RESUMO
Silver (Ag) is a non-essential heavy metal with substantial environmental toxicity but an excellent promotor for plant organogenesis. It is used as an elicitor for secondary metabolite production and for in planta synthesis of metal nanoparticles (MNPs). In the present study, the Ag accumulation and reduction capability of in vitro shoots of Withania somnifera and the toxicity and elicitation effect of Ag on in vitro shoots were explored. In vitro shoot cultures of W. somnifera were treated with different concentrations of silver nitrate for a specific treatment period. Growth index, withaferin A, elemental and electron microscopy analyses were done on silver-treated in vitro shoots of W. somnifera. 1 mM silver nitrate treatment for 12 days period was found to give increased growth index (1.425 ± 0.05c) and withaferin A (2.568 ± 0.08e mg g-1) content. The concentration of bioaccumulated Ag in 1 mM silver nitrate treated in vitro shoot was found to be 50.8 ppm. The presence of nano-Ag was also found in the leaves of 1 mM silver nitrate-treated in vitro shoots. In summary, this is the first report portraying the bioaccumulation and in planta reduction capability of the in vitro shoot system of W. somnifera, which makes it a potential medicinal plant of commercial value for silver contaminated soils.
Assuntos
Nanopartículas Metálicas , Brotos de Planta , Prata , Withania , Vitanolídeos , Withania/metabolismo , Withania/crescimento & desenvolvimento , Withania/efeitos dos fármacos , Vitanolídeos/metabolismo , Brotos de Planta/efeitos dos fármacos , Brotos de Planta/metabolismo , Brotos de Planta/crescimento & desenvolvimento , Nanopartículas Metálicas/química , Prata/farmacologia , Nitrato de Prata/farmacologiaRESUMO
Withania somnifera (L.) Dunal is an important indigenous medicinal plant with extensive pharmaceutical potential. The root is the main source of major bioactive compounds of this plant species including withanolides, withanine, phenolic acids, etc. Hairy root culture (HRC) is a crucial method for low-cost production of active compounds on a large scale. Four different Agrobacterium rhizogenes strains have been used for the hairy root induction. Maximum transformation efficiency (87.34 ± 2.13%) was achieved with A4 bacterial strain-mediated transformed culture. The genetic transformation was confirmed by using specific primers of seven different genes. Seven HR (Hairy root) lines were selected after screening 29 HR lines based on their fast growth rate and high accumulation of withanolides and phenolic acids content. Two biotic and three abiotic elicitors were applied to the elite root line to trigger more accumulation of withanolides and phenolic acids. While all the elicitors effectively increased withanolides and phenolic acids production, among the five different elicitors, salicylic acid (4.14â¯mgâ¯l-1) induced 11.49 -fold increase in withanolides (89.07 ± 2.75â¯mgâ¯g-1 DW) and 5.34- fold increase in phenolic acids (83.69 ± 3.11â¯mgâ¯g-â¯1 DW) after 5 days of elicitation compared to the non-elicited culture (7.75 ± 0.63â¯mgâ¯g-1 DW of withanolides and 15.66 ± 0.92â¯mgâ¯g-1 DW of phenolic acids). These results suggest that elicitors can tremendously increase the biosynthesis of active compounds in this system; thus, the HRC of W. somnifera is cost-effective and can be efficiently used for the industrial production of withanolides and phenolic acids.
Assuntos
Agrobacterium , Hidroxibenzoatos , Raízes de Plantas , Withania , Vitanolídeos , Withania/metabolismo , Withania/genética , Withania/crescimento & desenvolvimento , Hidroxibenzoatos/metabolismo , Vitanolídeos/metabolismo , Raízes de Plantas/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/genética , Agrobacterium/genética , Agrobacterium/metabolismo , Transformação GenéticaRESUMO
Withania somnifera (L.) Dunal, generally well-known as Ashwagandha, is part of Indian traditional medicinal systems like Ayurveda, Siddha, and Unani for over 3000 years for treating an array of disorders. The chief bioactive component of this plant is the withanolides, a group of C28-steroidal lactone triterpenoids. These compounds are present in very low concentrations and hence cell culture methods have been used to enhance their production. Low-level laser irradiation has been reported to have elicited the seed germination, agronomical characters, biosynthesis of bioactive compounds in some plants. Therefore, the objective of the study was to investigate the effect of red (He-Ne) laser irradiation on seed germination, growth characters, pigment contents and withanolide content in W. somnifera. The seeds were inoculated onto two different combinations of Murashige and Skoog (MS) media and incubated for germination. The highest germination percentage was observed in ½ MS with pH 6.5 and GA3 presoaking followed by ½ MS with different pH. Four different doses of Helium-Neon (He-Ne) laser (10, 15, 20 and 25 J/cm2) were used to irradiate the seeds at 632.8 nm and germinated in vitro on ½ MS with pH 6.5. The maximum germination percentage, 63.88% was noted from seeds irradiated with 25 J/cm2 (P = 0.04). The highest total length of 13.33 cm was observed in the seedlings irradiated with 25 J/cm2 groups (P = 0.008). The highest total chlorophyll content of 329.5 µg/g fresh weight (FW) was observed for seedlings irradiated with 15 J/cm2 (P = 0.02) and the highest carotenoid content of 49.6 µg/g FW was observed for 25 J/cm2 treated seedlings. Further, primary root length was measured and found to be highest (11.14 cm) in seedlings irradiated with 10 J/cm2 and the highest number of lateral roots were observed for 15 and 25 J/cm2 groups. The significant amount of Withanolide A (WA) 0.52 µg/g dry weight (DW) and 0.60 µg/g DW was noted in 15 (P = 0.01) and 20 J/cm2 (P = 0.002) groups, respectively than control. The present investigation thus reveals the positive impact of red laser on the germination of seeds, growth characters and withanolide contents under in vitro environment.
Assuntos
Germinação/efeitos da radiação , Extratos Vegetais/metabolismo , Plântula/efeitos da radiação , Sementes/efeitos da radiação , Withania/efeitos da radiação , Vitanolídeos/metabolismo , Carotenoides/análise , Carotenoides/metabolismo , Técnicas de Cultura de Células , Clorofila/análise , Clorofila/metabolismo , Relação Dose-Resposta à Radiação , Lasers , Extratos Vegetais/efeitos da radiação , Raízes de Plantas/metabolismo , Raízes de Plantas/efeitos da radiação , Doses de Radiação , Plântula/metabolismo , Sementes/metabolismo , Withania/crescimento & desenvolvimento , Vitanolídeos/efeitos da radiaçãoRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Withania somnifera is one of the most extensively delved Ayurvedic medicine. Apart from rejuvenation and increasing longevity, it has several other properties such as immunomodulation, anti-cancer, anti-stress and neuroprotection. Because of its prevailing use and increasing demand, it becomes prudent to scientifically evaluate and document both its propagation and production of desired phytoconstituents. AIM OF THE STUDY: This review aims to highlight the research progress achieved on various biotechnological and tissue culture aspects of Withania somnifera and to cover up-to-date information regarding in-vitro propagation and production of withanolides. MATERIALS AND METHODS: Significant published studies were identified for the years 2000-2018 using Elsevier-Science Direct, Pubmed and Google scholar and several research studies in our laboratory. Following keywords such as "plant extracts", "in vitro cultures", "callus and suspension culture", "micropropagation", "hairy root cultures" were used. Further, "Withania somnifera", "secondary metabolites specially withanolides", "molecular techniques" and "in vitro conservation" were used to cross-reference the keywords. RESULTS: Ashwagandha comprises a broad spectrum of phytochemicals with a wide range of pharmacological properties. W. somnifera seeds have reduced viability and germination rates; thus, its regular cultivation method fails to achieve commercial demands mainly for the production of desired phytoconstituents. Cultivation of plant cells/tissues under in vitro conditions and development of various biotechnological strategies will help to build an attractive alternative to provide adequate quality and quantity raw materials. Recently, a large number of in vitro protocols has developed for W. somnifera not only for its propagation but for the production of secondary metabolites as well. Present work highlights a variety of biotechnological strategies both for prompt propagation and production of different bioactive secondary metabolites. CONCLUSION: The present review focuses on the development and opportunities in various biotechnological approaches to accomplish the global demand of W. somnifera and its secondary metabolites. This review underlines the advances in plant biotechnological approaches for the propagation of W. somnifera and production of its bioactive compounds.
Assuntos
Biotecnologia/métodos , Extratos Vegetais/química , Withania/química , Withania/crescimento & desenvolvimento , Técnicas de Cultura/métodos , Ayurveda , Compostos Fitoquímicos/química , Compostos Fitoquímicos/metabolismo , Metabolismo Secundário , Withania/metabolismo , Vitanolídeos/química , Vitanolídeos/metabolismoRESUMO
KEY MESSAGE: Withania coagulans (L.) Dunal bio-synthesized silver nanoparticles (WcAgNPs) worked as an abiotic elicitor or auto-catalyst that enhanced root regeneration and withanolides production in in-vitro regenerated W. coagulans. Rapid development in the production / consumption of silver nanoparticles (AgNPs) raised serious concern over its effects on the growth of natural plant community. The knowledge related to impact of AgNPs on plant growth and biocompatibility is increasing day by day, but comprehensive mechanism and gaps regarding their impacts on plant health have yet to be addressed. In the present study, we investigated the impact of Withania coagulans biosynthesized AgNPs (WcAgNPs) on in-vitro plant growth and withanolides production. Obtained results showed that the low concentrations of WcAgNPs significantly induced the plant growth by regulating oxidative stress via anti-oxidative defense system. Physiological, morphology and anatomical features also reflected healthy plant growth under low WcAgNPs exposure. While higher concentrations of WcAgNPs have a negative impact on W. coagulans plant growth due to induced lipid peroxidation, ROS accumulation, and root cell death. At lower concentrations, WcAgNPs have shown a positive effect on in-planta withanolides biosynthesis stimulating withanolide A and withaferin A up to 11.15-22.8-fold, respectively. Furthermore, the expression of withanolides biosynthetic genes were also quantified upon WcAgNPs exposure and terpenes biosynthetic genes showed over-expression. Thus, the present study concludes that the lower concentrations of WcAgNPs positively induced plant growth via improved root organogenesis and also have potential to act as an elicitor for withanolides production.
Assuntos
Nanopartículas Metálicas/química , Prata/química , Withania/metabolismo , Vitanolídeos/metabolismo , Morte Celular , Expressão Gênica , Peroxidação de Lipídeos , Estresse Oxidativo , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Withania/genética , Withania/crescimento & desenvolvimentoRESUMO
Among various in vitro plant culture systems, hairy root systems seem to be one of the most appealing methods of recombinant protein production due to their advantages in combining both whole-plant cultivation and suspension cell culture platform. This is a report on production and secretion of a recombinant pharmaceutically active protein from hairy roots cultures of Withania somnifera to improve the economic potential of this plant for the production pharmaceutical compounds. In this study, we selected and synthesized a codon-optimized globular adiponectin (gAd) gene with a calreticulin signal peptide and cloned the sequence into a plant expression binary vector containing a nptII gene as a selectable marker gene. The transgenic hairy roots were produced by Agrobacterium rhizogenes-mediated transformation protocol developed by our group. Among ten established nptII positive hairy roots lines, six colons significantly accumulated gAd protein in the biomass and extracellular medium. The presence of gAd was confirmed by western blot analysis of root extracts. The maximum level of hairy root biomass, growth rate (GR), intra- and extracellular gAd expressions were obtained after 25-26 days of culture on MS medium. The maximum level of intra- and extracellular gAd proteins were found to be 15.19 µg/gFW and 215.7 µg/L, respectively, which resulted in a significant decrease in the amount of intra- and extracellular withanolide A and withaferin A production. The addition of PVP, KNO3 and NaCl significantly increased the level of extracellular gAd by approximately 13 folds. This improvement could significantly increase the amount of intra- and extracellular withanolide A and withaferin A production, too. The recombinant gAd produced from W. somnifera is functional as proved by induction the phosphorylation of ACC in C2C12 muscle cells, as its functional amount was 5.1-fold more than gAd produced from E. coli and 45 % lower than CHO cells.
Assuntos
Adiponectina/genética , Adiponectina/metabolismo , Preparações Farmacêuticas/metabolismo , Raízes de Plantas/metabolismo , Withania/genética , Withania/metabolismo , Agrobacterium , Animais , Sistemas de Secreção Bacterianos , Biomassa , Cricetulus , Meios de Cultura/metabolismo , Escherichia coli/genética , Dosagem de Genes , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas , Withania/química , Withania/crescimento & desenvolvimento , Vitanolídeos/metabolismoRESUMO
The present study was undertaken to delineate genotype-environment interactions and stability status of 16 genotypes of ashwagandha (Withania somnifera (L.) Dunal) in context to the 12 characters, namely plant height, number of primary branches, number of secondary branches, days to flowering, days to maturity, number of berries, number of seeds/berry, root length, root diameter, root branches, dry root yield and total alkaloid content (%). Experiment was carried out in a randomized complete block design with three replicationsover three different locations (S. K. Nagar, Jagudan and Bhiloda) in north Gujarat for three years (2016-17, 2017-18 and 2018-19). Pooled analysis of variance revealed that the mean squares due to genotypes and genotype 9 environment interaction along with linear and nonlinear components were highly significant (P<0.01) for most of the traits under study. Stability parameters for component traits through Eberhart and Russell model showed that genotypes that can be used directly in breeding programme are SKA-4 for early flowering, SKA-21 for early maturity and SKA-1, SKA-4, SKA-6 and SKA-17 for shorter plant height. Further, SKA-21 could be used for improving number of primary branches per plant, SKA-11 and SKA-17 for number of secondary branches per plant, SKA-19 for number of berries per plant, SKA-6, SKA-21, SKA-27 and AWS-1 for root branches and SKA-17 for root length as these genotypes were found to be moststable across the environments for mentioned traits. The result revealed that some reliable predictions about genotype 9 environment interaction and its unpredictable components were involved significantly in determining the stability of genotypes. Hence, the present investigation can be exploited for the identification of more productive genotypes in specific environments, leading to significant increase in root productivity of ashwagandha.
Assuntos
Interação Gene-Ambiente , Melhoramento Vegetal , Raízes de Plantas/genética , Withania/anatomia & histologia , Withania/genética , Genótipo , Fenótipo , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/crescimento & desenvolvimento , Withania/crescimento & desenvolvimentoRESUMO
Despite the vast exploration of endophytic microbes for growth enhancement in various crops, knowledge about their impact on the production of therapeutically important secondary metabolites is scarce. In the current investigation, chitinolytic bacterial endophytes were isolated from selected medicinal plants and assessed for their mycolytic as well as plant growth promoting potentials. Among them the two most efficient bacterial endophytes namely Bacillus amyloliquefaciens (MPE20) and Pseudomonas fluorescens (MPE115) individually as well as in combination were able to modulate withanolide biosynthetic pathway and tolerance against Alternaria alternata in Withania somnifera. Interestingly, the expression level of withanolide biosynthetic pathway genes (3-hydroxy-3-methylglutaryl co-enzyme A reductase, 1-deoxy-D-xylulose-5-phosphate reductase, farnesyl di-phosphate synthase, squalene synthase, cytochrome p450, sterol desaturase, sterol Δ-7 reductase and sterol glycosyl transferases) were upregulated in plants treated with the microbial consortium under A. alternata stress. In addition, application of microbes not only augmented withaferin A, withanolide A and withanolide B content (1.52-1.96, 3.32-5.96 and 12.49-21.47 fold, respectively) during A. alternata pathogenicity but also strengthened host resistance via improvement in the photochemical efficiency, normalizing the oxidized and non-oxidized fraction, accelerating photochemical and non-photochemical quantum yield, and electron transport rate. Moreover, reduction in the passively dissipated energy of PSI and PSII in microbial combination treated plants corroborate well with the above findings. Altogether, the above finding highlights novel insights into the underlying mechanisms in application of endophytes and emphasizes their capability to accelerate biosynthesis of withanolides in W. somnifera under biotic stress caused by A. alternata.
Assuntos
Bactérias/metabolismo , Vias Biossintéticas , Endófitos/metabolismo , Withania/microbiologia , Vitanolídeos/metabolismo , Alternaria/patogenicidade , Antibiose , Antifúngicos , Bacillus amyloliquefaciens/enzimologia , Bacillus amyloliquefaciens/genética , Bacillus amyloliquefaciens/isolamento & purificação , Bacillus amyloliquefaciens/metabolismo , Bactérias/enzimologia , Bactérias/genética , Bactérias/isolamento & purificação , Vias Biossintéticas/genética , DNA Bacteriano/análise , Endófitos/enzimologia , Endófitos/genética , Fungos/efeitos dos fármacos , Fungos/patogenicidade , Interações Hospedeiro-Patógeno , Índia , Plantas Medicinais , Pseudomonas fluorescens/enzimologia , Pseudomonas fluorescens/genética , Pseudomonas fluorescens/metabolismo , Estresse Fisiológico , Regulação para Cima , Withania/crescimento & desenvolvimentoRESUMO
Withania somnifera (Ashwagandha) has recently been studied extensively for its health-supplementing and therapeutic activities against a variety of ailments. Several independent studies have experimentally demonstrated pharmaceutical potential of its active Withanolides, Withaferin A (Wi-A), Withanone (Wi-N) and Withanolide A (Wil-A). However, to promote its use in herbal industry, an environmentally sustainable cultivation and high yield are warranted. In modern agriculture strategies, there has been indiscriminate use of chemical fertilizers to boost the crop-yield, however the practice largely ignored its adverse effect on the quality of soil and the environment. In view of these, we attempted to recruit Vermicompost (Vcom, 20-100%) as an organic fertilizer of choice during the sowing and growing phases of Ashwagandha plants. We report that (i) pre-soaking of seeds for 12 h in Vermicompost leachate (Vcom-L) and Vermicompost tea (Vcom-T) led to higher germination, (ii) binary combination of pre-soaking of seeds and cultivation in Vcom (up to 80%) resulted in further improvement both in germination and seedling growth, (iii) cultivated plants in the presence of Vcom+Vcom-L showed higher leaf and root mass, earlier onset of flowering and fruiting and (iv) leaves from the Vcom+Vcom-L cultivated plants showed higher level of active Withanolides, Withanone (Wi-N), Withanolide A (Wil-A) and Withaferin A (Wi-A) and showed anticancer activities in cell culture assays. Taken together, we report a simple and inexpensive method for improving the yield and pharmaceutical components of Ashwagandha leaves.
Assuntos
Biomassa , Triterpenos/metabolismo , Withania/química , Vitanolídeos/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Compostagem , Fertilizantes , Humanos , Extratos Vegetais/química , Folhas de Planta/química , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Sementes/crescimento & desenvolvimento , Solo/química , Triterpenos/química , Triterpenos/farmacologia , Withania/crescimento & desenvolvimento , Withania/metabolismo , Vitanolídeos/química , Vitanolídeos/farmacologiaRESUMO
Withanolides are a collection of naturally occurring, pharmacologically active, secondary metabolites synthesized in the medicinally important plant, Withania somnifera. These bioactive molecules are C28-steroidal lactone triterpenoids and their synthesis is proposed to take place via the mevalonate (MVA) and 2-C-methyl-d-erythritol-4-phosphate (MEP) pathways through the sterol pathway using 24-methylene cholesterol as substrate flux. Although the phytochemical profiles as well as pharmaceutical activities of Withania extracts have been well studied, limited genomic information and difficult genetic transformation have been a major bottleneck towards understanding the participation of specific genes in withanolide biosynthesis. In this study, we used the Tobacco rattle virus (TRV)-mediated virus-induced gene silencing (VIGS) approach to study the participation of key genes from MVA, MEP and triterpenoid biosynthesis for their involvement in withanolide biosynthesis. TRV-infected W. somnifera plants displayed unique phenotypic characteristics and differential accumulation of total Chl as well as carotenoid content for each silenced gene suggesting a reduction in overall isoprenoid synthesis. Comprehensive expression analysis of putative genes of withanolide biosynthesis revealed transcriptional modulations conferring the presence of complex regulatory mechanisms leading to withanolide biosynthesis. In addition, silencing of genes exhibited modulated total and specific withanolide accumulation at different levels as compared with control plants. Comparative analysis also suggests a major role for the MVA pathway as compared with the MEP pathway in providing substrate flux for withanolide biosynthesis. These results demonstrate that transcriptional regulation of selected Withania genes of the triterpenoid biosynthetic pathway critically affects withanolide biosynthesis, providing new horizons to explore this process further, in planta.
Assuntos
Vias Biossintéticas/genética , Inativação Gênica , Genes de Plantas , Vírus de Plantas/fisiologia , Plantas Medicinais/genética , Withania/genética , Vitanolídeos/metabolismo , Carotenoides/metabolismo , Clorofila/metabolismo , Regulação para Baixo/genética , Eritritol/análogos & derivados , Eritritol/metabolismo , Regulação da Expressão Gênica de Plantas , Ácido Mevalônico/metabolismo , Fenótipo , Folhas de Planta/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/anatomia & histologia , Plantas Geneticamente Modificadas , Plantas Medicinais/anatomia & histologia , Plantas Medicinais/crescimento & desenvolvimento , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fosfatos Açúcares/metabolismo , Withania/anatomia & histologia , Withania/crescimento & desenvolvimentoRESUMO
Withania somnifera (L.) Dunal (family: Solanaceae), commonly known as "Indian Ginseng", is a medicinally and industrially important plant of the Indian subcontinent and other warmer parts of the world. The plant has multi-use medicinal potential and has been listed among 36 important cultivated medicinal plants of India that are in high demand for trade due to its pharmaceutical uses. The medicinal importance of this plant is mainly due to the presence of different types of steroidal lactones- withanolides in the roots and leaves. Owing to low seed viability and poor germination, the conventional propagation of W. somnifera falls short to cater its commercial demands particularly for secondary metabolite production. Therefore, there is a great need to develop different biotechnological approaches through tissue and organ culture for seasonal independent production of plants in large scale which will provide sufficient raw materials of uniform quality for pharmaceutical purposes. During past years, a number of in vitro plant regeneration protocols via organogenesis and somatic embryogenesis and in vitro conservation through synthetic seed based encapsulation technology have been developed for W. somnifera. Several attempts have also been made to standardize the protocol of secondary metabolite production via tissue/organ cultures, cell suspension cultures, and Agrobacterium rhizogenes-mediated transformed hairy root cultures. Employment of plant tissue culture based techniques would provide means for rapid propagation and conservation of this plant species and also provide scope for enhanced production of different bioactive secondary metabolites. The present review provides a comprehensive report on research activities conducted in the area of tissue culture and secondary metabolite production in W. somnifera during the past years. It also discusses the unexplored areas which might be taken into consideration for future research so that the medicinal properties and the secondary metabolites produced by this plant can be exploited further for the benefit of human health in a sustainable way.
Assuntos
Técnicas de Cultura de Tecidos , Withania/crescimento & desenvolvimento , Withania/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/metabolismo , Técnicas de Embriogênese Somática de Plantas , Metabolismo Secundário , Sementes/crescimento & desenvolvimentoRESUMO
Withania somnifera is a highly valued medicinal plant in traditional home medicine and is known for a wide range of bioactivities. Its commercial cultivation is adversely affected by poor seed viability and germination. Infestation by various pests and pathogens, survival under unfavourable environmental conditions, narrow genetic base, and meager information regarding biosynthesis of secondary metabolites are some of the other existing challenges in the crop. Biotechnological interventions through organ, tissue, and cell culture provide promising options for addressing some of these issues. In vitro propagation facilitates conservation and sustainable utilization of the existing germplasms and broadening the genetic base. It would also provide means for efficient and rapid mass propagation of elite chemotypes and generating uniform plant material round the year for experimentation and industrial applications. The potential of in vitro cell/organ cultures for the production of therapeutically valuable compounds and their large-scale production in bioreactors has received significant attention in recent years. In vitro culture system further provides distinct advantage for studying various cellular and molecular processes leading to secondary metabolite accumulation and their regulation. Engineering plants through genetic transformation and development of hairy root culture system are powerful strategies for modulation of secondary metabolites. The present review highlights the developments and sketches current scenario in this field.
Assuntos
Biotecnologia/métodos , Técnicas de Cultura de Células/métodos , Compostos Fitoquímicos/química , Técnicas de Cultura de Tecidos/métodos , Withania/química , Biomassa , Reatores Biológicos , Proliferação de Células , Germinação , Extratos Vegetais/química , Raízes de Plantas/química , Brotos de Planta/química , Brotos de Planta/crescimento & desenvolvimento , Plantas Medicinais/química , Plantas Medicinais/crescimento & desenvolvimento , Substâncias Protetoras/química , Withania/crescimento & desenvolvimento , Vitanolídeos/químicaRESUMO
Ashwagandha (Withania somnifera) is an Ayurvedic herb commonly used in world-renowned traditional Indian home medicine system. Roots of Ashwagandha have been traditionally known to possess a variety of therapeutic and health promoting potentials that have not been sufficiently supported by laboratory studies. Nevertheless, most, if not all, of the preventive and therapeutic potentials have been assigned to its bioactive components, steroidal alkaloids and lactones. In contrast to the traditional use of roots, we have been exploring bioactivities in leaves of Ashwagandha. Here, we report that the leaves possess higher content of active Withanolides, Withaferin-A (Wi-A) and Withanone (Wi-N), as compared to the roots. We also established, for the first time, hydroponic cultivation of Ashwagandha and investigated the effect of various cultivation conditions on the content of Wi-A and Wi-N by chemical analysis and bioassays. We report that the Withanone/Withaferin A-rich leaves could be obtained by manipulating light condition during hydroponic cultivation. Furthermore, we recruited cyclodextrins to prepare extracts with desired ratio of Wi-N and Wi-A. Hydroponically grown Ashwagandha and its extracts with high ratio of withanolides are valuable for cancer treatment.
Assuntos
Hidroponia/métodos , Extratos Vegetais/química , Folhas de Planta/química , Withania/química , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Etanol/química , Humanos , Luz , Ayurveda , Extratos Vegetais/farmacologia , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/efeitos da radiação , Reprodutibilidade dos Testes , Fatores de Tempo , Triterpenos/análise , Triterpenos/isolamento & purificação , Withania/crescimento & desenvolvimento , Withania/efeitos da radiação , Vitanolídeos/análise , Vitanolídeos/isolamento & purificaçãoRESUMO
Withania somnifera (L.) Dunal known as Ashwagandha is commonly used in traditional Indian medicine system. It possesses immense therapeutic value against a large number of ailments such as mental diseases, asthma, inflammation, arthritis, rheumatism, tuberculosis, and a variety of other diseases including cancer. The therapeutic potential of W. somnifera is due to the presence of secondary metabolites mainly, tropane alkaloids and withanolides (steroidal lactones). The growing realization of commercial value of the plant has initiated a new demand for in vitro propagation of elite chemotypes of Withania. Micropropagation which is an important tool for rapid multiplication requires optimization of number of factors such as nutrient medium, status of medium (solid and liquid), type of explant, and plant growth regulators. Similarly, an efficient and reproducible in vitro regeneration system which is a prerequisite for the development of genetic transformation protocol requires precise manipulation of various intrinsic and extrinsic factors.
Assuntos
Técnicas de Cultura/métodos , Withania/crescimento & desenvolvimento , Meios de Cultura/metabolismo , Técnicas Histológicas/métodos , Reguladores de Crescimento de Plantas/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/fisiologia , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/fisiologia , Regeneração , Withania/fisiologia , Vitanolídeos/metabolismoRESUMO
Withania coagulans (Stocks) Dunal (Solanaceae), also known as 'Panir Bandh' is an important medicinal plant that is extensively used as a home remedy for several diseases in the Indian subcontinent. The plant possesses specific steroidal lactones known as withanolides which show high level of pharmaceutical activity against a broad spectrum of microorganisms. Natural propagation of the plant occurs through Seed but due to unisexual nature of the flowers; chances of Seed setting are very limited and the plant is on the verge of extinction because of overexploitation and reproductive failure. Plant tissue culture techniques offer opportunities for ex situ conservation and mass multiplication of endangered plant species through micropropagation and also enhancement of in vitro biosynthesis of bioactive compounds. In this chapter we present protocols for the mass multiplication of W. coagulans, assessment of clonal fidelity by RAPD, and estimation of bioactive compounds (withanolides) by thin layer chromatography (TLC) and reverse phase HPLC developed in our laboratory.
Assuntos
Lactonas/análise , Brotos de Planta/crescimento & desenvolvimento , Técnicas de Cultura de Tecidos/métodos , Withania/crescimento & desenvolvimento , Vitanolídeos/análise , Cromatografia Líquida de Alta Pressão/métodos , Cromatografia em Camada Fina/métodos , Lactonas/metabolismo , Brotos de Planta/genética , Brotos de Planta/metabolismo , Técnica de Amplificação ao Acaso de DNA Polimórfico/métodos , Withania/genética , Withania/metabolismo , Vitanolídeos/metabolismoRESUMO
Plant tissue culture offers several techniques for rapid clonal propagation, germplasm conservation, regeneration of genetically manipulated superior clones, production of phyto-constituents, and ex vitro conservation of valuable phytodiversity. An improved and efficient micropropagation protocol for Withania somnifera (L.), a drug-producing medicinal plant, using juvenile explants (nodal explants) has been developed. Highest multiplication and subsequent elongation of shoots is observed on MS medium containing BA and NAA. The regenerated microshoots roots best on ½ MS medium containing NAA, established in earthen pots containing garden soil and are maintained in the greenhouse with 95 % survival rate. Genetic uniformity of micropropagated plants is confirmed by PCR-based DNA fingerprinting techniques, viz., RAPD and ISSR. No variation is observed in DNA fingerprinting patterns among the micropropagated plants, which are similar to that of the donor plant illustrating their genetic uniformity.
Assuntos
Withania/crescimento & desenvolvimento , Withania/genética , Técnicas de Cultura de Células/métodos , Meios de Cultura/metabolismo , Impressões Digitais de DNA , DNA de Plantas/genética , Variação Genética , Melhoramento Vegetal/métodos , Reguladores de Crescimento de Plantas/metabolismo , Reação em Cadeia da Polimerase/métodos , Plântula/citologia , Plântula/genética , Plântula/crescimento & desenvolvimento , Withania/citologiaRESUMO
Soil contaminated by Petroleum oil cannot be utilized for agricultural purposes due to hydrocarbon toxicity. Oil contaminated soil induces toxicity affecting germination, growth and productivity. Several technologies have been proposed for bioremediation of oil contaminated sites, but remediation through biosurfactant producing plant growth promontory rhizobacteria (PGPR) is considered to be most promising methods. In the present study the efficacy of seed priming on growth and pigment of Withania somnifera under petroleum toxicity is explored. Seeds of W. somnifera were primed with biosurfactant producing Pseudomonas sp. AJ15 with plant growth promoting traits having potentiality to utilized petroleum as carbon source. Results indicates that plant arose from priming seeds under various petroleum concentration expressed high values for all the parameters studied namely germination, shoot length, root length, fresh and dry weight and pigments (chlorophyll and carotenoid) as compared to non primed seed. Hence, the present study signifies that petroleum degrarding biosurfactant producing PGPR could be further used for management and detoxification of petroleum contaminated soils for growing economically important crops.
Assuntos
Petróleo/análise , Microbiologia do Solo , Poluentes do Solo/análise , Solo/química , Withania/efeitos dos fármacos , Biodegradação Ambiental , Germinação/efeitos dos fármacos , Petróleo/toxicidade , Pseudomonas/crescimento & desenvolvimento , Rhizobium/crescimento & desenvolvimento , Sementes/efeitos dos fármacos , Sementes/crescimento & desenvolvimento , Poluentes do Solo/toxicidade , Withania/crescimento & desenvolvimentoRESUMO
CONTEXT: Cancer, being a cause of death for major fraction of population worldwide, is one of the most studied diseases and is being investigated for the development of new technologies and more accurate therapies. Still the currently available therapies for cancer have many lacunae which affect the patient's health severely in the form of side effects. The natural drugs obtained from the medicinal plants provide a better alternative to fight against this devastating disease. Withania somnifera L. Dunal (Solanaceae), a well-known Ayurvedic medicinal plant, has been traditionally used to cure various ailments for centuries. OBJECTIVES: Considering the immense potential of W. somnifera, this review provides a detail account of its vital phytoconstituents and summarizes the present status of the research carried out on its anticancerous activities, giving future directions. METHODS: The sources of scientific literature were accessed from various electronic databases such as PubMed, Google Scholar, Science Direct, and library search. RESULTS: Various parts of W. somnifera especially the roots with its unique contents have been proved effective against different kinds of cancers. The most active components withanolides and withaferins along with a few other metabolites including withanone (WN) and withanosides have been reported effective against different types of cancer cell lines. CONCLUSION: This herb holds an important place among various anticancer medicinal plants. It is very essential to further screen and to investigate different formulations for anticancer therapy in vitro as well as in vivo in combination with established chemotherapy.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Descoberta de Drogas , Extratos Vegetais/farmacologia , Withania/química , Animais , Antineoplásicos Fitogênicos/isolamento & purificação , Descoberta de Drogas/métodos , Descoberta de Drogas/tendências , Humanos , Estrutura Molecular , Extratos Vegetais/isolamento & purificação , Folhas de Planta/química , Folhas de Planta/crescimento & desenvolvimento , Raízes de Plantas/química , Raízes de Plantas/crescimento & desenvolvimento , Withania/crescimento & desenvolvimentoRESUMO
Sterol glycosyltransferases (SGTs) belong to family 1 of glycosyltransferases (GTs) and are enzymes responsible for synthesis of sterol-glucosides (SGs) in many organisms. WsSGTL1 is a SGT of Withania somnifera that has been found associated with plasma membranes. However its biological function in W.somnifera is largely unknown. In the present study, we have demonstrated through RNAi silencing of WsSGTL1 gene that it performs glycosylation of withanolides and sterols resulting in glycowithanolides and glycosylated sterols respectively, and affects the growth and development of transgenic W.somnifera. For this, RNAi construct (pFGC1008-WsSGTL1) was made and genetic transformation was done by Agrobacterium tumefaciens. HPLC analysis depicts the reduction of withanoside V (the glycowithanolide of W.somnifera) and a large increase of withanolides (majorly withaferin A) content. Also, a significant decrease in level of glycosylated sterols has been observed. Hence, the obtained data provides an insight into the biological function of WsSGTL1 gene in W.somnifera.
Assuntos
Glicosiltransferases/metabolismo , Proteínas de Plantas/metabolismo , Interferência de RNA , Withania/enzimologia , Withania/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Vetores Genéticos/metabolismo , Glicosilação , Filogenia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Regeneração , Esteróis/biossíntese , Transformação Genética , Withania/genética , Vitanolídeos/metabolismoRESUMO
We studied the influence of sucrose and nitrogen concentration on in vitro flowering and fruit setting in elongated shoots of Withania somnifera. BA (1.5 mg/l) and IAA (0.3 mg/l) on MS medium supplemented with 4% sucrose showed 67% of in vitro flower induction frequency, 9 flowers/shoot, 4 fruits/shoot and 11 seeds/fruit in elongated-shoots. Different concentrations of nitrogen sources (L-glutamine, adenine sulphate, ammonium nitrate, potassium nitrate and sodium nitrate 5-25 mg/l) were tested in combination with 4% sucrose and BA at 1.5 mg/l and IAA at 0.3 mg/l. Highest number of flowers (20 flowers/shoot; 2.2-fold) and fruits (16 fruits/shoot; 3.39-fold), fruit setting (12 seeds/fruit; 1.08-fold) at a higher frequency (88%) were achieved on MS medium augmented with 15 mg/l adenine sulphate with same PGRs and sucrose concentration. The maximum production of withanolide A (0.68 mg/g DW) and withanolide B (0.77 mg/g DW) was recorded in in vitro fruits. Highest accumulation of withaferin A (2 mg/g DW) was quantified from in vitro flowers, whereas, it was low in in vitro fruits (0.49 mg/g DW withaferin A). However, withanone (0.23 mg/g DW) was found accumulated uniformly in both in vitro flowers and fruits compared to control.