Phyto-therapeutic potential of Withania somnifera: Molecular mechanism and health implications.

Withania somnifera, the plant named Indian ginseng, Ashwagandha, or winter cherry, has been used since ancient times to cure various health ailments. Withania somnifera is rich in constituents belonging to chemical classes like alkaloids, saponins, flavonoids, phenolic acids, and withanolides. Several chemotypes were identified based on their phytochemical composition and credited for their multiple bioactivities. Besides, exhibiting neuroprotective, immunomodulatory, adaptogenic, anti-stress, bone health, plant has shown promising anti-cancer properties. Several withanolides have been reported to play a crucial role in cancer; they target cancer cells by different mechanisms such as modulating the expression of tumor suppressor genes, apoptosis, telomerase expression, and regulating cell signaling pathway. Though, many treatments are available for cancer; however, to date, no assured reliable cure for cancer is made available. Additionally, synthetic drugs may lead to development of resistance in time; therefore, focus on new and natural drugs for cancer therapeutics may prove a longtime effective alternative. This current report is a comprehensive combined analysis upto 2023 with articles focused on bio-activities of plant Withania somnifera from various sources, including national and international government sources. This review focuses on understanding of various mechanisms and pathways to inhibit uncontrolled cell growth by W. somnifera bioactives, as reported in literature. This review provides a recent updated status of the W. somnifera on pharmacological properties in general and anti-cancer in particular and may provide a guiding resource for researchers associated with natural product-based cancer research and healthcare management.

New withanolides from leaves along with alkyl glucoside from roots and stem as well as variations in withanolide contents in leaves of accessions of Withania somnifera, Ashwagandha- the Indian ginseng$.

Based on the major components in the leaves, the ashwagandha has been found to exist in several chemotypic forms in India. From the leaves of various accessions of Withania somnifera, which were maintained in our institute, three new steroids namely, 4-acetoxy-20ß-hydroxy-1-oxo-witha-2,5,24-trienolide (7), 24,25-dihydro-14α-hydroxy withanolide D (9), 5α,6ß,17α,27-tetrahydroxy-1-oxo-witha-2,24-dienolide (12) together with thirteen known withanolides were identified by spectroscopic methods. From the roots and stem of one accession and leaves of another, a new alkyl ester glucoside (4) has also been isolated. The new withanolides 7, 9 and 12 have been tentatively named as withanolide 135 A, withanolide 135B and withanolide 108, respectively.

Cloning and homologous characterization of geranylgeranyl pyrophosphate synthase (GGPPS) from Withania somnifera revealed alterations in metabolic flux towards gibberellic acid biosynthesis.

MAIN CONCLUSION: Overexpression of a novel geranylgeranyl pyrophosphate synthase gene (WsGGPPS) in planta resulted in increased levels of gibberellic acid and decrease in withanolide content. Withania somnifera (L.) Dunal, the herb from family Solanaceae is one of the most treasured medicinal plant used in traditional medicinal systems owing to its unique stockpile of pharmaceutically active secondary metabolites. Phytochemical and pharmacological studies in this plant were well established, but the genes affecting the regulation of biosynthesis of major metabolites were not well elucidated. In this study cloning and functional characterization of a key enzyme in terpenoid biosynthetic pathway viz. geranylgeranyl pyrophosphate synthase (EC 2.5.1.29) gene from Withania somnifera was performed. The full length WsGGPPS gene contained 1,104 base pairs that encode a polypeptide of 365 amino acids. The quantitative expression analysis suggested that WsGGPPS transcripts were expressed maximally in flower tissues followed by berry tissues. The expression levels of WsGGPPS were found to be regulated by methyl jasmonate (MeJA) and salicylic acid (SA). Amino acid sequence alignment and phylogenetic studies suggested that WsGGPPS had close similarities with GGPPS of Solanum tuberosum and Solanum pennellii. The structural analysis provided basic information about three dimensional features and physicochemical parameters of WsGGPPS protein. Overexpression of WsGGPPS in planta for its functional characterization suggested that the WsGGPPS was involved in gibberellic acid biosynthesis.

Molecular cloning and characterization of Triterpenoid Biosynthetic Pathway Gene HMGS in Centella asiatica (Linn.).

BACKGROUND: The flux of isoprenoids and the total accumulation of triterpenoid saponins known as centellosides in C. asiatica are controlled by the key genes of the Mevalonate pathway (MVA). These genes were reported to have positive regulation of the pathway in providing isoprenoid moieties. Though, some information is available on the pathway and secondary metabolites. However, most of the pathway steps are not characterized functionally. METHODOLOGY AND RESULTS: For the study, full-length pathway gene Hydroxymethyl glutaryl-CoA-synthase (CaHMGS; GenBank accession number: MZ997833), was isolated from previously annotated transcriptome data of Centella asiatica leaves. HMGS has been successfully cloned and heterologously expressed in bacteria E. coli strain DH5α. The cloned gene has been sequenced and further characterized through in silico studies by different bioinformatics tools. Also, the gene sequences have been submitted in NCBI. In silico studies of isolated gene sequence revealed the nature, characteristics of genes. The ORF of HMGS is 1449 bp encoding 482 amino acids. Predicted molecular weight (MW) of HMGS was 48.09 kDa and theoretical pI was 5.97. Blast results and Multiple sequence alignments of the gene showing the similarity with HMGS of other plants of their respective families. The Molecular Evolutionary Genetic Analysis (MEGA) version 10.1.6 was used to construct a phylogenetic tree. Differential tissue-specific expression of different plant parts was also checked. Tissue expression patterns unveiled that the highest expression level of the CaHMGS had been seen in the roots and lowest in the node of the plant. Functional complementation experiment of the CaHMGS in Saccharomyces cerevisiae wild strain YSC1021 and haploid strain YSC1021 which lack HMGS protein confirmed that the CaHMGS gene encodes functional CaHMGS that catalyzed the biosynthesis of mevalonate in yeast. CONCLUSIONS: The gene was reported, cloned and characterized first time in Centella asiatica. Understanding this biosynthetic pathway gene will further help in the improvement of plants for enhanced secondary metabolites production.

Comparative transcriptome analysis to identify putative genes related to trichome development in Ocimum species.

Genus Ocimum is known to have species possessing important therapeutic essential oil. The major phytoconstituents of essential oil in Ocimum species are phenylpropanoids and terpenoids. The essential oil is accumulated in the trichomes; the specialized structures predominantly found on leaves and other tissues. The development of trichome is integrated with development of plant and leaf and also tightly coordinated with the primary and secondary metabolic pathways producing essential oil constituents. In continuation to our studies on elucidating/understanding the mechanism of biosynthesis of  essential oil pathways in Ocimum species, we have performed comparative transcriptome analysis to investigate the role of trichome-related gene expression in the regulation of biosynthetic pathways of essential oil. The essential oil biogenesis is tightly integrated with primary metabolic activities, the analysis for the expression pattern of genes related to primary metabolism and its relationship with secondary metabolism was evaluated in comparative manner. Physiological parameters in relation to primary metabolism such as photosynthetic pigment content, soluble sugar content, and invertase enzymes along with morphological parameters were analysed in O. basilicum and O. sanctum. Differential expression profiling uncovered about 8116 and 2810 differentially expressed transcripts in O. basilicum and O. sanctum, respectively. Enrichment of differentially expressed genes were analysed in relation to metabolic pathways, primary metabolism and secondary metabolism. Trichome related genes identified from the Ocimum species vis-à-vis their expression profiles suggested higher expression in O. basilicum. The findings in this study provide interesting insights into the role of trichome-related transcripts in relation to essential oil content in Ocimum species. The study is valuable as this is the first study on revealing the transcripts and their role in trichome development and essential oil biogenesis in two major species of Ocimum.

WRKY1-mediated regulation of tryptophan decarboxylase in tryptamine generation for withanamide production in Withania somnifera (Ashwagandha).

KEY MESSAGE: WsWRKY1-mediated transcriptional modulation of Withania somnifera tryptophan decarboxylase gene (WsTDC) helps to regulate fruit-specific tryptamine generation for production of withanamides. Withania somnifera is a highly valued medicinal plant. Recent demonstration of novel indolyl metabolites called withanamides in its fruits (berries) prompted us to investigate its tryptophan decarboxylase (TDC), as tryptophan is invariably a precursor for indole moiety. TDC catalyzes conversion of tryptophan into tryptamine, and the catalytic reaction constitutes a committed metabolic step for synthesis of an array of indolyl metabolites. The TDC gene (WsTDC) was cloned from berries of the plant and expressed in E. coli. The recombinant enzyme was purified and characterized for its catalytic attributes. Catalytic and structural aspects of the enzyme indicated its regulatory/rate-limiting significance in generation of the indolyl metabolites. Novel tissue-wise and developmentally differential abundance of WsTDC transcripts reflected its preeminent role in withanamide biogenesis in the fruits. Transgenic lines overexpressing WsTDC gene showed accumulation of tryptamine at significantly higher levels, while lines silenced for WsTDC exhibited considerably depleted levels of tryptamine. Cloning and sequence analysis of promoter of WsTDC revealed the presence of W-box in it. Follow-up studies on isolation of WsWRKY1 transcription factor and its overexpression in W. somnifera revealed that WsTDC expression was substantially induced by WsWRKY1 resulting in overproduction of tryptamine. The study invokes a key role of TDC in regulating the indolyl secondary metabolites through enabling elevated flux/supply of tryptamine at multiple levels from gene expression to catalytic attributes overall coordinated by WsWRKY1. This is the first biochemical, molecular, structural, physiological and regulatory description of a fruit-functional TDC.

Over-expression of DXS gene enhances terpenoidal secondary metabolite accumulation in rose-scented geranium and Withania somnifera: active involvement of plastid isoprenogenic pathway in their biosynthesis.

Rose-scented geranium (Pelargonium spp.) is one of the most important aromatic plants and is well known for its diverse perfumery uses. Its economic importance is due to presence of fragrance rich essential oil in its foliage. The essential oil is a mixture of various volatile phytochemicals which are mainly terpenes (isoprenoids) in nature. In this study, on the geranium foliage genes related to isoprenoid biosynthesis (DXS, DXR and HMGR) were isolated, cloned and confirmed by sequencing. Further, the first gene of 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway, 1-deoxy-d-xylulose-5-phosphate synthase (GrDXS), was made full length by using rapid amplification of cDNA ends strategy. GrDXS contained a 2157 bp open reading frame that encoded a polypeptide of 792 amino acids having calculated molecular weight 77.5 kDa. This study is first report on heterologous expression and kinetic characterization of any gene from this economically important plant. Expression analysis of these genes was performed in different tissues as well as at different developmental stages of leaves. In response to external elicitors, such as methyl jasmonate, salicylic acid, light and wounding, all the three genes showed differential expression profiles. Further GrDXS was over expressed in the homologous (rose-scented geranium) as well as in heterologous (Withania somnifera) plant systems through genetic transformation approach. The over-expression of GrDXS led to enhanced secondary metabolites production (i.e. essential oil in rose-scented geranium and withanolides in W. somnifera). To the best of our knowledge, this is the first report showing the expression profile of the three genes related to isoprenoid biosynthesis pathways operated in rose-scented geranium as well as functional characterization study of any gene from rose-scented geranium through a genetic transformation system.

Comparative transcripts profiling of fruit mesocarp and endocarp relevant to secondary metabolism by suppression subtractive hybridization in Azadirachta indica (neem).

Azadirachta indica (neem) is a medicinally important plant that is valued for its bioactive secondary metabolites. Higher levels of the bioactive phytochemicals are accumulated in fruits than in other tissues. In the present study, a total of 387 and 512 ESTs, respectively, from endocarp and mesocarp of neem fruits were isolated and analyzed. Out of them 318 ESTs (82.17%) clones from endocarp and 418 ESTs (81.64%) from mesocarp encoded putative proteins that could be classified into three major gene ontology categories: biological process, molecular function and cellular component. From the analyses of contigs, 73 unigenes from the forward subtracted library and 35 unigenes from the reverse subtracted library were obtained. The ESTs from mesocarp encoded cytochrome P450 enzymes, which indicated hydroxylation to be a major metabolic event and that biogeneration of hydroxylated neem fruit phytochemicals was differentially regulated with developmental stage-specificity of synthesis. Through this study, we present the first report of any gene expression data in neem tissues. Neem hydroxy-methyl glutaryl-coenzyme A reductase (NHMGR) gene was used as expressing control vis-a-vis subtracted tissues. NHMGR was present in fruit, endocarp and mesocarp tissues, but absent in subtractive libraries, revealing that it was successfully eliminated during subtraction. Eight genes of interest from subtracted libraries were profiled for their expression in fruit, mesocarp and endocarp. Expression profiles validated the quality of the libraries and functional diversity of the tissues. The subtractive cDNA library and EST database described in this study represent a valuable transcript sequence resource for future research aimed at improving the economically important medicinal plant.

Expanding roles of cross-talk between hydrogen sulfide and nitric oxide under abiotic stress in plants.

Abiotic stress such as salt, heavy metals, drought, temperature, and others can affect plants from seed germination to seedling growth to reproductive maturity. Abiotic stress increases reactive oxygen species and lowers antioxidant enzymes in plants resulted the plant tolerance ability against stress conditions decrease. Hydrogen sulfide (H2S) and nitric oxide (NO) are important gasotransmitters involved in seed germination, photosynthesis, growth and development, metabolism, different physiological processes and functions in plants. In plants, various enzymes are responsible for the biosynthesis of both H2S and NO via both enzymatic and non-enzymatic pathways. They also mediate post-translation modification, such as persulfidation, and nitrosylation, which are protective mechanisms against oxidative damage. They also regulate some cellular signalling pathways in response to various abiotic stress. H2S and NO also stimulate biochemical reactions in plants, including cytosolic osmoprotectant accumulation, reactive oxygen species regulation, antioxidant system activation, K+ uptake, and Na+ cell extrusion or vacuolar compartmentation. In this review, we summarize how H2S and NO interact with each other, the function of both H2S and NO, the mechanism of biosynthesis, and post-translational modification under different abiotic stress. Our main emphasis was to find the cross-talk between NO and H2S and how they regulate genes in plants under abiotic stress.

Differential regulation of key triterpene synthase gene under abiotic stress in Withania somnifera L. Dunal and its co-relation to sterols and withanolides.

Withania somnifera (Ashwagandha), is one of the most reputed Indian medicinal plants, having immense pharmacological activities due to the occurrence of withanolides. The withanolides are biosynthesized through triterpenoid biosynthetic pathway with the involvement of WsCAS leading to cyclization of 2, 3 oxidosqualene, which is a key metabolite to further diversify to a myriad of phytochemicals. In contrast to the available reports on the studies of WsCAS in withanolide biosynthesis, its involvement in phytosterol biosynthesis needs investigation. Present work deals with the understanding of role of WsCAS triterpenoid synthase gene in the regulation of biosynthesis of phytosterols & withanolides. Docking studies of WsCAS protein revealed Conserved amino acids, DCATE motif, and QW motif which are involved in efficient substrate binding, structure stabilization, and catalytic activity. Overexpression/silencing of WsCAS leading to increment/decline of phytosterols confers its stringent regulation in phytosterols biosynthesis. Differential regulation of WsCAS on the metabolic flux towards phytosterols and withanolide biosynthesis was observed under abiotic stress conditions. The preferential channelization of 2, 3 oxidosqualene towards withanolides and/or phytosterols occurred under heat/salt stress and cold/water stress, respectively. Stigmasterol and ß-sitosterol showed major contribution in high/low temperature and salt stress, and campesterol in water stress management. Overexpression of WsCAS in Arabidopsis thaliana led to the increment in phytosterols in general. Thus, the WsCAS plays important regulatory role in the biosynthetic pathway of phytosterols and withanolides under abiotic stress conditions.

A new source of elemol rich essential oil and existence of multicellular oil glands in leaves of the Dioscorea species.

Dioscorea species is a very important food and drug plant. The tubers of the plant are extensively used in food and drug purposes owing to the presence of steroidal constituent's diosgenin in the tubers. In the present study, we report for the first time that the leaves of Dioscorea composita and Dioscorea floribunda grown under the field conditions exhibited the presence of multicellular oil glands on the epidermal layers of the plants using stereomicroscopy (SM) and scanning electron microscopy (SEM). Essential oil was also isolated from the otherwise not useful herbage of the plant, and gas chromatographic-mass spectroscopy analysis revealed confirmation of the essential oil constituents. Out of the 76 compounds detected in D. floribunda and 37 from D. composita essential oil, major terpenoids which are detected and reported for Dioscorea leaf essential oil are α -terpinene, nerolidol, citronellyl acetate, farnesol, elemol, α -farnesene, valerenyl acetate, and so forth. Elemol was detected as the major constituent of both the Dioscorea species occupying 41% and 22% of D. Floribunda and D. composita essential oils, respectively. In this paper, we report for the first time Dioscorea as a possible novel bioresource for the essential oil besides its well-known importance for yielding diosgenin.

Withanolide biosynthesis recruits both mevalonate and DOXP pathways of isoprenogenesis in Ashwagandha Withania somnifera L. (Dunal).

Withanolides are pharmaceutically important C(28)-phytochemicals produced in most prodigal amounts and diversified forms by Withania somnifera. Metabolic origin of withanolides from triterpenoid pathway intermediates implies that isoprenogenesis could significantly govern withanolide production. In plants, isoprenogenesis occurs via two routes: mevalonate (MVA) pathway in cytosol and non-mevalonate or DOXP/MEP pathway in plastids. We have investigated relative carbon contribution of MVA and DOXP pathways to withanolide biosynthesis in W. somnifera. The quantitative NMR-based biosynthetic study involved tracing of (13)C label from (13)C(1)-D-glucose to withaferin A in withanolide producing in vitro microshoot cultures of the plant. Enrichment of (13)C abundance at each carbon of withaferin A from (13)C(1)-glucose-fed cultures was monitored by normalization and integration of NMR signal intensities. The pattern of carbon position-specific (13)C enrichment of withaferin A was analyzed by a retro-biosynthetic approach using a squalene-intermediated metabolic model of withanolide (withaferin A) biosynthesis. The pattern suggested that both DOXP and MVA pathways of isoprenogenesis were significantly involved in withanolide biosynthesis with their relative contribution on the ratio of 25:75, respectively. The results have been discussed in a new conceptual line of biosynthetic load-driven model of relative recruitment of DOXP and MVA pathways for biosynthesis of isoprenoids. Key message The study elucidates significant contribution of DOXP pathway to withanolide biosynthesis. A new connotation of biosynthetic load-based role of DOXP/MVA recruitment in isoprenoid biosynthesis has been proposed.

1,4-Dioxane and ergosterol derivatives from Withania somnifera roots.

The chemical investigation on the n-hexane extract of Withania somnifera roots has yielded octacosane, oleic and stearic fatty acids, stigmasterone, stigmasterol, sitostanone, oleanolic acid along with the ergosterol and 1,4-dioxane derivatives as new compounds. The isolation of alkenyl-1,4-dioxane compound is rare, whereas the ergosterol derivative may have biogenetic significance in the lactone formation in the E ring of withanolides. The presence of a 1,4-dioxane derivative in the nonpolar extract of roots assumes importance as this type of compound has not been reported earlier from W. somnifera. The structures of new compounds were elucidated by spectroscopic methods and chemical transformations.

Standardized root extract of Withania somnifera and Withanolide A exert moderate vasorelaxant effect in the rat aortic rings by enhancing nitric oxide generation.

ETHNO-PHARMACOLOGICAL RELEVANCE: Withania somnifera (L.) Dunal, commonly known as Ashwagandha, belongs to the family Solanaceae. In Ayurveda, Ashwagandha has been defined as one of the most important herb and is considered to be the best adaptogen. It is also an excellent rejuvenator, a general health tonic and cure for various disorders such as cerebrovascular, insomnia, asthma, ulcers, etc. Steroidal lactones (Withanolides: Withanolide A, Withaferin A, Withanolide D, Withanone, etc) isolated from this plant, possess promising medicinal properties such as anti-inflammatory, immune-stimulatory etc. Standardized root extract of the plant NMITLI-118R (NM) was prepared at CSIR-CIMAP, and was investigated for various biological activities at CSIR-CDRI. Among the notable medicinal properties, NM exhibited excellent neuroprotective activity in the middle cerebral artery occlusion (MCAO) rat model. AIM OF THE STUDY: Endothelial dysfunction is the primary event in the cerebrovascular or cardiovascular disorders, present study was thus undertaken to evaluate vasoprotective potential of NM and its biomarker compound Withanolide A (WA) using rat aortic rings and EA.hy926 endothelial cells. MATERIAL AND METHODS: Transverse aortic rings of 10 weeks old Wistar rats were used to evaluate effect of NM and WA on the vasoreactivity. While, mechanism of NM and WA mediated vasorelaxant was investigated in Ea.hy926 cell line by measuring NO generation, nitrite content, Serine 1177 phosphorylation of eNOS, reduced/oxidized biopterin levels and expression of endothelial nitric oxide synthase (eNOS) mRNA and protein. RESULTS: Fingerprinting of NM using HPLC identified presence of WA in the extract. NM as well as WA exerted moderate vasorelaxant effect in the endothelium intact rat aortic rings which was lesser than acetylcholine (ACh). NM and WA augmented ACh induced relaxation in the rat aortic rings. NM and WA dependent vasorelaxation was blocked by N-nitro-L-arginine methyl ester (L-NAME) or 1H-[1,2,4] oxadiazolo [4,3,-a]quinoxalin-1-one (ODQ), indicating role of NO/cGMP. Further Ea.hy926 cells treated with NM and WA showed accumulation of nitrite content, enhanced NO levels, eNOS expression and eNOS phosphorylation (Serine 1177). CONCLUSION: Altogether NM and WA dependent improvement in the NO availability seems to be mediated by the enhanced eNOS phosphorylation. WA, seems to be one of the active constituent of NM, and presence of other vasoactive substances cannot be ruled out. The data obtained imply that the vasorelaxant property of NM is beneficial for its neuroprotective potential.

Elucidation of plasma protein binding, blood partitioning, permeability, CYP phenotyping and CYP inhibition studies of Withanone using validated UPLC method: An active constituent of neuroprotective herb Ashwagandha.

ETHNOPHARMACOLOGICAL RELEVANCE: Withanone (WN), an active constituent of Withania somnifera commonly called Ashwagandha has remarkable pharmacological responses along with neurological activities. However, for a better understanding of the pharmacokinetic and pharmacodynamic behavior of WN, a comprehensive in-vitro ADME (absorption, distribution, metabolism, and excretion) studies are necessary. AIM OF THE STUDY: A precise, accurate, and sensitive reverse-phase ultra-performance liquid chromatographic method of WN was developed and validated in rat plasma for the first time. The developed method was successfully applied to the in-vitro ADME investigation of WN. MATERIAL AND METHODS: The passive permeability of WN was assayed using PAMPA plates and the plasma protein binding (PPB) was performed using the equilibrium dialysis method. Pooled liver microsomes of rat (RLM) and human (HLM) were used for the microsomal stability, CYP phenotyping, and inhibition studies. CYP phenotyping was evaluated using the specific inhibitors. CYP inhibition study was performed using specific probe substrates along with WN or specific inhibitors. RESULTS: WN was found to be stable in the simulated gastric and intestinal environment and has a high passive permeability at pH 4.0 and 7.0 in PAMPA assay. The PPB of WN at 5 and 20 µg/mL concentrations were found to be high i.e. 82.01 ± 1.44 and 88.02 ± 1.15%, respectively. The in vitro half-life of WN in RLM and HLM was found to be 59.63 ± 2.50 and 68.42 ± 2.19 min, respectively. CYP phenotyping results showed that WN was extensively metabolized by CYP 3A4 and1A2 enzymes in RLM and HLM. However, the results of CYP Inhibition studies showed that none of the CYP isoenzymes were potentially inhibited by WN in RLM and HLM. CONCLUSION: The in vitro results of pH-dependent stability, plasma stability, permeability, PPB, blood partitioning, microsomal stability, CYP phenotyping, and CYP inhibition studies demonstrated that WN could be a better phytochemical for neurological disorders.

Bioconversion of artemisinin to its nonperoxidic derivative deoxyartemisinin through suspension cultures of Withania somnifera Dunal.

Biotransformation of artemisinin was investigated with two different cell lines of suspension cultures of Withania somnifera. Both cell lines exhibited potential to transform artemisinin into its nonperoxidic analogue, deoxyartemisinin, by eliminating the peroxo bridge of artemisinin. The enzyme involved in the reaction is assumed to be artemisinin peroxidase, and its activity in extracts of W. somnifera leaves was detected. Thus, the non-native cell-free extract of W. somnifera and suspension culture-mediated bioconversion can be a promising tool for further manipulation of pharmaceutical compounds.

Withanolides from Withania somnifera roots.

Two new and seven known withanolides along with beta-sitosterol, stigmasterol, beta-sitosterol glucoside, stigmasterol glucoside, alpha+beta glucose were isolated from the roots of Withania somnifera. Among the known compounds, Viscosa lactone B, stigmasterol, stigmasterol glucoside and alpha+beta glucose are being reported from the roots of W. somnifera for the first time. One of the new compounds contained the rare 16beta-acetoxy-17(20)-ene the other contained unusual 6alpha-hydroxy-5,7alpha-epoxy functional groups in the withasteroid skeleton. The structures were elucidated by spectroscopic methods and chemical transformations.

In vitro withanolide production by Withania somnifera L. cultures.

In vitro multiple shoots, root, callus and cell suspension cultures of Withania somnifera exhibited the potentiality to produce pharmacologically active withanolides. Multiple shoots cultures exhibited an increase in withanolide A accumulation compared to shoots of the mother plant. In vitro generated root cultures as well as callus and suspension cultures also produced withanolides albeit at lower levels.

Prolonged exposure to salt stress affects specialized metabolites-artemisinin and essential oil accumulation in Artemisia annua L.: metabolic acclimation in preferential favour of enhanced terpenoid accumulation accompanying vegetative to reproductive phase transition.

Artemisia annua accumulates substantial quantities of unique and highly useful antimalarial sesquiternoid artemisinin and related phytomolecules as well as its characteristic essential oil in its glandular trichomes. The phytomolecules are mainly produced in its leaves and inflorescences. Artemisia annua plants were grown under NaCl salinity (50, 100 and 200 mM) stress conditions imposed throughout the entire life cycle of the plant. Results revealed that specialized metabolites like artemisinin, arteannuin-B, artemisinic acid + dihydroartemisinic acid and essential oil accumulation were positively modulated by NaCl salinity stress. Interestingly, total content of monoterpenoids and sesquiterpenoids of essential oil was induced by NaCl salinity treatment, contrary to previous observations. Production of camphor, the major essential oil constituent was induced under the influence of treatment. The metabolic acclimation and manifestations specific to terpenoid pathway are analysed vis-a-vis vegetative to reproductive periods and control of the modulation. WRKY and CYP71AV1 play a key role in mediating the responses through metabolism in glandular trichomes. The distinctness of the salinity induced responses is discussed in light of differential mechanism of adaptation to abiotic stresses and their impact on terpenoid-specific metabolic adjustments in A. annua. Results provide potential indications of possible adaptation of A. annua under saline conditions for agrarian techno-economic benefaction.

Plant secondary metabolism linked glycosyltransferases: An update on expanding knowledge and scopes.

The multigene family of enzymes known as glycosyltransferases or popularly known as GTs catalyze the addition of carbohydrate moiety to a variety of synthetic as well as natural compounds. Glycosylation of plant secondary metabolites is an emerging area of research in drug designing and development. The unsurpassing complexity and diversity among natural products arising due to glycosylation type of alterations including glycodiversification and glycorandomization are emerging as the promising approaches in pharmacological studies. While, some GTs with broad spectrum of substrate specificity are promising candidates for glycoengineering while others with stringent specificity pose limitations in accepting molecules and performing catalysis. With the rising trends in diseases and the efficacy/potential of natural products in their treatment, glycosylation of plant secondary metabolites constitutes a key mechanism in biogeneration of their glycoconjugates possessing medicinal properties. The present review highlights the role of glycosyltransferases in plant secondary metabolism with an overview of their identification strategies, catalytic mechanism and structural studies on plant GTs. Furthermore, the article discusses the biotechnological and biomedical application of GTs ranging from detoxification of xenobiotics and hormone homeostasis to the synthesis of glycoconjugates and crop engineering. The future directions in glycosyltransferase research should focus on the synthesis of bioactive glycoconjugates via metabolic engineering and manipulation of enzyme's active site leading to improved/desirable catalytic properties. The multiple advantages of glycosylation in plant secondary metabolomics highlight the increasing significance of the GTs, and in near future, the enzyme superfamily may serve as promising path for progress in expanding drug targets for pharmacophore discovery and development.