culture media by 4- to 6-fold. the results su ...200314514063
peroxidase from catharanthus roseus (l.) g. don and the biosynthesis of alpha-3',4'-anhydrovinblastine: a specific role for a multifunctional enzyme.we have characterized a basic peroxidase with alpha-3',4'-anhydrovinblastine (avlb) synthase activity, which was purified from catharanthus roseus leaves. this enzyme was the single peroxidase isoenzyme detected in c. roseus leaves, and the single avlb synthase activity detected in c. roseus extracts. it was observed that the monomeric substrates of avlb, vindoline and catharanthine, are both suitable electron donors for the oxidizing intermediates of the basic peroxidase, compounds i and ii. re ...200314513315
jasmonate-induced epoxidation of tabersonine by a cytochrome p-450 in hairy root cultures of catharanthus roseus.methyl jasmonate, a chemical inducer of secondary metabolism, was shown to promote tabersonine 2 biosynthesis in hairy root cultures of catharanthus roseus. tabersonine 6,7-epoxidase activity was detected in total protein extract of jasmonate-induced hairy root cultures using labeled 14c-tabersonine 2. this enzyme converted tabersonine 2 to lochnericine 3 by selective epoxidation at positions 6 and 7 via a reaction dependent on nadph and molecular oxygen. carbon monoxide, clotrimazole, miconazol ...200312943756
natural products of plant origin as anticancer agents.natural products have been used as effective remedies for the treatment of various ailments. numerous plant products in the form of decoction, tincture, tablets and capsules have been clinically used for the treatment of different kinds of cancer. this review covers some of the important plants with clinically proven anticancer activity, including catharanthus roseus, podophyllum peltatum, taxus brevifolia, camptothecin acuminata, cephalotaxus harringtonia, viscum album, onchrosia elliptica, ann ...200112806432
role of the non-mevalonate pathway in indole alkaloid production by catharanthus roseus hairy roots.the 1-deoxy-d-xylulose-5-phosphate (dxp) pathway (non-mevalonate pathway) leading to terpenoids via isopentenyl diphosphate (ipp) has been shown to occur in most bacteria and in all higher plants. treatment with the antibiotic fosmidomycin, a specific inhibitor of dxp reductoisomerase, considerably inhibited the accumulation of the alkaloids ajmalicine, tabersonine, and lochnericine by catharanthus roseus hairy root cultures in the exponential growth phase. however, fosmidomycin did not signific ...201312790690
studies on production of ajmalicine in shake flasks by multiple shoot cultures of catharanthus roseus.the effects of different concentrations of indole-3-acetic acid (iaa) and benzyladenine (ba) on production of ajmalicine by multiple shoot cultures of catharanthus roseus (c. roseus) were studied. by supplementing murashige and skoog's (ms) medium with a high concentration of iaa (11.42 microm) and a low concentration of ba (2.22 microm), shoot cultures accumulated high levels of ajmalicine. when culture medium was fortified with a low concentration of iaa (2.85 microm) and a high concentration ...201612790683
screening of medicinal plants used in south african traditional medicine for genotoxic effects.dichloromethane and 90% methanol extracts from 51 south african medicinal plants were evaluated for potential genotoxic effects using the bacterial ames and vitotox tests with and without metabolic activation. dichloromethane extracts from bulbs of crinum macowanii showed mutagenicity in strain ta98 with and without metabolic activation, whereas extracts from leaves of chaetacme aristata and foliage of plumbago auriculata showed mutagenicity and/or toxicity. extracts from the leaves of catharant ...200312749823
production of anthocyanins by catharanthus roseus.a stable cell suspension line of catharanthus roseus producing anthocyanin was obtained. in this strain it was found that approximately 30% of cells regularly accumulated these metabolites and that anthocyanin accumulation occurred between the second half of log phase and the stationary phase of the culture growth cycle. the anthocyanins in the suspension cultures were compared with those biosynthetized in the flowers both of regenerated by somatic embryogenesis and field-grown plants. six antho ...200312628396
quantification of metabolic flux in plant secondary metabolism by a biogenetic organizational approach.metabolic engineering represents a promising approach to enhance the yield of valuable natural products from plants. a method to quantify flux through metabolite measurements is necessary for the analysis of native and modified pathways. rather than focusing only on the accumulation of the final products, analyzing a wide range of secondary metabolites has significant advantages. we propose a model that organizes the flux analysis by grouping metabolites of similar biosynthetic origin. to this e ...200212616695
immunocytolocalization of tryptophan decarboxylase in catharanthus roseus hairy roots.we investigated the intracellular distribution of tryptophan decarboxylase (tdc) (ec in catharanthus roseus hairy roots using immunofluorescence and immunogold techniques. tdc was detected by immunofluorescence localization in the cytosol and in the apoplastic region of the meristematic cells of the roots, with a slight enrichment in the epidermal cells of the root cap and in the meristematic region. in the enlargement zone, tdc was localized only in the first three layers of the corte ...200312611265
inhibition of the plant cytokinin transduction pathway by bacterial histidine kinase inhibitors in catharanthus roseus cell cultures.we describe the isolation of two catharanthus roseus cdnas encoding proteins putatively involved in the final steps of a 'histidine-to-aspartate' phosphorelay in cytokinin (ck) signaling. the expression of one of these genes, crrr1, was specifically up-regulated by cks in c. roseus cell suspensions. we used this system as a biological model to test the activity of bacterial histidine kinase inhibitors. our data demonstrate that these inhibitors are active on the ck transduction pathway and repre ...200312606039
salt-tolerant mutants in glycophytic salinity response (gsr) genes in catharanthus roseus.the periwinkle catharanthus roseus shares glycophytic properties of crop plants. to contribute towards an understanding of the glycophytic response to salinity, large populations of m(2) seeds having an origin in nitroso-methyl urea and ethyl methane sulphonate treatments were screened for germination with 250 mm of nacl. out of the nine mutant lines so recovered, which tolerated salt stress due to loss of the normal glycophytic salinity response ( gsr), the characteristics of six gsr mutants ar ...200312582847
biotransformation of 2alpha,5alpha,10beta,14beta-tetraacetoxy-4(20),11-taxadiene by cell suspension cultures of catharanthus roseus.catharanthus roseus cell suspension cultures were employed for the biotransformation of 2alpha,5alpha,10beta,14beta-tetraacetoxy-4(20),11-taxadiene ( 1), and four metabolites were obtained. based on their physical and chemical data, the structures of the four metabolites were respectively elucidated as 10beta-hydroxy-2alpha,5alpha,14beta-triacetoxy-4(20),11-taxadiene (2), 5alpha-hydroxy-2alpha,10beta, 14beta-triacetoxy-4(20),11-taxadiene (3), 6alpha,10beta-dihydroxy-2alpha,5alpha,14beta-triaceto ...200212494340
characterization of an inducible promoter system in catharanthus roseus hairy roots.transgenic hairy root cultures of catharanthus roseus were established with a glucocorticoid-inducible promoter controlling the expression of green fluorescent protein (gfp), and gfp expression was characterized. the inducible system shows a tightly controlled, reversible, and dosage-dependent response to the glucocorticoid dexamethasone in c. roseus hairy roots. full induction was noted after 12-18 h in the mature regions of the root tips and after 6 h in the meristem tissue. upon removal of th ...201312467449
transcription factors: tools to engineer the production of pharmacologically active plant metabolites.plants produce a variety of secondary metabolites, some of which are used as pharmaceuticals or are health promoting as food components. recent genetic studies on the flavonoid biosynthetic pathway show that transcription factors are efficient new molecular tools for plant metabolic engineering to increase the production of valuable compounds. the use of specific transcription factors would avoid the time-consuming step of acquiring knowledge about all enzymatic steps of a poorly characterized b ...200212457774
an early c-22 oxidation branch in the brassinosteroid biosynthetic pathway.the natural occurrence of 22-hydroxylated steroids in cultured catharanthus roseus cells and in arabidopsis seedlings was investigated. using full-scan gas chromatography-mass spectrometry analysis, (22s)-22-hydroxycampesterol (22-ohcr), (22s,24r)-22-hydroxyergost-4-en-3-one (22-oh-4-en-3-one), (22s,24r)-22-hydroxy-5alpha-ergostan-3-one (22-oh-3-one), 6-deoxocathasterone (6-deoxoct), 3-epi-6-deoxoct, 28-nor-22-ohcr, 28-nor-22-oh-4-en-3-one, 28-nor-22-oh-3-one, 28-nor-6-deoxoct, and 3-epi-28-nor- ...200212376657
continuous selective extraction of secondary metabolites from catharanthus roseus hairy roots with silicon oil in a two-liquid-phase bioreactor.a two-liquid-phase bioreactor was designed to extract indole alkaloids from catharanthus roseus hairy roots with silicon oil. partition studies between silicon oil and culture medium showed that the silicon oil did not alter the availability of nutrients. the affinity of tabersonine and löchnericine for silicon oil is nine times higher than for the aqueous phase. cultures were elicited with 25 mg/l of jasmonic acid. the growth of the hairy roots was not significantly modified by the presence of ...201512363351
vindoline biosynthesis is transcriptionally blocked in catharanthus roseus cell suspension cultures.catharanthus roseus cell cultures were exposed to different conditions in order to induce alkaloid metabolism. the exposure to jasmonate and fungal elicitors resulted in the transcriptional activation of tryptophan decarboxylase and in the accumulation of the monoterpenoid indole alkaloids ajmalicine and catharanthine, but not of vindoline. the inability of the cell cultures to produce vindoline was related to a lack of expression of the desacetoxyvindoline 4-hydroxylase (d4h) gene. southern blo ...200212353909
supercritical fluid extraction and liquid chromatography-electrospray mass analysis of vinblastine from catharanthus roseus.supercritical fluid extraction using carbon dioxide modified with methanol, methanol-diethylamine, or methanol-triethylamine was used to extract vinblastine from the aerial portions of catharanthus roseus. an hplc-electrospray ionization (esi)/ms analysis method was also developed to quantify the alkaloids in these extracts. of the supercritical solvents evaluated, carbon dioxide-methanol-triethylamine (80 : 18 : 2) at 80 degrees c and 34.0 mpa greatly improved the supercritical fluid extraction ...200212237558
the effect of ajmalicine spiking and resin addition timing on the production of indole alkaloids from catharanthus roseus cell cultures.the potential for the feedback inhibition of indole alkaloid synthesis was investigated by spiking suspension cultures of catharanthus roseus with 0, 9, or 18 mg/l ajmalicine on day 0. the production of ajmalicine, catharanthine, and serpentine were inhibited in a dose-dependent manner. the inhibition was transient as the exogenous ajmalicine was ultimately either metabolized in the medium or within the cell. the addition of neutral resin has previously been shown to enhance ajmalicine productio ...200212115404
tryptophan decarboxylase from transformed roots of catharanthus roseus.tryptophan decarboxylase (tdc, ec from catharanthus roseus hairy roots was purified 80-fold. antibodies against tdc were obtained and they recognized only one protein of 55 kda in crude extracts from hairy root cultures. elicitation of transformed root cultures with macerozyme yielded a marked increase in tdc activity, which was accompanied by a similar increase in the amount of immunoreactive tdc protein. these results suggest that the alkaloid accumulation, produced by elicitation, r ...200212102544
screening of catharanthus roseus secondary metabolites by high-performance liquid chromatography.two direct hplc analytical methods for the screening of the major indole alkaloids of catharanthus roseus hairy roots and their iridoid precursors have been developed. photodiode array and fluorescence detection were performed. the separation was achieved on a reversed-phase c18 column. the first method allowed the separation of catharanthine, serpentine, tabersonine, vindoline, vinblastine, and vincristine in 20 min. ajmalicine, tryptophan, tryptamine and secologanine were separated using the s ...200212061566
effect of precursor feeding on alkaloid accumulation by a tryptophan decarboxylase over-expressing transgenic cell line t22 of catharanthus obtain more insight into the regulation of terpenoid indole alkaloid (tia) biosynthesis in catharanthus roseus (l.) g. don cell cultures and particularly to identify possible rate limiting steps, a transgenic cell line over-expressing tryptophan decarboxylase (tdc), and thus having a high level of tryptamine, was fed with various amounts of precursors (tryptophan, tryptamine, loganin and secologanin) in different time schedules and analyzed for tia production. when these precursors were added ...200212039535
inheritance of flower color in periwinkle: orange-red corolla and white eye.the commonly found flower colors in periwinkle (catharanthus roseus)--pink, white, red-eyed, and pale pink center--are reported to be governed by the epistatic interaction between four genes--a, r, w, and i. the mode of inheritance of an uncommon flower color, orange-red corolla and white eye, was studied by crossing an accession possessing this corolla color with a white flowered variety (nirmal). the phenotype of the f(1) plants and segregation data of f(2) and backcross generations suggested ...200212011178
t-dna activation tagging as a tool to isolate regulators of a metabolic pathway from a genetically non-tractable plant species.t-dna activation tagging is a method used to generate dominant mutations in plants or plant cells by the insertion of a t-dna which carries constitutive enhancer elements that can cause transcriptional activation of flanking plant genes. we applied this approach to the species catharanthus roseus (l.) g. don (madagascar periwinkle), in an attempt to isolate regulators of genes that are involved in the biosynthesis of secondary metabolites of the terpenoid indole alkaloid (tia) class. several tia ...200111817539
predicting the substrates of cloned plant o-methyltransferases.plant o-methyltransferases (omts) have important roles in secondary metabolite biosynthesis. sequencing projects and homology-based cloning strategies yield sequences for proteins with similarities to known omts, but the identification of the physiological substrates is not trivial. we investigated with a cdna cloned from catharanthus roseus the possibilities for predicting the substrates of omts, using the information from previous work and two newly identified motifs that were based on informa ...200211754938
deracemization of racemic 4-pyridyl-1-ethanol by catharanthus roseus cell cultures.immobilized cells of catharanthus roseus synthesized (r)-4-pyridyl-1-ethanol from the corresponding racemate in 100% yield by stereoinversion of the (s)-alcohol in the racemate to the (r)-alcohol. furthermore, in the reduction of 4-acetylpyridine, we could synthesize both (r) and (s)-4-pyridyl-1-ethanol during a short or a long incubation.199811711075
symmetry c18 column: a better choice for the analysis of indole alkaloids of catharanthus virtue of the different elution patterns of vindoline, catharanthine, vincristine and vinblastine present in the leaves of catharanthus roseus plants, a symmetry c18 column provided a better resolution for all of these compounds as compared with other c18 columns. the binary gradient system with a linear gradient profile, employed in a method developed using a symmetry c18 column, gave excellent peak symmetry, resolution and reproducibility. detection was performed at 220 nm which presented b ...200611705028
a new structural class of bisindole alkaloids from the seeds of catharanthus roseus: vingramine and methylvingramine.two new bisindole alkaloids, vingramine (1) and methylvingramine (2), were isolated from the seeds of catharanthus roseus, apocynaceae. the structures were determined by hrfabms as well as one- and two-dimensional nmr experiments. they possess a new bisindole skeleton involving an indole alkaloid part b with loss of 5',6'-ethylene, a c7'-c16' linkage, a 14'-o-19'-tetrahydrofuran, and a n-4'-isobutyramide group. the 12-methyl vincorine part a and part b are connected via an 11,10'-biphenyl linkag ...199811672355
assay of 2,3-dihydroxybenzoic acid and related compounds in plant materials by high-performance liquid chromatography.salicylic acid and its putative biosynthetic precursors were assayed isocratically by rp-hplc with uv detection at 280 nm. optimum resolution was provided by an hplc mobile phase consisting of meoh-1% aqueous hoac (40:60, v/v), at ph 4. furthermore, for the analysis of 2,3-dihydroxybenzoic acid (2,3-dhba) in catharanthus roseus cell cultures after elicitation, a mobile phase consisting of acetonitrile-1% aqueous hcooh containing 0.25% trichloroacetic acid (1:5, v/v), at ph 2, was used. the recov ...200111572396
aryltetralin lignans: chemistry, pharmacology and biotransformations.podophyllotoxin derivatives like etoposide 7a, etophos 7b, and teniposide 7c are used clinically as potent chemotherapeutic agents for a variety of tumors including small cell lung carcinoma, testicular cancer, and malignant lymphoma. these compounds derived from a series of modifications which converted podophyllotoxin 1a from an entity that interacted with tubulin and blocks mitosis to one that induced a block in late s or early g2 by interacting with topoisomerase ii. synthetic studies on pod ...200111562272
effects of stress factors, bioregulators, and synthetic precursors on indole alkaloid production in compact callus clusters cultures of catharanthus roseus.compact callus cluster (ccc) cultures established from catharanthus roseus consist of cohesive callus aggregates displaying certain levels of cellular or tissue differentiation. ccc cultures synthesize about two-fold more indole alkaloids than normal dispersed-cell cultures. our studies here show that additions of kcl, mannitol, and a variety of synthetic precursors and bioregulators to the ccc cultures markedly improved indole alkaloid production and release of these alkaloids into the medium. ...200111525616
studies on plant-mediated fate of the explosives rdx and hmx.the fate of the explosives rdx and hmx on exposure to plants was investigated in 'natural' aquatic systems of myriophyllum aquaticum for 16 days, and in axenic hairy root cultures of catharanthus roseus for > or = 9 weeks. exposure levels were: hmx, 5 mg/l; and rdx, approximately 8 mg/l. exposure outcomes observed include: hmx, no transformation by aquatic plants, and minimal biological activity by axenic roots; and rdx, removal by both plant systems. in the case of rdx exposure to axenic roots, ...200111513416
herbal medicine in oncology.over the years cancer has been more complicated by 'wonder cures' than perhaps any other condition. remedies such as laetril, shark cartilage, high dose vitamins and many alternative medical practices have been used with little evidence of improvement and sometimes worsened outcome. it is disconcerting that attempts to scientifically discredit these treatments sometimes result in the development of conspiracy theories. however, some plants do contain anticancer agents, the vinca alkaloids (deriv ...200111458587
molecular characterization of the salutaridinol 7-o-acetyltransferase involved in morphine biosynthesis in opium poppy papaver somniferum.salutaridinol 7-o-acetyltransferase (ec ) catalyzes the conversion of the phenanthrene alkaloid salutaridinol to salutaridinol-7-o-acetate, the immediate precursor of thebaine along the morphine biosynthetic pathway. we have isolated a cdna clone that corresponds to the internal amino acid sequences of the native enzyme purified from a cell suspension culture of opium poppy papaver somniferum. the recombinant enzyme acetylated the 7-hydroxyl moiety of salutaridinol in the presence of acetyl-coa. ...200111404355
catharanthus roseus g-box binding factors 1 and 2 act as repressors of strictosidine synthase gene expression in cell cultures.the enzyme encoded by the strictosidine synthase (str) gene catalyses a key step in the biosynthesis of therapeutically valuable terpenoid indole alkaloids. in catharanthus roseus the str gene was shown to be regulated by a wide variety of signals including auxin, methyl jasmonate and fungal elicitors in cell suspension cultures and by tissue-specific control in plant organs. the str promoter contains a functional g-box (cacgtg) cis-regulatory sequence. in order to understand better the mechanis ...200111352466
phospholipase c activity from catharanthus roseus transformed roots: aluminum effect.the effect of aluminum on the activity of plc was examined in transformed roots from catharanthus roseus (l) g. don. when added in vitro to the reaction mixture, al inhibited the enzymatic activity in a concentration and time-dependent fashion. this effect is very similar for both activities (soluble and membrane-associated). when roots were treated in vivo with al 0.1 mm for short periods (0-4 h), plc activity was also inhibited. aluminum (1 mm) diminished root growth in approximately 50% when ...200111352226
enhanced catharanthine production in catharanthus roseus cell cultures by combined elicitor treatment in shake flasks and bioreactors.chemical and fungal elicitors were added to catharanthus roseus cell suspension cultures so as to improve the production of indole alkaloids. a synergistic effect on alkaloid accumulation was observed in c. roseus cell cultures when treated with some combined elicitors of fungal preparations and chemicals. among them, the combination of tetramethyl amminium bromide and aspergillum niger mycelial homogenate gave the highest ajmalicine yield (63 mg l(-1)) and an improved catharanthine accumulation ...200111339952
selection of fungal elicitors to increase indole alkaloid accumulation in catharanthus roseus suspension cell culture.various fungal elicitors derived from 12 fungi were tested to improve indole alkaloid production in catharanthus roseus cell suspension cultures. results show that different fungal mycelium homogenates stimulate different kinds of indole alkaloid (ajmalicine, serpentine and catharanthine) accumulation, which ranged from 2- to 5-fold higher than the control. some fungal culture filtrates also efficiently elicited the biosynthesis of different indole alkaloids. the optimal elicitor addition and ex ...200111339951
pleiotropic morphological and abiotic stress resistance phenotypes of the hyper-abscisic acid producing abo- mutant in the periwinkle catharanthus roseus.the pleiotropic properties of a abo abo (abo-) gamma-ray induced mutant of catharanthus roseus cv. nirmal, selected among the m2 generation seeds for ability to germinate at 45 degrees c, are described. the mutant produced seeds possessing tricotyledonous embryos, unlike the typically dicotyledonous embryos present in the wild type abo+ seeds. in comparison to abo+ adults, the mutant plants had short stature and lanceolate leaves. the vascular bundles in the leaves and stem were poorly developed ...200111255514
monitoring the production yields of vincristine and vinblastine in catharanthus roseus from somatic embryogenesis. semiquantitative determination by flow-injection electrospray ionization mass spectrometry.a semiquantitative determination of two bis-indole antitumor alkaloids, vincristine and vinblastine, has been performed by flow-injection electrospray ionization mass spectrometric analysis of the extracts of catharanthus roseus. leaves and flowers of two different phenotypes (pink flower and white flower) obtained from somatic embryogenesis were thus examined and compared with the field-grown mother plant. different amounts of vincristine and vinblastine were detected depending on the examined ...200111241768
light activation of vindoline biosynthesis does not require cytomorphogenesis in catharanthus roseus seedlings.upon illumination, the cotyledons of catharanthus roseus seedlings readily synthesise vindoline from late biosynthetic intermediates, which accumulate in etiolated seedlings. the cellular localisation of tryptophan decarboxylase (tdc) and desacetoxyvindoline 4-hydroxylase (d4h), which catalyse the first and penultimate reactions of vindoline biosynthesis, was identified by immunocytochemistry in developing seedlings. the expression of tdc was restricted to the upper epidermis of cotyledons, wher ...200011130662
phosphate uptake across the tonoplast of intact vacuoles isolated from suspension-cultured cells of catharanthus roseus (l.) g. don.transport of inorganic orthophosphate (pi) across the tonoplast membrane was studied using intact vacuoles isolated from suspension-cultured cells of catharanthus roseus. orthophosphate uptake was strongly stimulated in the presence of mg-atp and mg-pyrophosphate and inhibited by bafilomycin and concanamycin which are potent inhibitors of the vacuolar h+-atpase. these results indicated that the build-up of an electrochemical gradient by the h - pumps was essential for the uptake of pi. potassium ...200010987558
in vivo 31p and multilabel 13c nmr measurements for evaluation of plant metabolic pathways.reliable measurements of intracellular metabolites are useful for effective plant metabolic engineering. this study explored the application of in situ 31p and 13c nmr spectroscopy for long-term measurements of intracellular ph and concentrations of several metabolites in glycolysis, glucan synthesis, and central carbon metabolic pathways in plant tissues. an nmr perfusion reactor system was designed to allow catharanthus roseus hairy root cultures to grow for 3-6 weeks, during which time nmr sp ...199910935751
orca3, a jasmonate-responsive transcriptional regulator of plant primary and secondary metabolism.biosynthesis of many classes of secondary metabolites in plants is induced by the stress hormone jasmonate. the gene for orca3, a jasmonate-responsive apetala2 (ap2)-domain transcription factor from catharanthus roseus, was isolated by transferred dna activation tagging. orca3 overexpression resulted in enhanced expression of several metabolite biosynthetic genes and, consequently, in increased accumulation of terpenoid indole alkaloids. regulation of metabolite biosynthetic genes by jasmonate-r ...200010894776
purification and characterization of mevalonate kinase from suspension-cultured cells of catharanthus roseus (l.) g. don.mevalonate kinase was purified to homogeneity from catharanthus roseus (l.) g. don suspension-cultured cells. the purified enzyme had an m(r) of 104,600 and a subunit size of about 41,500. kinetic studies indicated an ordered sequential mechanism of action, in which mevalonate was the first substrate to bind and adp was the last product to leave the enzyme. true values for the kinetic constants were determined for mevalonate, with k(ma) = 76 microm and k(ia) = 74 microm, and for atp, with k(mb) ...200010860546
regulation of cytoplasmic ph under extreme acid conditions in suspension cultured cells of catharanthus roseus: a possible role of inorganic phosphate.changes in cytoplasmic ph of suspension-cultured cells of catharanthus roseus under extreme acid conditions were measured with the ph-dependent fluorescence dye; 2',7'-bis-(2-carboxyethyl)-5 (and-6) carboxyfluorescein (-acetoxymethylester) (bcecf). when cells were treated with 1 mm hcl (ph 3 solution), the cytoplasmic ph first decreased then returned to the original level. treatment with 10 mm hcl (ph 2 solution) acidified the cytoplasm to a greater extent, and the acidification continued at a c ...200010845455
determination of metabolic rate-limitations by precursor feeding in catharanthus roseus hairy root cultures.precursors from the terpenoid and tryptophan branches were fed to catharanthus roseus to determine which of the two branches limits metabolic flux to indole alkaloids. the feeding of tryptophan at 17 days of the culture cycle produced auxin-like effects. addition of low levels of auxin or tryptophan resulted in significant increases in flux to the indole alkaloids. conversely, feeding higher levels of auxin or tryptophan resulted in increased branching and thickening of the hairy root cultures. ...200010812182
effects of buffered media upon growth and alkaloid production of catharanthus roseus hairy roots.the influence of buffered media upon the growth and alkaloid productivity of catharanthus roseus hairy root culture was examined. as expected, the buffers minimized shifts in the ph of the media and had slightly negative effects upon growth. the growth of the hairy roots remained optimal in unbuffered media. the specific yield of lochnericine was significantly lower in response to the addition of buffers, while tabersonine was significantly higher. in contrast, the specific yields of ajmalicine, ...200010772463
biosynthesis of brassinosteroids in cultured cells of catharanthus roseus.precursor administration experiments with 2h-labeled 6-oxocampestanol, 6-deoxocastasterone and 6alpha-hydroxycastasterone in cultured cells of catharanthus roseus were performed and the metabolites were analyzed by gc-ms. [2h6]cathasterone was identified as a metabolite of [2h6]6-oxocampestanol, whereas [2h6]6alpha-hydroxycastasterone and [2h6]castasterone were identified as metabolites of [2h6]6-deoxocastasterone, and [2h6]castasterone was identified as a metabolite of [2h6]6alpha-hydroxycastas ...200010724179
the effect of inoculum density and conditioned medium on the production of ajmalicine and catharanthine from immobilized catharanthus roseus cells.the effect of the cell-inoculum size and the addition of conditioned medium on ajmalicine and catharanthine production were studied using immobilized catharanthus roseus cells. higher specific-uptake rates of ammonium, nitrate, and sugars were observed in the low-inoculum-density cultures (50 g fw/l) compared to the high-inoculum-density cultures (100 g fw/l). alkaloid production was not correlated with the exhaustion of a particular nutrient from the medium. the high-inoculum-density cultures p ...200010581436
a simple structured model for maintenance, biomass formation, and ajmalicine production by nondividing catharanthus roseus cells.the stoichiometry of maintenance and carbohydrate storage as well as ajmalicine production kinetics of non-dividing catharanthus roseus cells in the second stage of a two-stage batch process were investigated. for the mathematical description of these processes, a simple structured model with 5 parameters is proposed. in the model the biomass is divided in two compartments: active biomass and storage carbohydrates. in induction medium (standard medium without phosphate, nitrogen and hormones), b ...199910577468
isolation of indole alkaloids from catharanthus roseus by centrifugal partition chromatography in the ph-zone refining mode.centrifugal partition chromatography (cpc) in the ph-zone refining mode allowed a preparative and efficient isolation of vindoline, vindolinine, catharanthine and vincaleukoblastine from a crude mixture of catharanthus roseus alkaloids. the separation protocol was tested with a synthetic mixture of vindoline, catharanthine and vincaleukoblastine. the fraction content was analyzed and the results compared with theoretical chromatograms obtained by numerical simulation. the increase in injected sa ...199910457440
purification of xyloglucan endotransglycosylase based on affinity sorption of the active glycosyl-enzyme intermediate complex to cellulose.xyloglucan endotransglycosylase (xet) catalyzes the cleavage of xyloglucan (xg) molecules by a transglycosylation mechanism involving two steps: (a) endocleavage of the beta-(1,4)-linked polyglucosyl backbone of the xyloglucan molecule with formation of a glycosyl-enzyme intermediate; (b) transfer of the glycosyl residue from the intermediate to the c-4 position of the nonreducing end glucosyl unit of another molecule of xg or an xg-derived oligosaccharide with liberation of the enzyme (z. sulov ...199910419819
arabidopsis det2 is defective in the conversion of (24r)-24-methylcholest-4-en-3-one to (24r)-24-methyl-5alpha-cholestan-3-one in brassinosteroid biosynthesis.previously, we have shown that the arabidopsis det2 (deetiolated2) mutant is defective in the biosynthesis of brassinosteroids (br) and that det2 (a steroid 5alpha-reductase) acts early in the proposed br biosynthetic pathway. in this paper we present further biochemical characterization of det2. we have undertaken metabolic experiments with 2h-labeled substrates of intermediates involved in the formation of campestanol from campesterol, and quantitative analysis of intermediates in arabidopsis ...199910398719
multicellular compartmentation of catharanthus roseus alkaloid biosynthesis predicts intercellular translocation of a pathway intermediatein situ rna hybridization and immunocytochemistry were used to establish the cellular distribution of monoterpenoid indole alkaloid biosynthesis in madagascar periwinkle (catharanthus roseus). tryptophan decarboxylase (tdc) and strictosidine synthase (str1), which are involved in the biosynthesis of the central intermediate strictosidine, and desacetoxyvindoline 4-hydroxylase (d4h) and deacetylvindoline 4-o-acetyltransferase (dat), which are involved in the terminal steps of vindoline biosynthes ...199910330473
microplate enzyme-coupled assays of mevalonate and phosphomevalonate kinase from catharanthus roseus suspension cultured cells.a microplate assay for mevalonate and 5-phosphomevalonate kinase activities has been developed using an enzyme-coupled system of pyruvate kinase and lactate dehydrogenase. mevalonate and 5-phosphomevalonate kinase activities were measured in crude and partially purified enzyme preparations from catharanthus roseus suspension-cultured plant cells. the assay was validated with respect to protein and substrate concentration. mevalonate and 5-phosphomevalonate kinase showed michaelis-menten kinetics ...199910221996
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
alkaloid accumulation in catharanthus roseus suspension cultures. 199910081005
dimethylallyl pyrophosphate is not the committed precursor of isopentenyl pyrophosphate during terpenoid biosynthesis from 1-deoxyxylulose in higher plants.cell cultures of catharanthus roseus were supplied with [2-13c, 3-2h]-deoxyxylulose or [2-13c,4-2h]1-deoxyxylulose. lutein and chlorophylls were isolated from the cell mass, and hydrolysis of the chlorophyll mixtures afforded phytol. isotope labeling patterns of phytol and lutein were determined by 2h nmr and 1h,2h-decoupled 13c nmr. from the data it must be concluded that the deuterium atom in position 3 of deoxyxylulose was incorporated into both isopentenyl pyrophosphate (ipp) and dimethylall ...19999990020
phytohormone regulation of isoperoxidases in catharanthus roseus suspension cultures.peroxidase (pod) activity was investigated in catharanthus roseus cell suspensions cultured under different hormonal conditions. depletion of 2,4-dichlorophenoxyacetic acid (2,4-d) from the culture medium enhanced pod activity in cells and spent medium. addition of phytohormones, in particular the auxin 2,4-d, reduced pod activity in medium and cellular compartments and enhanced ionically cell-wall bound pod. the differential modulation of pod is due to hormone effects on synthesis and/or accumu ...19989842727
jasmonate modulates development- and light-regulated alkaloid biosynthesis in catharanthus roseus.methyl jasmonate, a chemical inducer of secondary metabolism, has been shown to promote vindoline biosynthesis in developing seedlings, as a result of induction of tryptophan decarboxylase (tdc) and desacetylvindoline 4-hydroxylase (d4h). the present studies suggest that jasmonate-based induction of tdc and d4h activities involves modulation of transcriptional, post-transcriptional and post-translational controls. the effects of jasmonate on both enzymes were transient with maximum tdc activity ...19989747537
the iridoid glucoside secologanin is derived from the novel triose phosphate/pyruvate pathway in a catharanthus roseus cell culture.secologanin is the iridoid building block of the majority of the terpenoid indole alkaloids. in the biosynthesis of secologanin, mevalonate was considered to be the exclusive precursor of isopentenyl diphosphate. after [1-(13)c]glucose feeding to a cell culture of catharanthus roseus, its incorporation into secologanin was studied by 13c nmr spectroscopy. the data showed that the novel triose phosphate/pyruvate and not the mevalonate pathway was the major route for the biosynthesis of secologani ...19989742965
developmental and light regulation of desacetoxyvindoline 4-hydroxylase in catharanthus roseus (l.) g. don. . evidence of a multilevel regulatory mechanismthe expression of desacetoxyvindoline 4-hydroxylase (d4h), which catalyzes the second to the last reaction in vindoline biosynthesis in catharanthus roseus, appears to be under complex, multilevel developmental and light regulation. developmental studies with etiolated and light-treated seedlings suggested that although light had variable effects on the levels of d4h transcripts, those of d4h protein and enzyme activity could be increased, depending on seedling development, up to 9- and 8-fold, ...19989701591
purification and characterization of alpha-3',4'-anhydrovinblastine synthase (peroxidase-like) from catharanthus roseus (l.) g. h2o2-dependent enzyme capable of coupling catharanthine and vindoline into alpha-3',4'-anhydrovinblastine (avlb) was purified to apparent homogeneity from catharanthus roseus leaves. the enzyme shows a specific avlb synthase activity of 1.8 nkat/mg, and a molecular weight of 45.40 kda (sds-page). in addition to avlb synthase activity, the purified enzyme shows peroxidase activity, and the vis spectrum of the protein presents maxima at 404, 501 and 633 nm, indicating that it is a high spin fer ...19989654153
effect of elicitor dosage and exposure time on biosynthesis of indole alkaloids by catharanthus roseus hairy root cultures.late exponential phase hairy root cultures of catharanthus roseus were elicited with pectinase and jasmonic acid. the effects of elicitor concentration and exposure time on growth and levels of several compounds in the indole alkaloid biosynthetic pathway were monitored. pectinase decreased the fresh weight to dry weight ratio of the roots, while addition of jasmonic acid had no significant effect. selective effects on indole alkaloid yields were observed upon addition of elicitors. an increase ...20129622525
[new antitumor derivatives of vinblastine].the alkaloids of catharanthus roseus vincristine 1 and vinblastine 2 are widely used in the chemotherapy of cancer. their nitro-derivatives have also antitumour activity [2]. the dinitro-derivatives 4, 6-9 were prepared for pharmacological investigation. some new hydroxymethyl derivatives 12-14, 17-18, 23-24 of the antitumour indole-indoline alkaloids vinblastine 2 and leurosine 5 were synthetised in two different ways a) by the selective reduction of 2, 5 or deacetoxy-vinblastine 16, b) by the ...19989592933
influence of precursor availability on alkaloid accumulation by transgenic cell line of catharanthus roseuswe have used a transgenic cell line of catharanthus roseus (l.) g. don to study the relative importance of the supply of biosynthetic precursors for the synthesis of terpenoid indole alkaloids. line s10 carries a recombinant, constitutively overexpressed version of the endogenous strictosidine synthase (str) gene. various concentrations and combinations of the substrate tryptamine and of loganin, the immediate precursor of secologanin, were added to suspension cultures of s10. our results indica ...19989490777
expression of extensin genes is dependent on the stage of the cell cycle and cell proliferation in suspension-cultured catharanthus roseus isolate cdnas expressed at a specific phase of the cell cycle in a higher plant, we performed differential screening of a cdna library prepared from the s-phase cells of synchronized cultures of catharanthus roseus. sequence analysis shows that two of the identified cdnas, cyc15 and cyc17, encode extensins that represent a family of cell wall hydroxyproline-rich glycoproteins. protein sequences deduced from the two cdnas contain the characteristic pentapeptide repeat sequence, ser-pro-pro-pro ...19989484475
direct fluorometry of phase-extracted tryptamine-based fast quantitative assay of l-tryptophan decarboxylase from catharanthus roseus assay for the enzyme l-tryptophan decarboxylase (tdc; ec is described. it is based on direct fluorometry of the enzymatic reaction product (tryptamine) selectively recovered in ethyl acetate from the reaction mixture. catalytically formed tryptamine from tryptophan in the incubation mixture is selectively (free from tryptophan) physically separated as ethyl acetate solution under basic (ph > or = 11) conditions and subjected to direct fluorescence measurement in the organic solvent ...19989448840
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 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
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
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
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