Advances in Biotechnological Production and Metabolic Regulation of Astragalus membranaceus.

Legume medicinal plants Astragalus membranaceus are widely used in the world and have very important economic value, ecological value, medicinal value, and ornamental value. The bioengineering technology of medicinal plants is used in the protection of endangered species, the rapid propagation of important resources, detoxification, and the improvement of degraded germplasm. Using bioengineering technology can effectively increase the content of secondary metabolites in A. membranaceus and improve the probability of solving the problem of medicinal plant resource shortage. In this review, we focused on biotechnological research into A. membranaceus, such as the latest advances in tissue culture, including callus, adventitious roots, hairy roots, suspension cells, etc., the metabolic regulation of chemical compounds in A. membranaceus, and the research progress on the synthetic biology of astragalosides, including the biosynthesis pathway of astragalosides, microbial transformation of astragalosides, and metabolic engineering of astragalosides. The review also looks forward to the new development trend of medicinal plant biotechnology, hoping to provide a broader development prospect for the in-depth study of medicinal plants.

Application of Data Modeling, Instrument Engineering and Nanomaterials in Selected Medid the Scientific Recinal Plant Tissue Culture.

At present, most precious compounds are still obtained by plant cultivation such as ginsenosides, glycyrrhizic acid, and paclitaxel, which cannot be easily obtained by artificial synthesis. Plant tissue culture technology is the most commonly used biotechnology tool, which can be used for a variety of studies such as the production of natural compounds, functional gene research, plant micropropagation, plant breeding, and crop improvement. Tissue culture material is a basic and important part of this issue. The formation of different plant tissues and natural products is affected by growth conditions and endogenous substances. The accumulation of secondary metabolites are affected by plant tissue type, culture method, and environmental stress. Multi-domain technologies are developing rapidly, and they have made outstanding contributions to the application of plant tissue culture. The modes of action have their own characteristics, covering the whole process of plant tissue from the induction, culture, and production of natural secondary metabolites. This paper reviews the induction mechanism of different plant tissues and the application of multi-domain technologies such as artificial intelligence, biosensors, bioreactors, multi-omics monitoring, and nanomaterials in plant tissue culture and the production of secondary metabolites. This will help to improve the tissue culture technology of medicinal plants and increase the availability and the yield of natural metabolites.

Production of anthraquinones from cell and organ cultures of Morinda species.

Since ancient times, Morinda species, particularly Morinda citrifolia, have been used for their therapeutic benefits. Iridoids, anthraquinones, coumarins, flavonoids, lignans, phytosterols, and carotenoids are examples of natural substances with bioactivity. Anthraquinone derivatives are the most significant of these chemicals since they are utilized as natural coloring agents and have a wide range of medicinal functions. Utilizing cell and organ cultures of Morinda species, various biotechnological methods have been developed for the bioproduction of anthraquinone derivatives. The generation of anthraquinone derivatives in cell and organ cultures is summarized in this article. The methods used to produce these chemicals in bioreactor cultures have also been examined. KEY POINTS: • This review investigates the potential of cell and organ cultures for anthraquinone synthesis. • The overproduction of anthraquinones has been addressed using a variety of techniques. • The use of bioreactor technologies for anthraquinone manufacturing is highlighted.

Evaluation of growth and some unexplored bioactivities of bioreactor grown adventitious root culture of ginseng (Panax ginseng C.A. Meyer).

The purpose of the present study was to evaluate the growth potential and some rarely reported bioactivities (antioxidant, thrombolytic, anticoagulant, and anthelmintic) of Panax ginseng C.A. Meyer adventitious roots. To demonstrate the growth, shake flask and laboratory-scale bioreactor cultures have been employed. The obtained biomass was dried and extracted with water, ethanol, and methanol. The growth ratio (12.62 ± 1.03) observed in the bioreactor was significantly higher than in the shake flask culture. The presence of 10 different phytochemical classes, including carbohydrates, saponins, glycosides, and terpenoids were detected in qualitative estimation. Significant quantities of phenolics, flavonoids, proteins, and tannins were determined. Dose-dependent antioxidant activities were observed, and the IC50 values of methanolic and ethanolic extracts were very similar to the standard. The highest (29.26 ± 5.31%) thrombolytic potential was shown by the methanolic extract. The ethanolic extract significantly extended the coagulation times up to 2.5 fold. The highest anthelmintic properties in terms of paralyzing (2.21 ± 0.31 min) and killing (3.69 ± 0.41 min) of the parasitic worms were displayed by the aqueous extract. The in vitro root growth implies the commercial feasibility of ginseng production in Bangladesh and the demonstration of potential bioactivities strengthens medicinal implications and also offering new research areas.

Anthraquinone Production from Cell and Organ Cultures of Rubia Species: An Overview.

The Rubia genus includes major groups of medicinal plants such as Rubia cordifolia, Rubia tinctorum, and Rubia akane. They contain anthraquinones (AQs), particularly alizarin and purpurin, which have pharmacological effects that are anti-inflammatory, antioxidant, anticancer, hemostatic, antibacterial, and more. Alizarin and purpurin have been utilized as natural dyes for cotton, silk, and wool fabrics since the dawn of time. These substances have been used in the cosmetics and food industries to color products. The amount of AQs in different Rubia species is minimal. In order to produce these compounds, researchers have established cell and organ cultures. Investigations have been conducted into numerous chemical and physical parameters that affect the biomass and accumulation of secondary metabolites in a cell, callus, hairy root, and adventitious root suspension cultures. This article offers numerous techniques and approaches used to produce biomass and secondary metabolites from the Rubia species. Additionally, it has been emphasized that cells can be grown in bioreactor cultures to produce AQs.

A comprehensive in silico exploration of pharmacological properties, bioactivities and COX-2 inhibitory potential of eleutheroside B from Eleutherococcus senticosus (Rupr. & Maxim.) Maxim.

Eleutherococcus senticosus (Rupr. & Maxim.) Maxim., popularly known as 'Siberian ginseng', is an important medicinal plant. Pharmacologically active compounds of this plant are called eleutherosides and among them, eleutheroside B is the most prevalent. The E. senticosus has been reported to have many medicinal properties however; very few studies are reported to understand the medicinal properties of eleutheroside B. Consequently, in the present study various computational tools have been used to predict the drug-likeness, bioactivities, and pharmacokinetic properties of eleutheroside B. Besides, the inhibitory potential of eleutheroside B has been investigated against cyclooxygenase 2 (COX-2) enzyme. This study suggests that eleutheroside B is a drug-like compound with bioactivity score (-0.08 to 0.38), having satisfactory pharmacokinetic values. Metabolism and toxicities were further studied using FAME3, GLORY, pred-hERG and Endocrine Disruptome tools. No severe toxicities (Ames, hepatotoxicity, cardiotoxicity, skin sensitization) were predicted. Rat acute toxicity, ecotoxicity and cell line cytotoxicity were evaluated based on GUSAR and CLC-pred. The compound has been predicted as non-toxic (class 5), non-hERG inhibitor and less likely to cause adverse drug interactions. Molecular docking against COX-2 enzyme revealed strong hydrogen bonds (SER530, TYR355, LEU352, SER353, VAL349, TYR385, MET522) and hydrophobic interaction (LEU352) with eleutheroside B. The docking score (-6.97 kcal/mol) suggested that this molecule can be utilized as an anti-inflammatory agent as well as a potential anticancer drug in the future. Hence, this is a comprehensive integrated in silico approach to establish the anti-inflammatory mechanism of eleutheroside B in the background of its potential in future drug development.Communicated by Ramaswamy H. Sarma.

Attributes of Polygonum multiflorum to transfigure red biotechnology.

A vast array of plant-based compounds has enriched red biotechnology to serve the human health and food. A peculiar medicinal plant which was an element of traditional Chinese medicine for centuries as a liver and kidney tonic, for life longevity and hair blackening, is Polygonum multiflorum Thunb. (PM) which is popularly known as "He shou wu" or "Fo-ti" and is rich in chemical components like stilbenes, quinones, and flavonoids which have been used as anti-aging, anti-alopecia, anti-cancer, anti-oxidative, anti-bacterial, anti-hyperlipidemia, anti-atherosclerosis, and immunomodulating and hepatoprotective agents in the modern medicine. The health benefits from PM are attained since long through commercial products such as PM root powder, extract, capsules, tincture, shampoo, and body sprays in the market. Currently, the production of these pharmaceuticals and functional foods possessing stilbenes, quinones, and flavonoids is through cell and organ cultures to meet the commercial demand. However, hepatotoxic effects of PM-based products are the stumbling blocks for its long-term usage. The current review encompasses a comprehensive account of bioactive compounds of PM roots, their biological activities as well as efficacy and toxicity issues of PM ingredients and future perspectives.

Quality, safety and efficacy profiling of ginseng adventitious roots produced in vitro.

Ginseng (Panax ginseng C. A. Meyer, Family Araliaceae) is one of the major medicinal and nutraceutical plants, which is native to oriental region. It is used worldwide as a popular herbal medicine because of its pharmacological effects like anti-oxidative, anti aging, anti-cancer, adaptogenic, and other health-improving activities. Chief components of ginseng identified till date are ginsenosides, a group of saponins with triterpenoid structure. Ginseng is cultivated under controlled conditions, and for harvesting of fully grown roots of the plant, the cultivation takes long duration of about 5-7 years and cultivated ginseng roots are inferior in quality and ginsenoside content. Wild Mountain ginseng is superior in quality and ginsenoside content but is scarce in nature. Therefore, for obtaining the useful compounds of this plant at commercial scale, cell and organ cultures especially adventitious roots have been established by using superior clones of wild mountain ginseng, ginseng biomass is produced by applying large scale bioreactors. In this paper, an effort has been made to shed light on the scientific literature and to decipher the evidences for quality, safety, and efficacy of ginseng adventitious roots produced from in vitro cultures.

Biotic elicitation of ginsenoside metabolism of mutant adventitious root culture in Panax ginseng.

Biotic elicitation is an important biotechnological strategy for triggering the accumulation of secondary metabolites in adventitious root cultures. These biotic elicitors can be obtained from safe, economically important strains of bacteria found in the rhizosphere and fermented foods. Here, we assayed the effects of filtered cultures of five nitrogen-fixing bacteria and four types of fermentation bacteria on mutant adventitious Panax ginseng root cultures induced in a previous study by colchicine treatment. The biomass, pH, and electrical conductivity (EC) of the culture medium were altered at 5 days after treatment with bacteria. The saponin content was highest in root cultures treated with Mesorhizobium amorphae (GS3037), with a concentration of 105.58 mg g-1 dry weight saponin present in these cultures versus 74.48 mg g-1 dry weight in untreated root cultures. The accumulation of the ginsenosides Rb2 and Rb3 dramatically increased (19.4- and 4.4-fold, and 18.8- and 4.8-fold) 5 days after treatment with M. amorphae (GS3037) and Mesorhizobium amorphae (GS336), respectively. Compound K production increased 1.7-fold after treatment with M. amorphae (GS3037) compared with untreated root cultures. These results suggest that treating mutant adventitious root cultures with biotic elicitors represents an effective strategy for increasing ginsenoside production in Panax ginseng.

Panax ginseng Adventitious Root Suspension Culture: Protocol for Biomass Production and Analysis of Ginsenosides by High Pressure Liquid Chromatography.

Panax ginseng C.A. Meyer (Korean ginseng) is a popular herbal medicine. It has been used in Chinese and Oriental medicines since thousands of years. Ginseng products are generally used as a tonic and an adaptogen to resist the adverse influence of a wide range of physical, chemical and biological factors, and to restore homeostasis. Ginsenosides or ginseng saponins are the principal active ingredients of ginseng. Since ginseng cultivation process is very slow and needs specific environment for field cultivation, cell and tissue cultures are sought as alternatives for the production of ginseng biomass and bioactive compounds. In this chapter, we focus on methods of induction of adventitious roots from ginseng roots, establishment of adventitious root suspension cultures using bioreactors, procedures for processing of adventitious roots, and analysis of ginsenosides by high pressure liquid chromatography.

Anti-inflammatory potential of saponins derived from cultured wild ginseng roots in lipopolysaccharide-stimulated RAW 264.7 macrophages.

Ginseng, namely the root of Panax ginseng Meyer, is a well-known traditional medicine that has been used in Asian countries for thousands of years. Ginseng saponins have been shown to exert a variety of prominent pharmacological effects in a number of diseases. The aim of the present study was to identify the anti-inflammatory effects of total saponins extracted from cultured wild ginseng roots (TSWG) on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. An elevated production of nitric oxide (NO) was detected in the RAW 264.7 cells in response to stimulation with LPS, as shown by NO detection assay using Griess reagent. However, pre-treatment with TSWG inhibited the production of NO through the suppression of inducible NO synthase gene expression. Furthermore, the LPS-induced gene expression and production of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) were significantly reduced by treatment with TSWG, as shown by ELISA, and western blot analysis and RT-PCR, respectively. In the LPS-stimulated RAW 264.7 cells, nuclear factor-κB (NF-κB) was translocated from the cytosol to the nucleus, while pre-treatment with TSWG induced the sequestration of NF-κB in the cytosol through the inhibition of the inhibitor of κB degradation, as shown by immunofluorescence staining. TSWG also contributed to the downregulation of mitogen-activated protein kinases and Akt in the LPS-stimulated RAW 264.7 cells. Additionally, in the TSWG-treated RAW 264.7 cells, we observed the activation of nuclear factor (erythroid-derived 2)-like 2 and an increase in heme oxygenase-1 expression; these effects were associated with the inhibition of the generation of reactive oxygen species. The results from the present study indicate that TSWG exerts anti-inflammatory and antioxidant effects, suggesting that TSWG may be an effective therapeutic agent for inflammatory diseases and prevent cellular damage induced by oxidative stress.

Hypericins: biotechnological production from cell and organ cultures.

Hypericum perforatum L. (St. John's wort), a perennial flowering plant native to Europe, is widely used as a medicinal plant and has a long history of its use in the treatment of various ailments. Currently, H. perforatum is widely used as an herbal remedy for the treatment of mild to moderate depression. Hypericins are natural napthodianthrone compounds produced from H. perforatum (St. John's wort) which are having antitumor, antiviral (i.e., against human immunodeficiency and hepatitis C virus), antineoplastic, and antidepressant properties. Currently, field-grown plant materials are generally used for the commercial production of hypericins. It has been reported that hypericin accumulation in natural plants is influenced by different ecological and environmental conditions including light intensity, nitrogen availability, temperature, seasons, and growing regions. Therefore, up to 17-fold and 13-fold differences in hypericin and pseudohypericin amounts, respectively, are reported in different phytopharmaceutical preparations. Plant cell and organ cultures are effective systems for producing natural products, and attempts were made for the production of biomass and stable concentrations of hypericins through in vitro cultures of H. perforatum. Cell, callus, shoot, plantlet, and adventitious root cultures have been established and various chemical and physical factors which influence the biomass and secondary metabolite accumulation have been investigated. Large-scale plantlet and adventitious root cultures have also been attempted in H. perforatum in bioreactors, and various strategies have been applied for the production of higher biomass and secondary products. This review describes the biotechnological approaches employed for the production of hypericins and focuses upon the challenges and future prospects.

Biotechnological production of caffeic acid derivatives from cell and organ cultures of Echinacea species.

Caffeic acid derivatives (CADs) are a group of bioactive compounds which are produced in Echinacea species especially Echinacea purpurea, Echinacea angustifolia, and Echinacea pallida. Echinacea is a popular herbal medicine used in the treatment of common cold and it is also a prominent dietary supplement used throughout the world. Caffeic acid, chlorogenic acid (5-O-caffeoylquinic acid), caftaric acid (2-O-caffeoyltartaric acid), cichoric acid (2, 3-O-dicaffeoyltartaric acid), cynarin, and echinacoside are some of the important CADs which have varied pharmacological activities. The concentrations of these bioactive compounds are species specific and also they vary considerably with the cultivated Echinacea species due to geographical location, stage of development, time of harvest, and growth conditions. Due to these reasons, plant cell and organ cultures have become attractive alternative for the production of biomass and caffeic acid derivatives. Adventitious and hairy roots have been induced in E. pupurea and E. angustifolia, and suspension cultures have been established from flask to bioreactor scale for the production of biomass and CADs. Tremendous progress has been made in this area; various bioprocess methods and strategies have been developed for constant high-quality productivity of biomass and secondary products. This review is aimed to discuss biotechnological methods and approaches employed for the sustainable production of CADs.

Pilot-scale culture of Hypericum perforatum L. adventitious roots in airlift bioreactors for the production of bioactive compounds.

Hypericum perforatum L. (St. John's Wort) is an important medicinal plant which is widely used in the treatment for depression and irritable bowel syndrome. It is also used as a dietary supplement. Major bioactive phytochemicals of H. perforatum are phenolics and flavonoids. Quality of these phytochemicals is dramatically influenced by environmental and biological factors in the field grown plants. As an alternative, we have developed adventitious root cultures in large-scale bioreactors for the production of useful phytochemicals. Adventitious roots of H. perforatum were cultured in 500 l pilot-scale airlift bioreactors using half-strength Murashige and Skoog medium with an ammonium and nitrate ratio of 5:25 mM and supplemented with 1.0 mg l(-1) indole butyric acid, 0.1 mg l(-1) kinetin, and 3 % sucrose for the production of bioactive phenolics and flavonoids. Then 4.6 and 6.3 kg dry biomass were realized in the 500 l each of drum-type and balloon-type bioreactors, respectively. Accumulation of 66.9 mg g(-1) DW of total phenolics, 48.6 mg g(-1) DW of total flavonoids, 1.3 mg g(-1) DW of chlorogenic acid, 0.01 mg g(-1) DW of hyperin, 0.04 mg g(-1) DW of hypericin, and 0.01 mg g(-1) DW of quercetin could be achieved with adventitious roots cultured in 500 l balloon-type airlift bioreactors. Our findings demonstrate the possibilities of using H. perforatum adventitious root cultures for the production of useful phytochemicals to meet the demand of pharmaceutical and food industry.

Biotechnological production of eleutherosides: current state and perspectives.

Eleutherosides, the phenylpropanoid and lignan glycosides, are the active ingredients accumulated in the roots and stems of Eleutherococcus species and in Eleutherococcus senticosus in particular. Syringin (=eleutheroside B) and (-) syringaresinol-di-O-ß-D-glucoside (=eleutheroside E) appear as the most important bioactive compounds which are used as adaptogens, besides their abundant antidiabetic and anticancer properties. As the availability of "Eleuthero" is becoming increasingly limited because of its scanty natural distribution, the production of these compounds by biotechnological means has become an attractive alternative. In E. senticosus and other closely related species, Eleutherococcus sessiliflorus, Eleutherococcus chiisanensis, and Eleutherococcus koreanum, organogenic cultures have been induced for the production of eleutherosides. Bioreactor cultures have been established and various parameters, which influence on the accumulation of biomass and secondary metabolites, have been thoroughly investigated. Pilot-scale cultures have also been accomplished for the large-scale production of somatic embryos containing abundant amounts of eleutherosides. This review describes the biotechnological approaches and challenges for the production of eleutherosides.

Ginsenosides: prospective for sustainable biotechnological production.

Panax ginseng C.A. Meyer (ginseng) is a well-known medicinal plant that has been traditionally used in the oriental countries for centuries. Wild ginseng is a scarce and rare commodity. Field cultivation of the ginseng plant is a time-consuming and labor-intensive process. Ginsenosides, a group of glycosylated triterpenes, also known as saponins, are the principal bioactive constituents of ginseng. The use of cell and organ culture processes has been sought as a potential alternative for the efficient mass production of ginseng raw material. Various bioprocessing strategies have been developed to date. Cells and adventitious roots have been cultured in large-scale bioreactors and various strategies have been developed accordingly for the enhancement of biomass and ginsenoside accumulation. This review highlights the recent progress in the cultivation of ginseng cell and organ cultures for the production of ginsenosides from bioreactor cultures. In addition, the metabolism and biochemistry of ginsenoside biosynthesis, genomic and proteomic studies in ginseng, metabolic engineering, biosafety, toxicological evaluation, and efficacy assessment of ginseng raw material are also summarized and thoroughly discussed.

Pilot-scale culture of somatic embryos of Eleutherococcus senticosus in airlift bioreactors for the production of eleutherosides.

PURPOSE OF WORK: To establish pilot scale bioreactor cultures of somatic embryos of Siberian ginseng for the production of biomass and eleutherosides. Somatic embryos of Eleutherococcus senticosus were cultured in airlift bioreactors using Murashige and Skoog medium with 30 g sucrose l(-1) for the production of biomass and eleutherosides. Various parameters including the type of bioreactor, aeration volume, and inoculum density were optimized for 3 l capacity bioreactors. Balloon-type airlift bioreactors, utilizing a variable aeration volume of 0.1-0.3 vvm and an inoculum of 5 g l(-1), were suitable for biomass and eleutheroside production. In 500 l balloon-type airlift bioreactors, 11.3 g dry biomass l(-1), 220 µg eleutheroside B l(-1), 413 µg eleutheroside E l(-1), and 262 µg eleutheroside E1 l(-1) were produced.

Efficacy of ginseng adventitious root extract on hyperglycemia in streptozotocin-induced diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginseng has various bioactive effects on human health including its potential activity of improving the glucose homeostasis and insulin sensitivity. MATERIALS AND METHODS: Tissue culture raised mountain ginseng adventitious root (TCMGARs) extract enriched with ginsenosides was used as experimental material. Streptozotocin-induced diabetic 'Sprague Dawley' male rats were used as experimental systems and were fed with Tissue culture raised mountain ginseng adventitious root extract. Field cultivated Korean ginseng root extract fed rats were used as positive control and several indices such as body weight, blood glucose level and other serological indicators were tested. RESULTS: Chemical profile showed TCMGARs were rich in varied ginsenosides especially Rb1, Rb2, Rc, Rd, Rg3, and Rh2 when compared to field cultivated Korean ginseng. TCMGARs extract at dosage levels of 250 and 500 mg/kg body weight significantly lowered the blood glucose, total cholesterol and triglyceride content in streptozotocin-induced diabetic rats. CONCLUSION: The data of in vivo experiments on anti-glycemic effects of TCMGARs proves their efficacy and also their use as dietary supplement.

A strategy for enrichment of the bioactive sphingoid base-1-phosphates produced by Hypericum perforatum L. in a balloon type airlift reactor.

An efficient enrichment method using immobilized metal affinity chromatography (IMAC) was developed for selective extraction of bioactive sphingoid base-1-phosphates (SB1Ps) from adventitious roots of Hypericum perforatum cultured in bioreactor. The phosphate-selective IMAC enrichment coupled with LC-MS/MS enabled sensitive analysis of low-abundance SB1Ps present in the root biomass, which would not be feasible otherwise due to severe interferences from complex biological matrices. The time-dependent growth rate and production of SB1Ps from adventitious roots were investigated. The level of phytosphingosine-1-phosphate, which was found to be the major SB1Ps, reached a maximum amount of 635.6pmolpergram of dry weight after 3weeks of culture and decreased between 3 and 5weeks of culture subsequently. On the other hand, sphingosine-1-phosphate and sphinganine-1-phosphate were present at levels of 18.91 and 73.15pmolpergram of dry weight, respectively, after a week of culture and their level decreased thereafter.

Production of biomass and useful compounds from adventitious roots of high-value added medicinal plants using bioreactor.

The increasing global demand for biomass of medicinal plant resources reflects the issues and crisis created by diminishing renewable resources and increasing consumer populations. Moreover, diverse usage of plants and reduced land for cultivation in the world accelerated the deficiency of plant resources. In addition, the preparation of safety of plant based medicine whips up demand for biomass of valuable medicinal plants. As one of alternative approach to upswing the productivity of plant-based pharmaceutical compounds, automation of adventitious root culture system in air-lift bioreactor was adopted to produce cosmic amount of root biomass along with enriched diverse bioactive molecules. In this review, various physiological, engineering parameters, and selection of proper cultivation strategy (fed-batch, two-stage etc.) affecting the biomass production and secondary metabolite accumulation have been discussed. In addition, advances in adventitious root cultures including factors for process scale-up as well as recent research aimed at maximizing automation of the bioreactor production processes are also highlighted. Examples of the scale-up of cultures of adventitious roots of Morinda citrifolia, Echinacea purpurea and angustifolia, Hypericum perforatum and Panax ginseng by applying 20 L to 10,000 L bioreactors in our lab were demonstrated with a view of commercial application.