Enhancing the biosynthesis of salidroside by biotransformation of p-tyrosol in callus culture of Rhodiola rosea L (original) (raw)
Acta Biologica Cracoviensia Series Botanica, 2000
The study examined tyrosol glucosyltransferase activity and the efficiency of salidroside production in natural and transformed root cultures of Rhodiola kirilowii (Regel) Regel et Maximowicz. Neither enzyme activity nor salidroside accumulation were detected in natural and transformed root cultures maintained in media without tyrosol. To induce TGase activity in biotransformation reactions, tyrosol was added to natural and transformed root cultures on the day of inoculation. The first peak of TGase activity (0.23 U/μg) was detected on day 9 in natural root culture, accompanied by the highest salidroside content (15.79 mg/g d.w.), but TGase activity was highest (0.27 U/μg) on day 15. In transformed root culture, day 18 showed the highest TGase activity (0.15 U/μg), which coincided with the highest salidroside content (2.4 mg/g d.w.). Based on these results, tyrosol was added to the medium on the days of highest previously detected activity of TGase: day 15 for natural root cultures and day 18 for transformed root cultures. This strategy gave significantly higher yields of salidroside than in the cultures supplemented with tyrosol on the day of inoculation. In natural root culture, salidroside production reached 21.89 mg/g d.w., while precursor feeding in transformed root cultures caused a significant increase in salidroside accumulation to 7.55 mg/g d.w. In all treatments, salidroside production was lower in transformed than in natural root cultures.
PLoS ONE, 2013
Tyrosine decarboxylase initializes salidroside biosynthesis. Metabolic characterization of tyrosine decarboxylase gene from Rhodiola crenulata (RcTYDC) revealed that it played an important role in salidroside biosynthesis. Recombinant 53 kDa RcTYDC converted tyrosine into tyramine. RcTYDC gene expression was induced coordinately with the expression of RcUDPGT (the last gene involved in salidroside biosynthesis) in SA/MeJA treatment; the expression of RcTYDC and RcUDPGT was dramatically upregulated by SA, respectively 49 folds and 36 folds compared with control. MeJA also significantly increased the expression of RcTYDC and RcUDPGT in hairy root cultures. The tissue profile of RcTYDC and RcUDPGT was highly similar: highest expression levels found in stems, higher expression levels in leaves than in flowers and roots. The gene expressing levels were consistent with the salidroside accumulation levels. This strongly suggested that RcTYDC played an important role in salidroside biosynthesis in R. crenulata. Finally, RcTYDC was used to engineering salidroside biosynthetic pathway in R. crenulata hairy roots via metabolic engineering strategy of overexpression. All the transgenic lines showed much higher expression levels of RcTYDC than non-transgenic one. The transgenic lines produced tyramine, tyrosol and salidroside at higher levels, which were respectively 3.21-6.84, 1.50-2.19 and 1.27-3.47 folds compared with the corresponding compound in non-transgenic lines. In conclusion, RcTYDC overexpression promoted tyramine biosynthesis that facilitated more metabolic flux flowing toward the downstream pathway and as a result, the intermediate tyrosol was accumulated more that led to the increased production of the end-product salidroside.
2019
Rhodiola imbricata belonging to Crassulaceae family, is a dioecious perennial plant. It is herb indigenous to the TransHimalayan region and exclusively found in Leh-Ladakh valleys of India and border regions of China and Tibet. Various pharmacological activities in R. imbricata viz. hepatoprotective, radioprotective, immunomodulatory etc., are attributed to presence of different phytochemicals such as ptyrosol, salidroside, rosavin and rosin. But the established plant cell cultures of Rhodiola imbricata usually encounter problems of low product yields and high cost, which discourage its commercialization. So, in order to enhance the secondary metabolite content in R. imbricata, we have studied the effect of different abiotic elicitors on marker compound production. Owing to the previous success of different elicitors on other species of Rhodiola, we have performed chemical and physical elicitations on shoot cultures of Rhodiola imbricata. The best shoot growth in elicited cultures w...
Salidroside content in "in vitro" propagated Rhodiola rosea L
The aim of this study was to determinate the quantity of salidroside in one and two year old Rhodiola rosea L. samples, propagated in vitro, in comparison with coeval ones, grown by seeds. The average content of the investigated substance in the rhizomes was 0.64% at the first and 0.61% at the second year and exceeds that of the marker individuals. These results indicate the in vitro propagation as a possible way to receive a plant material of good quality.
Plant Cell Reports, 2007
Salidroside is a novel effective adaptogenic drug extracted from the medicinal plant Rhodiola sachalinensis A. Bor. Because this plant is a rare resource and has low yield, there is great interest in enhancing the production of salidroside. In this study, a putative UDPglucosyltransferase (UGT) cDNA, UGT73B6, was isolated from Rhodiola sachalinensis using a rapid amplification of cDNA ends (RACE) method. The cDNA was 1,598 bp in length encoding 480 deduced amino acid residues with a conserved UDP-glucose-binding domain (PSPG box). Southern blot analysis of genomic DNA indicated that UGT73B6 existed as a single copy gene in the R. sachalinensis genome. Northern blot analysis revealed that transcripts of UGT73B6 were present in roots, calli and stems, but not in leaves. The UGT73B6 under 35S promoter with double-enhancer sequences from CaMV-W and TMV-W fragments was transferred into R. sachalinensis via Agrobacterium tumefaciens. PCR, PCR-Southern and Southern blot analyses confirmed that the UGT73B6 gene had been integrated into the genome of transgenic calli and plants. Northern blot analysis revealed that the UGT73B6 gene had been expressed at the transcriptional level. High performance liquid chromatography (HPLC) analysis indicated that the overexpression of the UGT73B6 gene resulted in an evident increase of salidroside content. These data suggest that the cloned UGT73B6 can regulate the conversion of tyrosol aglycon to salidroside in R. sachalinensis. This is the first cloned glucosyltransferase gene involved in salidroside biosynthesis. Keywords Rhodiola sachalinensis Á Salidroside Á Tyrosol Á UGT Á Overexpression Abbreviations UGT Uridine diphosphate dependent glycosyltransferase 6-BA N6-Benzyladenine NAA a-Naphthalene acetic acid 2,4-D 2,4-Dichlorophenoxyacetic acid Communicated by L. Jouanin.
Proceedings of the Bulgarian Academy of Sciences
Rhodiola rosea L. is a high-altitude medicinal plant with various health-beneficial effects, associated mainly with the phenolic substances salidroside and rosavins (rosavin, rosin and rosarin). The intensive collection of R. rosea rhizomes and roots to be used in folk medicine and pharmacy has led to a decrease of the species natural resources. New opportunities for species conservation and accelerated synthesis of biologically active substances have been provided by various systems for in vitro and ex vitro cultivation. In this study, the salidroside and rosavins content was determined using HPLC analysis in in vitro cultures of R. rosea and further monitored in adapted regenerants grown ex vitro. The targeted secondary metabolites were not detected in the calli and in the roots of 1-month-old regenerants obtained on different nutrient media. The roots/rhizomes of adapted regenerants grown ex vitro in a greenhouse for 1 year and in the mountain for 1, 2 and 3 years produced increa...
Enhanced Biotransformation Capacity of Rhodiola rosea Callus Cultures for Glycosid Production
Plant Cell, Tissue and Organ Culture, 2005
Rhodiola rosea is a promising medicinal plant that produces various glycosides. Recently we developed a successful method for cultivating it in liquid cultures of compact callus aggregates. In a previous study we reported the successful production of the glycosides of R. rosea by biotransformation of cinnamyl alcohol and tyrosol. In the present study we investigated the possibility of further increasing the yields of the biotransformation products by addition of glucose to the culture medium aside from sucrose, which was used earlier as carbon source. Surprisingly, glucose addition doubled the yield of cinnamyl alcohol glycosides. Rosavin was not produced at all when only sucrose was used. When glucose was added the accumulation dynamics of rosin and a recently described derivative glycoside (designed as compound 321) were similar. Both increased during the first days and then remained constant, while other glycoside compounds increased continuously throughout the cultivation. Rosavin reached its maximum concentration after nine days. In contrast to the beneficial effect on cinnamyl alcohol related glycosides the addition of glucose did not affect the accumulation of the tyrosol derivative salidroside.
Salidroside content in Rhodiola rosea L., dynamics and variability
Th e aim of this study was to investigate the dynamics of the salidroside content in the underground parts of Rhodiola rosea L. during two consecutive vegetation periods and its variability. Th e obtained annual trends of the salidroside content expressed similar pattern for both years and for both kinds of the investigated matherials (roots and rhizomes) of each sex. Th e signiffi cant infl uence on the content of salidroside exert the sex, plant part and season.
Study of Biotransformation Compounds in Callusar Culture of Rhodiola Rosea Specie
2015
Rhodiola rosea L. is a well known species of plants, which has been used medicinally for decades, but study of its pharmacological effects and the compounds responsible for it use still continues. We present data about induced accumulation of secondary metabolites and as well the results of biotransformation of cinamic alcohol in callus culture of R. rosea of Carpathian origin under the influence of same stress factors. The presence of secondary metabolites was investigated by HPLC-MS analysis. The obtained results can be used for selection of valuable genotypes and their future cultivation in artificial conditions.
Safety of a Sustainably Produced, Bioengineered, Nature-Identical Salidroside Compound
Nutrients
Bioactive phytochemicals such as salidroside have been studied to understand the beneficial effects of Rhodiola rosea, an herbaceous plant used in traditional medicine to increase energy and treat a variety of health issues. However, Rhodiola plants are often slow-growing, and many are endangered in their native habitats. Thus, there is a need for safe, alternative supplies of key phytochemicals from Rhodiola. The salidroside subject of this safety study is a synthetic biology product from fermentation of a bioengineered E. coli that produces salidroside. Here, we present comprehensive test results that support the safety of salidroside manufactured via a patented sustainable bioengineering manufacturing process. In vitro bacterial reverse mutation assays with the bioengineered salidroside show no mutagenicity in any of the concentrations tested. In vivo toxicity studies in rats show no adverse effects from the salidroside product. Based on the results of these studies, we conclude ...
International Journal of Biological Macromolecules, 2019
Roseroot (Rhodiola rosea L.) is a medicinal plant with adaptogenic properties and several pharmaceutically important metabolites. In this study, a full length cDNA encoding a UDPG gene of roseroot was identified, cloned and characterized. Its ORF (1425 bp) was transformed into E. coli, where the expression of the recombinant enzyme was confirmed. To monitor the enzyme activity, 3 precursors (tyramine, 4-hydroxyphenylpyruvate & tyrosol) of salidroside biosynthesis pathway were added to roseroot callus cultures and samples were harvested after 1, 6, 12, 24, 48 & 96h. Along with the controls (without precursor feeding), each sample was subjected to HPLC and qRT-PCR for phytochemical and relative UDP-glycosyltransferase gene expression analysis, respectively. The HPLC analysis showed that the salidroside content significantly increased; reaching 0.5% of the callus dry weight (26-fold higher than the control) after 96h when 2mM tyrosol was given to the media. The expression of the UDPglycosyltransferase increased significantly being the highest at 12h after the feeding. The effect of tyramine and 4-hydroxyphenylpyruvate was not as pronounced as of tyrosol. Here, we introduce a R. rosea specific UDPG gene and its expression pattern after biotransformation of intermediate precursors in in vitro roseroot callus cultures.
2008
s:135. 29. Krajewska-Patan A, Furmanowa M, Dreger M, Górska-Paukszta M, Łowicka A, Mścisz A, Mielcarek s, Baraniak M, Buchwald W, Mrozikiewicz PM. enhancing the biosynthesis of salidroside by biotransformation of p-tyrosol in callus culture of Rhodiola rosea L. herba Pol 2007; 53:55-64. 30. Krajewska-Patan A, Dreger M, Łowicka A, Górska-Paukszta M, Mścisz A, Mielcarek s, Baraniak M, Buchwald W, Furmanowa M, Mrozikiewicz PM. chemical investigations of biotransformed Rhodiola rosea callus tissue. herba Pol 2007; 53: 77-87. 31. György z, tolonen A, Pakonen M, neubauer P, hohtola A. enhancing the production of cinamyl glucosides in compact callus aggregate cultures of Rhodiola rosea by biotransformation of cinnamyl alcohol. Plant sci 2004, 166:229-36. 32. György z, tolonen A, neubauer P, hohtola A. enhanced biotransformation capacity of Rhodiola rosea callus cultures for glycoside production. Plant cell tiss organ cult 2005; 83:129-35. 33. Mikołajczak P, okulicz-Kozaryn i, Kamińska e, n...
Metabolites
Rhodiola semenovii Boriss. (Regel and Herder) might be a promising replacement for the well-known but endangered Rhodiola rosea L. In this research, the metabolic profile of R. semenovii, including drug-active and stress-resistant components, was studied in the context of source–sink interactions in situ in the dynamics of growth and development. Gas chromatography with mass spectrometric detection and liquid chromatography methods were used. The data obtained allow for assumptions to be made about which secondary metabolites determine the level of stress resistance in R. semenovii at different stages of ontogeny in situ. For the first time, an expansion in the content of salidroside in the above-ground organs, with its maximum value during the period of seed maturation, and a significant decrease in its content in the root were revealed in the dynamics of vegetation. These results allow us to recommend collecting the ground component of R. semenovii for pharmaceutical purposes thro...
Plant Science, 2004
Compact callus aggregates (CCA) of Rhodiola rosea were established. The growth curve of CCA culture was determined. No secondary compounds are produced in callus. Cinnamyl alcohol was added to the media in several concentrations between 0.05 and 5 mM in order to stimulate the production of cinnamyl glycosides. The optimal concentration of the precursor was determined. The consumption of cinnamyl alcohol and the production of cinnamyl glycosides were followed by daily sampling. The chemical analyses of the samples were performed by HPLC/MS. Among the cinnamyl glycosides rosin was produced in high amounts. A three to sixfold rosin content was achieved in the treated calli comparing to the content of wild growing plants. Rosavin was produced in traces only.
Plant Physiology
Edible berries are considered to be among nature's treasure chests as they contain a large number of (poly)phenols with potentially health-promoting properties. However, as berries contain complex (poly)phenol mixtures, it is challenging to associate any interesting pharmacological activity with a single compound. Thus, identification of pharmacologically interesting phenols requires systematic analyses of berry extracts. Here, raspberry (Rubus idaeus, var Prestige) extracts were systematically analyzed to identify bioactive compounds against pathological processes of neurodegenerative diseases. Berry extracts were tested on different Saccharomyces cerevisiae strains expressing disease proteins associated with Alzheimer's, Parkinson's, or Huntington's disease, or amyotrophic lateral sclerosis. After identifying bioactivity against Huntington's disease, the extract was fractionated and the obtained fractions were tested in the yeast model, which revealed that salidroside, a glycosylated phenol, displayed significant bioactivity. Subsequently, a metabolic route to salidroside was reconstructed in S. cerevisiae and Corynebacterium glutamicum. The best-performing S. cerevisiae strain was capable of producing 2.1 mM (640 mg L 21 ) salidroside from Glc in shake flasks, whereas an engineered C. glutamicum strain could efficiently convert the precursor tyrosol to salidroside, accumulating up to 32 mM (9,700 mg L 21 ) salidroside in bioreactor cultivations (yield: 0.81 mol mol 21 ). Targeted yeast assays verified that salidroside produced by both organisms has the same positive effects as salidroside of natural origin.
The Scientific World Journal, 2012
At present, more than 50 000 plant species are used in phytotherapy and medicine. About 2/3 of them are harvested from nature leading to local extinction of many species or degradation of their habitats. Biotechnological methods offer possibilities not only for faster cloning and conservation of the genotype of the plants but for modification of their gene information, regulation, and expression for production of valuable substances in higher amounts or with better properties.Rhodiola roseais an endangered medicinal species with limited distribution. It has outstanding importance for pharmaceutical industry for prevention and cure of cancer, heart and nervous system diseases, and so forth. Despite the great interest in golden root and the wide investigations in the area of phytochemistry, plant biotechnology remained less endeavoured and exploited. The paper presents research on initiation ofin vitrocultures inRhodiola roseaand some otherRhodiola species. Achievements in induction o...
Supercritical Carbon Dioxide Extraction of Salidroside from Rhodiola rosea L var. rosea Root
Journal of the Chinese Chemical Society, 2011
Salidroside from the root of Rhodiola rosea L var. rosea was extracted by supercritical carbon dioxide with and without methanol as modifier. Three parameters, i.e. temperature, pressure and different concentrations of methanol were optimized. Salidroside determinations were carried out using high-performance liquid chromatography (HPLC) with UV-Vis detector. An experimental design of response surface methodology (RSM) was used to map the effect of pressure (at 200, 300 and 400 bar), temperature (at 50, 60 and 70°C) and percentage of methanol modifier (at 80, 90 and 100%) on the extraction yield of the active compound and to determine the optimal conditions for the extraction of salidroside from the root of plant. The results showed that supercritical carbon dioxide failed to extract salidroside from the plant material without a methanol as modifier. The yield obtained after 1.5 h extraction with the rate of modifier 0.4 mL/min and 300 bar, 70°C, and 80 percent of methanol modifier condition was the highest (17.15 mg/g). The optimum conditions were 70°C, 295.49 bar and 80 percent of methanol as modifier with the yield of 16.17 mg/g. In addition, the yield obtained with supercritical fluid extraction (SFE) was compared with the Soxhlet extraction, whose yield was 8.64 mg/g.
PRODUCTION OF OLEANOLIC AND URSOLIC ACIDS BY CALLUS CULTURES OF SALVIA TOMENTOSA MILL.
Callus cultures of Salvia tomentosa Mill. were induced and analyzed for their capacity to produce oleanolic and ursolic acids. The obtained callus lines showed high variability in their biosynthetic potentials. Flow cytometric investigations showed that this variability was due to polyploidization of plant cells in calli. Polyploidization of callus cell seems to be promoted by the growth regulators used for callus induction. One octaploid line (consisting of 8C, 16C and 32C cells) was selected as prospective producer of oleanolic (991.57 µg/g DW) and ursolic (641.85 µg/g DW) acids. To our knowledge, this is the first report for obtaining of triterpenes producing callus culture of S. tomentosa Mill.
International Journal of Pharmacy and Pharmaceutical Sciences
Objective: Jatropha curcas (Euphorbiaceae) is native of Latin America origin and widely distributed throughout tropical and sub tropical regions. The present work was carried out to identify the chemical composition in the presence of various concentration of salicyclic acid by GC-MS analysis. Methods: In vitro callus was treated with 200, 300, 400, 500 μM of salicyclic acid in Murashige and Skoog (MS) medium under in vitro condition. Ethyl acetate extract of control and treated callus was analysed for their chemical constituents using Gas Chromatography-Mass Spectroscopy (GCMS). Results: The major constituents obtained from control and salicylic acid treatments are 1-docosene, 1-octadecene, 1-hexadecene, (E)-3-eicosene, (E)-5-eicosene, 1-hexacene, nonahexacontaoic acid and methyl ester-3-oxocyclohexane carboxylic acid. Salicylic acid treatment altered the chemical content and increased percentage of compounds in all treatments, and also resulted in the production of higher percenta...
Herba Polonica, 2009
S u m m a r y Rhodiola Kirilowii (Regel) Maxim. (Crassulaceae) is a traditional medicinal plant used in North Asia and China, especially in the cardiopulmonary disorders in the hypoxic conditions induced by high altitude. The presented results are the part of the investigations carried out in the Branch of Medicinal Plants of the Institute of Natural Fibres and Medicinal Plants in cooperation with the Department of Biology and Pharmaceutical Botany, Medical University in Warsaw on R. Kirilowii plants and tissue cultures. The aim of recent study was to determine the growth dynamics and active compounds production during the cultivation of callus tissues of R. Kirilowii on solid/liquid media. Tissue cultures of R. Kirilowii shown the ability to produce all the active compounds determined in the roots of plants of Polish origin. It is worth emphasizing, that rosavins, according to known literature, were not detected in roots of plants growing in Asia. The best time for collection the tissues from solid medium was 223 Vol. 55 No 3 2009 Callus tissues of Rhodiola Kirilowii (Regel) Maxim.-dynamic of growth and active compounds production fifth or sixth week of the culture-the tissues were growing dynamically and the contents of the main active compounds was high. The material from suspension should be collected in 12-15 days after inoculation. The obtained results will be applied in future investigations on the use of R. Kirilowii extracts in the experimental hypoxia in rats.