Genetic survey of Rhodiola rosea L. populations from the Swiss Alps based on SSR markers

Rhodiola rosea is a perennial adaptogenic medicinal plant found in the cool climates of the northern hemisphere. This species is highly variable both in morphological and phytochemical traits. The genetic diversity of five populations located in the Swiss Alps was studied with twelve SSR markers. However, only eight markers turned out to be informative in this study. The primer pairs for these eight SSR markers produced 37 fragments. The number of alleles per locus ranged from two to eight. The observed heterozygosity was between 0.09 and 1.0, whereas the expected heterozygosity was between 0.13 and 0.72. The genetic diversity was in the same range for all five populations. Principal coordinate analysis revealed that individuals from different populations did not cluster together, which confirmed that diversity within and among the populations were almost equivalent. The genetic fragmentation of this alpine species despite of its fragmented and isolated habitats, did not happened yet. The results of the present study on the genetic diversity were consistent with an earlier study on the chemical diversity with the same individuals.     

The in vitro and in vivo antiviral effects of salidroside from Rhodiola rosea L. against coxsackievirus B3

The aim of this study was to investigate the antiviral effects of salidroside, a major component of Rhodiola rosea L. First, the antiviral effects of salidroside against coxsackievirus B3 (CVB3) were determined in vitro and in vivo. Then, the effect of salidroside on the mRNA expression of some important cytokines was measured in hearts of infected BALB/c mice by RT-PCR. Salidroside exhibited obvious antiviral effects both in in vitro and in vivo experiments. Salidroside was found to modulate the mRNA expression of interferon-gamma (IFN-gamma), interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-alpha), and interleukin-2 (IL-2). In conclusion, salidroside possesses antiviral activities against CVB3 and it may represent a potential therapeutic agent for viral myocarditis.     

Micropropagation of a medicinal plant, Plantago major L.

An efficient micropropagation protocol was developed for an important medicinal plant, Plantago major L. For this purpose, it is recommended to culture shoot-tips on modified MS medium [412.5 mg dm-3 NH4NO3 and 340 mg dm-3 KH2PO4] supplemented with 50 g dm-3 glucose and 0.5 μM 6-benzylaminopurine. Maximum rooting frequency was obtained at 1 μM naphthaleneacetic acid. This revised version was published online in August 2006 with corrections to the Cover Date.     

A cell-free plant extract for accurate pre-tRNA processing, splicing and modification.

An intron-containing tobacco tRNA(Tyr) precursor synthesized in a HeLa cell nuclear extract has been used to develop a cell-free processing and splicing system from wheat germ. Removal of 5' and 3' flanking sequences, accurate excision of the intervening sequence, ligation of the resulting tRNA halves, addition of the 3'-terminal CCA sequence and modification of seven nucleosides were achieved in appropriate wheat germ S23 and S100 extracts. The maturation of pre-tRNA(Tyr) in these extracts resembles the pathway observed in vivo for tRNA biosynthesis in Xenopus oocytes and yeast in that processing of the flanks precedes intron excision. Most of the modified nucleosides (m2(2) G, psi 35, psi 55, m7G and m1A) are introduced into the intron-containing pre-tRNA with mature ends, whereas two others (m1G and psi 39) are only found in the mature tRNA(Tyr). Processing and splicing proceed very efficiently in the wheat germ extracts, leading to complete maturation of 5' and 3' ends followed by about 65% conversion to mature tRNA(Tyr) under our standard conditions. The activity of the wheat germ endonuclease is stimulated 3-fold by the non-ionic detergent Triton X-100. All previous attempts to demonstrate the presence of a splicing endonuclease in wheat germ had failed (Gegenheimer et al., 1983). Hence, this is the first cell-free plant extract which supports pre-tRNA processing and splicing in vitro.     

A new control strategy for nematodes of sheep using chlamydospores of a fungus,Clonostachys rosea f. rosea,and an ethanolic extract of a plant,Ananas comosus

An experiment was conducted at Ukulinga Research Farm using 24 Merino sheep to test a combination of plant Ananas comosus (Ac) and fungus Clonostachys rosea (Cr) in controlling nematode parasites. Four treatments were assigned: 100 mg kg^?1 body weight (BW) ethanol extract of Ac, 1 g of Cr product kg^?1 BW, the combination of both treatments (AcCr), and a Control. Sheep within a treatment were paired and penned in individual paddocks, and rectal faecal samples were taken for counting of eggs per gram (EPG) of faeces and third-stage (L3) larvae. Grass samples were taken every 21 days for counting of L3 larvae. A. comosus and AcCr reduced (P < 0.001) EPG. A. comosus, C.rosea and AcCr reduced (P < 0.001) L3 counts. The efficacy of treatments increased with time (P < 0.001). C. rosea and AcCr reduced (P < 0.001) L3 on grass. In conclusion, AcCr is better at controlling nematodes than either Ac or Cr individually.     

Diversity of antagonistic bacteria isolated from medicinal plant Peganum harmala L.

The antimicrobial activity of plant extract of Peganum harmala, a medicinal plant has been studied already. However, knowledge about bacterial diversity associated with different parts of host plant antagonistic to different human pathogenic bacteria is limited. In this study, bacteria were isolated from root, leaf and fruit of plant. Among 188 bacterial isolates isolated from different parts of the plant only 24 were found to be active against different pathogenic bacteria i.e. Escherichia coli, Methicillin-resistant Staphylococcus aureus (MRSA), Enterococcus faecium, Enterococcus faecalis and Pseudomonas aeruginosa. These active bacterial isolates were identified on the basis of 16S rRNA gene analysis. Total population of bacteria isolated from plant was high in root, following leaf and fruit. Antagonistic bacteria were also more abundant in root as compared to leaf and fruit. Two isolates (EA5 and EA18) exhibited antagonistic activity against most of the targeted pathogenic bacteria mentioned above. Some isolates showed strong inhibition for one targeted pathogenic bacterium while weak or no inhibition for others. Most of the antagonistic isolates were active against MRSA, following E. faecium, P. aeruginosa, E. coli and E. faecalis. Taken together, our results show that medicinal plants are good source of antagonistic bacteria having inhibitory effect against clinical bacterial pathogens.     

Systems pharmacology unravels the synergic target space and therapeutic potential of Rhodiola rosea L. for non-small cell lung cancer

BackgroundLung cancer is the most common and mortal cancer worldwide. Rhodiola rosea L. (RR), a well-known traditional Chinese medicine (TCM), has been turned out to be effective in anti-lung cancer therapy, but its molecular mechanism of action has not been clearly understood.PurposeIn this study, we aimed to elucidate the possible molecular mechanism underlying the effect of RR against non-small cell lung cancer (NSCLC) by systems pharmacology.MethodsThe effects of RR on NSCLC were examined in Lewis lung carcinoma (LLC) tumor-bearing mice models. The possible molecular mechanism was unraveled by systems pharmacology, which includes pharmacokinetics evaluation, active compounds screening, target prediction and network analysis. Cell proliferation was examined by cell counting kit-8 (CCK-8) assay; cell apoptosis was detected by flow cytometry; protein and proinflammatory cytokines expression were evaluated by Western blot and qRT-PCR.ResultsIn vivo, RR significantly inhibited the tumor growth and prolonged the survival of the tumor bearing mice. In silico, we identified 19 potential active molecules (e.g., salidroside and rhodiosin), 112 targets (e.g., COX-2 and AKT) and 27 pathways (e.g., PI3K/AKT signaling pathway and NF-κB signaling pathway) for RR. Additionally, targets analysis and networks construction further revealed that RR exerted anti-cancer effects by regulating apoptosis, angiogenesis and inflammation. In vitro, salidroside could significantly decrease expression of pro-angiogenic factors (e.g., VEGF and eNOS) and proinflammatory cytokines (e.g., COX-2, iNOS and TNF-α). Also, Bcl-2, an anti-apoptotic protein was decreased whereas Bax, a pro-apoptotic protein, was increased. Further flow cytometry analysis showed that salidroside could induce apoptosis in H1975 cells.ConclusionsMechanistically, the antitumor effect of RR on NSCLC was responsible for the synergy among anti-inflammatory, anti-angiogenic and pro-apoptotic.     

Effect of soil type, salinity, and allelochemicals on germination and seedling growth of a medicinal plant Lepidium sativum L.

The present work was conducted to elucidate the effect of soil type, salinity stress and allelochemicals from alfalfa on the germination efficiency, seedling growth and photosynthetic pigments of Medicago sativa L. The highest germination rate (69.9%) was recorded for seeds cultivated in sandy soil followed by that of clay soil (42%) and then loamy soil (19%). The maximum germination rate was attained at 50 mM NaCl concentration level, after that the values were decreased as the salinity levels increased. The maximum lengths (cm) of plumule and radicle and their growth rates (cm/day) were obtained at control level and found to decline significantly as the salinity increased. Aqueous extracts of alfalfa reduced the germination rate, radicle and plumule growth and seedling dry weight of L. sativum. However, chlorophyll and carotenoid content was increased, their maxima were recorded at 50% concentration level. The chlorophyll a/b ratio attained its maximum at the full strength concentration level.     

Repetitive somatic embryogenesis from leaves of the medicinal plant Petiveria alliacea L.

Leaf segments from in vitro-grown shoot cultures of Petiveria alliacea were incubated on Murashige and Skoog (MS) medium supplemented with different concentrations of zeatin, thidiazuron, 2,4-dichlorophenoxyacetic acid (2,4-D) or picloram (PIC). Direct somatic embryogenesis was induced in response to all tested concentrations of 2,4-D and PIC. Primary somatic embryos displayed highly repetitive embryogenesis, both on the induction medium and in liquid hormone-free MS medium. Plantlets were obtained from these secondary embryos at an estimated frequency of 5 %, after 180 days of culture on half-strength MS medium gelled with 0.2 % Phytagel. Simultaneous development of friable non-embryogenic callus was also observed on media containing PIC or 2,4-D at different concentrations. Cell suspension cultures initiated from these callus tissues did not show an increase in biomass. The embryogenic portions formed at the surface of the explants in response to 20.0 μM PIC were inoculated in hormone-free full-or half-strength liquid MS medium (MS0) and showed high rates of secondary embryogenesis, resulting in the production of a mean of 35 embryos for each embryo inoculated at the culture initiation. Embryos that started the conversion process in the liquid MS0 medium originated whole plants at a frequency of 100 % when transferred to MS0 medium solidified with 0.7 % agar. Acclimatization was achieved in 90 % of the converted plantlets, with the production of phenotypically normal plants. This system is potentially useful for the micropropagation of this species, as well as for the production of substances with pharmacological interest, such as dibenzyl trisulfide.     

A randomized trial of two different doses of a SHR-5 Rhodiola rosea extract versus placebo and control of capacity for mental work.

A randomized, double-blind, placebo-controlled, parallel-group clinical study with an extra non-treatment group was performed to measure the effect of a single dose of standardized SHR-5 Rhodiola rosea extract on capacity for mental work against a background of fatigue and stress. An additional objective was to investigate a possible difference between two doses, one dose being chosen as the standard mean dose in accordance with well-established medicinal use as a psychostimulant/adaptogen, the other dose being 50% higher. Some physiological parameters, e.g. pulse rate, systolic and diastolic blood pressure, were also measured. The study was carried out on a highly uniform population comprising 161 cadets aged from 19 to 21 years. All groups were found to have very similar initial data, with no significant difference with regard to any parameter. The study showed a pronounced antifatigue effect reflected in an antifatigue index defined as a ratio called AFI. The verum groups had AFI mean values of 1.0385 and 1.0195, 2 and 3 capsules respectively, whilst the figure for the placebo group was 0.9046. This was statistically highly significant (p < 0.001) for both doses (verum groups), whilst no significant difference between the two dosage groups was observed. There was a possible trend in favour of the lower dose in the psychometric tests. No such trend was found in the physiological tests.     

Efficient plant regeneration from cell suspension protoplasts of the woody medicinal plant Solanum dulcamara L. (bittersweet,woody nightshade)

Large populations of viable protoplasts were released from suspension cultured cells of the woody medicinal plant Solanum dulcamara (bittersweet, woody nightshade) when the cells were harvested 3 to 7 months after culture initiation and 4 to 5 days after transfer to fresh medium. A Bio-Gel p6 purified enzyme mixture enhanced the protoplast plating efficiency 6 fold compared to the unpurified mixture, without affecting protoplast yield. Agarose-solidified medium markedly improved protoplast division and colony formation, and enabled protoplasts to be plated at lower densities than in liquid medium. All protoplast-derived tissues produced shoots on MS based medium with 1.0 mgl^-1 zeatin. Shoots rooted readily on medium lacking phytohormones. Cytological examination revealed high chromosome stability of suspension cultured cells, of plants derived from such cells, and of protoplast-derived plants. The implication of these results is discussed in relation to the genetic manipulation of this pharmaceutically important plant.     

Zeatin and TDZ-induced Shoot Proliferation and Use of Bioreactor in Clonal Propagation of Medicinal Herb, Roseroot (Rhodiola rosea L)

A procedure for in vitro propagation of roseroots (Rhodiola rosea L), a medicinal plant, was developed using a RITA bioreactor system containing liquid medium, combined with a gelled medium. Wild roseroot clones: ‘RCi’, ‘RC2’ and ‘RC3’ were established on a basal medium (BM) from in vitro-germinated seedlings on half-strength Murashige and Skoog (MS) salts. TDZ at 2–4 μM supported shoot proliferation but inhibited shoot elongation of ‘RCi’ shoots on gelled medium. Clones differed significantly with respect to multiplication rate with ‘RCi’ producing the most shoots per explant on gelled BM with 2 μM zeatin. In a bioreactor system, TDZ supported rapid shoot proliferation at lower concentration (0.5 μM) but induced hyperhydricity at more than 0.5 μM. Bioreactor-multiplied hyperhydric shoots of all clones when transferred to gelled medium containing 1–2 μM zeatin produced normal shoots within 4 wk of culture. Shoots were rooted in vitro on BM void of growth regulators. Almost all (9U to 95%) in vitro plantlets survived when transferred to potting medium.     

Standardization of a protocol for micropropagation of Eupatorium glandulosum L. an important medicinal plant

The presence of residual female fertility in most of the parthenocarpic banana accessions encourages the banana breeder to develop new hybrids through conventional breeding. Desirable trait can be fixed in the first generation of hybrid progenies, but the evaluation of these hybrids in field is the time-consuming process owing to non-availability of uniform suckers/planting material. This can be overcome by developing multiple shoots from single embryo in a short period of time through embryo culture. A protocol for in vitro multiplication of plantlets from zygotic embryos was standardized in seeded accessions. Multiple shoots from zygotic embryos were achieved up to 55.2% and 64.1% in seeded accessions of Musa acuminata and M. velutina respectively in medium supplemented with 17.76 µM of BAP. The Single shoot derived (only germination) from zygotic embryos was decapitated and the apical meristem were disturbed for further multiple shoot formation in media supplemented with 17.76 µM of BAP. Present studies revealed that in total 75% and 91% of the M. acuminata and M.velutina embryos were able to produce multiple shoot from single embryo by manipulating the media composition and decortications technique. The above protocol was applied for zygotic embryos obtained from controlled pollination (18 cross combinations) and open pollination (nine accessions) of various genomic groups (ABB, AAB, AA). The multiple shoots derived from zygotic embryos and plantlet germinated from zygotic embryos was examined for genetic fidelity analysis by SSR markers.

     

Effect of aqueous extract from Nigella sativa L. on guinea pig isolated heart

A potent inhibitory effect of aqueous extract from N. sativa on calcium channel of guinea pig heart was found comparable and even greater than that of diltazem. The results may also indicate an opening effect for the plant on potassium channel of isolated heart.     

In vitro organogenesis and plant regeneration of Thymus serpyllum L.: an important aromatic medicinal plant

An efficient in vitro propagation system has been developed for the rapid micropropagation of Thymus serpyllum L. (Banajwain), an aromatic medicinal herb from nodal explant on MS medium. Phenolic leaching and high rate of contamination was the most significant problem in establishing in vitro culture of Thymus serpyllum which was overcome by preparing explants in an antioxidant ascorbic acid (1000 ppm) at 6°C for 45 min and addition of the same antioxidant (50 mgl⁻¹) to the MS medium. The frequency of shoot production was influenced by different cytokinins (Kn, BAP, and Kn + BAP) and 95.56% shoot induction was observed when MS medium was supplemented with 1.0 + 2.0 mgl⁻¹ (Kn + BAP). The maximum average number of shoots 16.93 ± 2.15 and average length (3.98 ± 0.55) was recorded when MS medium have 0.5 + 2.0 mgl⁻¹ (Kn + BAP). The in vitro regenerated microshoots were rooted on MS and half strength MS medium and there was significant difference in root induction on both media under the influence of auxins (IAA, IBA, and NAA). The maximum average number (11.67 ± 3.03) and average root length (3.88 ± 0.71) was reported in half MS medium having 1.0 mgl⁻¹ IBA. The complete regenerated plantlets were acclimatized under growth chamber before transferring to the earthen pots and showed 90% survival.

Graphical abstract

     

中国淫羊藿属药用植物一新种

记载了产于我国贵州的淫羊藿属（小檗科）药用植物一新种:黔北淫羊藿。     

Revised Methods for Purification of Ribosyl-trans-zeatin from Vinca rosea L. Crown Gall Tumor Tissue

Ribosyl-trans-zeatin has been purified from Vinca rosea L. crown gall tumor tissue by using two new sequences of isolation procedures. Identification of the compound has been established by mass spectrometry, ultraviolet absorbancy spectra, chromatographic values, and growth activity. The isolation sequences eliminate the exposures to pH extremes and the strong cation-exchange resin used in the purification reported earlier. The initial extraction procedures have been designed so as to avoid enzymatic alteration or production of active material and to prevent the inclusion in the extracts of nucleic acids which might serve as soures of the small active compounds. The production of ribosylzeatin by the tumor tissue is confirmed and the validity of isolation steps such as the use of cation exchangers is supported.     

Plant regeneration from mesophyll protoplasts of a medicinal plant, Phellodendron amurense rupr.

Summary A regeneration system from protoplast to plantlet for a medicinal plant species, Phellodendron amurense Rupr., has been developed. Leaves of micropropagated shoots or plantlets were selected as plant materials for protoplast isolation. The yield and viability of leaf protoplasts were greatly influenced by enzyme combination, treatment time and osmoticum. The highest viability (86%) with a yield of 7.1×10⁵ protoplasts per gram fresh weight was obtained with a 6-h digestion in 1% Cellulase Onozuka R-10 plus 1% Driselase-20. Sustained cell division and colony formation from the protoplasts were best supported at a plating density of 4×10⁵−6×10⁵ protoplasts per milliliter using a 0.2% gellan gum-solidified or liquid MS (Murashige and Skoog, 1962) medium containing 0.6M mannitol, 2.0μM 6-benzylaminopurine (BA) with 4.0 μM α-naphthaleneacetic acid (NAA), indole-3-butyric acid (IBA), or 2,4-dichlorophenoxyacetic acid (2,4-D). The protoplast-derived colonies formed green compact calluses when transferred to a solidified MS medium containing 2.0 μM BA with 4.0μM NAA of IBA. Shoot regeneration from protoplast-derived calluses was induced on MS medium supplemented with 2.0 μM BA and 1.0μM NAA or 2.5μM IBA. Shoot multiplication and elongation occurred on MS medium containing 1.0μM BA. In vitro-grown shoots were rooted on MS medium with either 0.5–4.0μM IBA or NAA. Regenerants were transferred to the Kanuma soil and successfully established under greenhouse conditions.